EP1654268A2 - Lincomycin derivatives possessing antibacterial activity - Google Patents

Lincomycin derivatives possessing antibacterial activity

Info

Publication number
EP1654268A2
EP1654268A2 EP04785949A EP04785949A EP1654268A2 EP 1654268 A2 EP1654268 A2 EP 1654268A2 EP 04785949 A EP04785949 A EP 04785949A EP 04785949 A EP04785949 A EP 04785949A EP 1654268 A2 EP1654268 A2 EP 1654268A2
Authority
EP
European Patent Office
Prior art keywords
propyl
methyl
pyran
methylsulfanyl
tetrahydro
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP04785949A
Other languages
German (de)
French (fr)
Inventor
Jason G. Lewis
Dinesh V. Patel
Sampath K. Anandan
Mikhail F. Gordeev
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Vicuron Pharmaceuticals LLC
Original Assignee
Vicuron Pharmaceuticals LLC
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US10/642,807 external-priority patent/US7164011B2/en
Priority claimed from US10/777,455 external-priority patent/US7199105B2/en
Application filed by Vicuron Pharmaceuticals LLC filed Critical Vicuron Pharmaceuticals LLC
Publication of EP1654268A2 publication Critical patent/EP1654268A2/en
Withdrawn legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
    • C07H15/02—Acyclic radicals, not substituted by cyclic structures
    • C07H15/14—Acyclic radicals, not substituted by cyclic structures attached to a sulfur, selenium or tellurium atom of a saccharide radical
    • C07H15/16—Lincomycin; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • This invention relates to lincomycin derivatives that exhibit antibacterial activity.
  • Lincomycin is a biosynthetic product that adversely affects growth of various microorganisms, in particular gram positive bacteria.
  • the characteristics and preparation of lincomycin are disclosed in U.S. Patent 3,086,912.
  • a variety of derivatives of lincomycin, which also have antimicrobial activity, have been prepared. These derivatives include, for example, clindamycin, which is described in U.S. Patent 3,496,163.
  • Lincomycin derivatives remain attractive targets for antibacterial drug discovery. Accordingly, lincomycin derivatives that possess antimicrobial activity are desired as potential antibacterial agents.
  • the present invention provides lincomycin derivatives that possess antibacterial activity.
  • m is 0. In another embodiment, m is 1. In another embodiment, m is 2. In another embodiment, m is 3.
  • R 1 is methyl
  • one of R 2 and R 3 is H, and the other is alkyl. In one embodiment, one of R 2 and R 3 is H, and the other is methyl. In one embodiment, one of R 2 and R 3
  • each R 9 is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, cycloalkylalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, - (CH ) n -OH, -(CH ) n -NR 4 R 5 , -alkylene-R a where R a is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R 4 and R 5 are H or alkyl.
  • R 9 is a substituted alkyl, and wherein the one or more substituents on the alkyl group are selected from the group consisting of halogen, oxygen, hydroxy, amine (primary), amine (secondary-amine substituted by alkyl above), amine (tertiary- amine substituted by alkyl as above), sulfur, -SH, and phenyl.
  • each R 9 is independently selected from the group consisting of halogen, alkyl, substituted alkyl, alkoxy, alkoxyalkoxy, substituted alkoxy, and cycloalkyl.
  • each R 9 is independently selected from the group consisting of H, alkyl, and substituted alkyl.
  • R 9 is selected from the group consisting of 1-ethylpropyl; 2-(4- fluorophenyl)ethyl; 2,2-difluoroethoxymethyl/fluoro; 2,2-difluoropentyl; 2-cyclobutyl-ethyl; 2- cyclopropyl-ethyl; 2-fluoroethoxy; 2-methoxyethoxy; 2-propoxy-ethyl; 3-(2-fluoro-ethoxy)- propyl; 3-(3-fluoro-propoxy)-propyl; 3-(4-chlorophenyl)propyl; 3-(cyclopropyl-methoxy)- propyl; 3,3,3-trifluoropropoxy; 3,3-difluorobutyl; 3,3-difluoropentyl; 3,3-difluoropropyl; 3,3- difluoropropyl/fluoro; 3-cyclobutyl-propy
  • R 9 is selected from the group consisting of 1-ethylpropyl; 2-(4- fluorophenyl)ethyl; 2,2-difluoropentyl; 2-cyclobutyl-ethyl; 2-cyclopropyl-ethyl; 2- methoxyethoxy; 3-(2-fluoro-ethoxy)-propyl; 3-(4-chlorophenyl)propyl; 3,3-difluorobutyl; 3,3- difluoropentyl; 3,3-difluoropropyl; 3-cyclobutyl-propyl; 3-cyclopentyl-propyl; 3-cyclopropyl- propyl; 3-ethoxy-propyl; 3-fluoropropyl; 3-hydroxy-3-ethylpentyl; 3-methoxy-propyl; 3- propoxy-propyl; 4,4-difluorobutyl; 4-fluorobutyl; 5,5-
  • R 9 not 2,2-difluoroethoxymethyl/fluoro; 2-fluoroethoxy; 2- propoxy-ethyl; 3-(3-fluoro-propoxy)-propyl; 3-(cyclopropyl-methoxy)-propyl; 3,3,3- trifluoropropoxy; 3,3-difluoropropyl/fluoro; 3-fluoropropoxy; 3-fluoro-propoxy-methyl; 3- fluoropropyl/fluoro; 4,4-difluoropentyl; 4-fluorobutoxy; 4-methoxy-butyl; ethyl/fluoro; fluoro/butyl; or fluoro/propyl.
  • this invention is directed to a prodrug compound of the formula (IN):
  • R 1 , R 2 , R 3 , and R 9 are as defined for formula I; wherein R 6 is as defined for formula I or is a suitable prodrug group which is cleavable in vivo; and wherein R 11 is H or a suitable prodrug group which is cleavable in vivo.
  • the compound is a prodrug wherein R 6 is selected from the group consisting of hydrogen; l-(acetyloxy)-ethyl-oxycarbonyl; l-amino-2-methyl-butyl- carbonyl; 1 -amino-2-methyl-butyl-carbonyl-oxy-methyl-oxycarbonyl; 1 -amino-2-methyl- propyl-carbonyl; l-amino-2-phenyl-ethyl-carbonyl; 1-amino-ethyl-carbonyl; 1-methyl-l, 2,3,6 tetrahydro-pyridin-4-yl-oxycarbonyl; 1-methyl-l, 4 dihydro-pyridin-3-yl-carbonyl; l-methyl-3- oxo-but-1-enyl; 5-methyl-[l,3]dioxol-2-one-4-yl-methoxy-carbonyl; 5-methyl-[l,3]
  • the compound is a prodrug wherein R 6 is selected from the group consisting of hydrogen; l-(acetyloxy)-ethyl-oxycarbonyl; l-amino-2-methyl-butyl- carbonyl; 1 -amino-2 -methyl-butyl-carbonyl-oxy-methyl-oxycarbonyl; 1 -amino-2-methyl- propyl-carbonyl; l-amino-2-phenyl-ethyl-carbonyl; 1-amino-ethyl-carbonyl; 1-methyl-l, 2,3, 6 tetrahydro-pyridin-4-yl-oxycarbonyl; l-methyl-1,4 dil ⁇ ydro-pyridin-3-yl-carbonyl; l-methyl-3- oxo-but- 1 -enyl; ethyl-carbonylamino-metliyl; piperidin-4-yl-carbonyl
  • the compound is a prodrug wherem R is not 5-methyl- [1 ,3]dioxol-2-one-4-yl-methoxy-carbonyl; 5-methyl-[l ,3]dioxol-2-one-4-yl-methyl; ethoxy- carbonyl; fluorenyl-methylene-oxy-carbonyl; or phenoxy-carbonyl.
  • the compound is a prodrug wherein R 6 is not 5-methyl- [1 ,3]dioxol-2-one-4-yl-methoxy-carbonyl; or 5-methyl-[l ,3]dioxol-2-one-4-yl-methyl.
  • the compound is a prodrug wherein R 11 is selected from the group consisting of hydrogen; 2-(N-(2-morpholin-4-yl-ethyl)-amino-carbonyl)-ethyl- carbonyl; -C(O)CH 2 CH 2 COOH; N,N-dimethyl-amino-methyl-carbonyl; pentadecyl- ' carbonyloxy; and -PO 3 H 2 .
  • this invention provides compounds of formula (II)
  • R 1 is alkyl
  • R 2 and R 3 are independently H, alkyl, or cyanoalkyl, with the proviso that both R 2 and R 3 are not H
  • R 6 is H, alkyl, or hydroxyalkyl
  • R 9 which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH ) n -OH, - (CH 2 ) n - ⁇ R 4 R 5 , -alkylene-R a where R a is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R 4 and R 5 are H or alkyl; and
  • R 1 is alkyl; R 2 and R 3 are fluoro; R is H, alkyl, or hydroxyalkyl; R 9 , which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH 2 ) n -OH, - (CH 2 ) n -NR 4 R 5 , -alkylene-R a where R a is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R 4 and R 5 are H or alkyl; and m is 1 or 2; and prodrugs and pharmaceutically acceptable salts thereof, with tl e proviso that
  • R 7 is H or alkyl
  • R 9 which can be singly or multiply substituted in tlie ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH 2 ) n -OH, - (CH ) n -NR 4 R 5 , -alkylene-R a where R a is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R 4 and R 5 are H or alkyl; and m is 1 or 2; and prodrugs, tautomers or pharmaceutically acceptable salts thereof; with the proviso that the compound of formula I has a minimum inhibition concentration of 32 ⁇ g/mL or less against at least
  • R and/or S isomers as either individual isomers or as a mixture
  • R and/or S isomers as either individual isomers or as a mixture
  • Specific compounds within the scope of this invention include the following compounds: 4-Ethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 1 -Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [3-cyano-2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Ethyl-piperidine-2 -carboxylic acid [2-hydroxy-2-methyl-l -(3,4,5-trihydroxy-6- methylsulf
  • pro-drug compounds within the scope of this invention include the following compounds: 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid 9H-fluoren-9-ylmethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid ethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid phenyl ester; Phosphoric acid mono-(4,5-
  • [0027] is a compound selected from the group consisting of: 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclobutylmethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy
  • compositions comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound defined herein.
  • the pharmaceutical compositions of the present invention may further comprise one or more additional antibacterial agents.
  • One or more of the antibacterial agents may be active against gram negative bacteria.
  • One or more of the antibacterial agents may be active against gram positive bacteria.
  • this invention is directed to a method for tlie treatment of a microbial infection in a mammal comprising administering to the mammal a therapeutically effective amount of a compound of this invention.
  • the compound of this invention may be administered to the mammal orally, parenterally, transdermally, topically, rectally, or intranasaUy.
  • this invention is directed to a method for the treatment of a microbial infection in a mammal comprising administering to the mammal a pharmaceutical composition comprising a therapeutically effective amount of a compound of this invention.
  • the pharmaceutical compositions of the present invention may further comprise one or more additional antibacterial agents, one of which may be active against gram negative bacteria and/or one of which may be active against gram positive bacteria.
  • the pharmaceutical composition may be administered to the mammal orally, parenterally, transdermally, topically, rectally, or intranasaUy.
  • the microbial infection being treated is a gram positive bacterial infection.
  • the infection may be a gram negative bacterial infection.
  • the infection may be a mycobacteria infection, a mycoplasma infection, or a chlamydia infection.
  • the present invention provides novel intermediates and processes for preparing' compounds of formula (I), (II), (III) and (IN).
  • this invention relates to lincomycin derivatives that exhibit antibacterial activity, in particular gram positive antibacterial activity.
  • antibacterial activity in particular gram positive antibacterial activity.
  • Acyl means the group -C(O)R' wherein R' is alkyl, alkenyl, alkynyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
  • Acyloxy means the group -C(O)OR', wherein R' is alkyl, alkenyl, alkynyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
  • alkenyl groups include, but are not limited to, allyl, vinyl, 2-butenyl, and the like.
  • Alkoxy refers to the group “alkyl-O-" which includes, by way of example, methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n- hexoxy, 1,2-dimethylbutoxy, and the like.
  • Alkoxyalkoxy refers to the group alkyl-O-alkylene-O-, wherein alkyl is as defined herein.
  • Alkyl means a linear saturated monovalent hydrocarbon radical of one to eight carbon atoms or a branched saturated monovalent hydrocarbon radical of three to eight carbon atoms.
  • alkyl groups include, but are not limited to, groups such as methyl, ethyl, n- propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, and the like.
  • Alkylene means a linear divalent hydrocarbon radical of one to eight carbon atoms or a branched divalent hydrocarbon group of three to eight carbon atoms. Examples of alkylene groups include, but are not limited to, methylene, ethylene, 2-methylpropylene, and the like.
  • Alkylthio refers to the group “alkyl-S-” which includes, by way of example, methylthio, butylthio, and the like.
  • Alkynyl means a linear monovalent hydrocarbon radical of two to eight carbon atoms or a branched monovalent hydrocarbon radical of three to eight carbon atoms containing at least one triple bond, (-C ⁇ C-).
  • alkynyl groups include, but are not limited to, ethynyl, propynyl, 2-butynyl, and the like.
  • Amino or “substituted nitrogen” refers to the group "-NR a R b " wherein R a and Rb are independently H, alkyl, haloalkyl, alkenyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
  • Aminocarboxyalkyl means a group “-RcC(O)NR a R b " where Rc is an alkylene, as defined above, and R a and R b are as defined above.
  • Aryl means a monovalent monocyclic or bicyclic aromatic carbocyclic group of six to fourteen ring atoms. Examples include, but are not limited to, phenyl, naphthyl, and anthryl.
  • the aryl ring may be optionally fused to a 5-, 6-, or 7-membered monocyclic non-aromatic ring optionally containing 1 or 2 heteroatoms independently selected from oxygen, nitrogen, or sulfur, the remaining ring atoms being C where one or two C atoms are optionally replaced by a carbonyl.
  • aryl groups with fused rings include, but are not limited to, 2,5- dihydro-benzo[b]oxepinyl, 2,3-dihydrobenzo[l,4]dioxanyl, chromanyl, isochromanyl, 2,3- dihydrobenzofuranyl, 1,3-dihydroisobenzofuranyl, benzo[l,3]dioxolyl, 1,2,3,4- tetrahydroisoquinolinyl, 1,2,3,4-tetrahydroquinolinyl, 2,3-dihydro-lH-indolyl, 2,3dihydro-lH- isoindolyl, benzimidazole-2-onyl, 2-H-benzoxazol-2-onyl, and the like.
  • Carbonyl means the group "C(O).
  • Carboxy means the group “C(O)O.”
  • Cyanoalkyl refers to an alkyl, wherein alkyl is as defined above, substituted with one or more cyano (-CN) groups provided that if two cyano groups are present they are not both on the same carbon atom.
  • cyanoalkyl groups include, for example, cyanomethyl,
  • Cycloalkyl refers to cyclic alkyl groups of from 3 to 20 carbon atoms having a single or multiple cyclic rings including, by way of example, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, adamantanyl, and the like. Cycloalkyl groups of the present invention also include fused multicyclic rings wherein one or more of the rings within the multicyclic ring system are aromatic, as long as the point of attachment to the core or backbone of the structure is on the non-aromatic ring, e.g., fluorenyl.
  • Cycloalkylalkyl means a group -R c R-j where Rc, is an alkylene group and Rd is a cycloalkyl group, as defined above. Examples include, but are not limited to, cyclopropylmethylene, cyclohexylethylene, and the like.
  • Halo or "Halogen” means fluoro, chloro, bromo, or iodo.
  • Haloalkyl means an alkyl, wherein alkyl is as defined above, substituted with one or more, preferably one to 6, of the same or different halo atoms.
  • haloalkyl groups include, for example, trifluoromethyl, 3-fluoropropyl, 2,2-dichloroethyl, and the like.
  • Heteroaryl means a monovalent monocyclic or bicyclic aromatic radical of 5 to 10 ring atoms containing one, two, or three ring heteroatoms selected from N, O, or S, the remaining ring atoms being C.
  • Representative examples include, but are not limited to, thienyl, benzothienyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinolinyl, quinoxalinyl, imidazolyl, furanyl, benzofuranyl, thiazolyl, isoxazolyl, benzisoxazolyl, benzimidazolyl, triazolyl, pyrazolyl, pyrrolyl, indolyl, 2-pyridonyl, 4-pyridonyl, N-alkyl-2-pyridonyl, pyrazinonyl, pyridazinonyl, pyrimidinonyl, oxazolonyl, and the like.
  • Heterocycle or “heterocyclic” refers to a saturated or unsaturated group having a single ring or multiple condensed rings, from 1 to 10 carbon atoms and from 1 to 4 heteroatoms selected from the group consisting of nitrogen, sulfur, or oxygen within the ring, wherein, in fused ring systems one or more of the rings can be aryl or heteroaryl as defined herein.
  • heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine, imidazolidine, imidazoline, piperidine, piperazine, indoline, phthalimide, 1,2,3,4- tetrahydro-isoquinoline, 4,5
  • Heterocycles may be optionally substituted with from one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxyl, carboxy, cyano, oxo, nitro, and alkylthio as these terms are defined herein.
  • Hydroxyalkyl refers to an alkyl, wherein alkyl is as defined above substituted with one or more -OH groups provided that if two hydroxy groups are present they are not both on the same carbon atom.
  • Examples of hydroxyalkyl groups include, for example, hydroxymethyl,
  • “Mammal” refers to all mammals including humans, livestock, and companion animals. - ⁇
  • aryl group optionally mono- or di- substituted with an alkyl group means that the alkyl may but need not be present, and the description includes situations where the aryl group is mono- or disubstituted with an alkyl group and situations where the aryl group is not substituted with the alkyl group.
  • “Pharmaceutically acceptable carrier” means a carrier that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic and neither biologically nor otherwise undesirable, and includes a carrier that is acceptable for veterinary use as well as human pharmaceutical use.
  • “A pharmaceutically acceptable carrier” as used in the specification and claims includes both one and more than one such carrier.
  • “Pharmaceutically acceptable salt” of a compound means a salt that is pharmaceutically acceptable and that possesses the desired pharmacological activity of the parent compound.
  • Such salts include, but are not limited to, (1) acid addition salts, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; or formed with organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2- hydroxyethanesulfonic acid,
  • Prodrugs mean any compound which releases an active parent drug according to a compound of the subject invention in.vivo when such prodrug is administered to a mammalian subject.
  • Prodrugs of a compound of the subject invention are prepared by modifying functional groups present in a compound of the subject invention in such a way that the modifications may be cleaved in vivo to release the parent compound.
  • Prodrugs include compounds of the subject invention wherein a hydroxy, sulfhydryl or amino group in the compound is bonded to any group that may be cleaved in vivo to regenerate the free hydroxyl, amino, or sulfhydryl group, respectively.
  • prodrugs include, but are not limited to esters (e.g., acetate, formate, and benzoate derivatives), carbamates (e.g., N,N-dimethylaminocarbonyl) of hydroxy functional groups in compounds of the subject invention, and the like.
  • esters e.g., acetate, formate, and benzoate derivatives
  • carbamates e.g., N,N-dimethylaminocarbonyl
  • -C(O)O-alkylene-cycloalkyl -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O] p -alkylene-heterocycle, -[C(O)O] p -alkylene-substituted heterocycle, wherein p is 0 or 1.
  • the -C(O)O-substituted alkyl does not include the following:
  • Substituted alkyl means an alkyl group, as defined above, in which one or more of the hydrogen atoms has been replaced by a halogen (i.e., CI, Br, F, or I), oxygen, hydroxy, amine (primary), amine (secondary-amine substituted by alkyl above), amine (tertiary-amine substituted by alkyl as above), sulfur, -SH, phenyl, substituted phenyl, cycloalkyl, alkoxy, substituted alkoxy.
  • a halogen i.e., CI, Br, F, or I
  • oxygen hydroxy
  • amine primary
  • amine secondary-amine substituted by alkyl above
  • amine tertiary-amine substituted by alkyl as above
  • sulfur -SH
  • phenyl substituted phenyl
  • cycloalkyl alkoxy, substituted alkoxy.
  • substituted alkyl groups include, but are not limited to, 1- fluoroethyl, 1-chloroethyl, 2-fluoroethyl, 2-chloroethyl, 1-bromopropyl, 2-iodopropyl, 1- chlorobutyl, 4-flurobutyl, 4-chlorobutyl, 2-cyclopropyl-ethyl, 3-cyclobutyl-propyl, 4- cyclopentyl-butyl, and 4-cyclohexyl-butyl.
  • Substituted alkoxy means substituted alkyl-O-, wherein substituted alkyl is as defined herein.
  • Substituted aryl means an aryl ring substituted with one or more substituents, preferably one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxy, carboxy, cyano, nitro, alkylthio, and thioalkyl.
  • the aryl ring may be optionally fused to a 5-, 6-, or 7-membered monocyclic non-aromatic ring optionally containing 1 or 2 heteroatoms independently selected from oxygen, nitrogen, or sulfur, the remaining ring atoms being carbon where one or two carbon atoms are optionally replaced by a carbonyl.
  • Substituted cycloalkyl means a cycloalkyl substituted with an alkyl group, wherein alkyl is as defined above or a group as defined above for substituted alkyl.
  • Substituted heteroaryl means a heteroaryl ring, wherein heteroaryl is as defined above, substituted with one or more substituents, preferably one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxy, carboxy, cyano, nitro, alkylthio, and thioalkyl, wherein said substituents are as defined herein.
  • Substituted oxygen refers to the group “-O-R d " wherein R d is alkyl, haloalkyl, alkenyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl, wherein said substituents are as defined herein.
  • Substituted phenyl means a phenyl ring wherein one or more of the hydrogen atoms has been replaced by a halogen, hydroxy, alkyl, amine (primary, secondary, and tertiary with the latter two alkyl substituted), -SH, and phenyl.
  • Representative examples include, but are not limited to, p-bromophenyl, m-iodophenyl, o-chlorophenyl, p-ethylphenyl, m-propylphenyl, o-methylphenyl, and p-octylphenyl.
  • Thioalkyl refers to an alkyl, wherein alkyl is as defined above, substituted with one or more -SH groups provided that if two hydroxy groups are present they are not both on the same carbon atom.
  • thioalkyl groups include, for example, thiomethyl, 2-thioethyl, 2-thiopropyl, and the like.
  • “Therapeutically effective amount” means the amount of a compound or composition that, when administered to a mammal for treating a disease, is sufficient to effect such treatment for the disease.
  • the “therapeutically effective amount” will vary depending on the compound or composition, the disease and its severity and the age, weight, etc., of the mammal to be treated.
  • "Treating" or “treatment” of a disease includes: (1) preventing the disease, i.e.
  • Tautomer refers to an isomer in which migration of a hydrogen atom results in two or more structures.
  • the compounds of the present invention are generally named according to the IUPAC or CAS nomenclature system. Abbreviations that are well known to one of ordinary skill in the art may be used (e.g. "Ph” for phenyl, “Me” for methyl, “Et” for ethyl, “Bn” for benzyl, “h” for hour and “rt” for room temperature).
  • the starting materials and reagents used in preparing these compounds are either available from commercial suppliers such as Toronto Research Chemicals (North York, ON Canada), Aldrich Chemical Co. (Milwaukee, Wisconsin, USA), Bachem (Torrance, California, USA), Emka-Chemie, or Sigma (St.
  • the compounds of this invention will typically contain one or more chiral centers. Accordingly, if desired, such compounds can be prepared or isolated as pure stereoisomers. All such stereoisomers (and enriched mixtures) are included within the scope of this invention, unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents well-known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chiral column chromatography, chiral resolving agents, and the like.
  • an appropriately 7-substititued lincosamine intermediate and an appropriately substituted pyrrolidinyl or piperidyl carboxylic acid are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base.
  • This reaction can be performed with any number of known coupling reagents, such as O-(7-azabenzotriazol-l- yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), 1-hydroxybenzotriazole hydrate (HOBT) with carbodiimides, isobutyl chloroformate, diphenylphosphoryl azide (DPP A), and the like.
  • Suitable organic bases include diisopropylethylamine (DIEA), triethylamine (TEA), pyridine, N-methyl morpholine, and the like.
  • Suitable inert organic solvents which can be used include, for example, N,N-dimethylformamide, acetonitrile, dichloromethane, and the like. This reaction is typically conducted using an excess of carboxylic acid to lincosamine at temperatures in the range of about 0°C to about 50°C. The reaction is continued until completion, which typically occurs in from about 2 to 12 h.
  • 7-substititued lincosamine intermediates as defined in the present invention (i.e., R 2 /R 3 ), are synthesized by methods well known to those of skill in the art from methyl 6-amino-6,8-dideoxy-l-mio-e-rythro- ⁇ -D-galacto-octopyranoside, which can be prepared as described by Hoeksema, et al., Journal of the American Chemical Society, 1967, 89 2448- 2452. Illustrative syntheses for 7-substituted lincosamine intermediates are shown below in Schemes 1-5.
  • Appropriately substituted pyrrolidinyl or piperidyl carboxylic acid intermediates are also synthesized by methods well known to those of skill in the art from prolines and pyridines.
  • the prolines and pyridines that can be used in the synthesis of the carboxylic acid intermediates of the present invention include, for example, 4- oxoproline and 4-substituted pyridines.
  • the prolines and pyridines used in the synthesis are commercially available from vendors such as Aldrich and Sigma. Alternatively, these prolines and pyridines can be prepared by methods well known in the art. Illustrative syntheses for appropriately substituted pyrrolidinyl or piperidyl carboxylic acid intermediates are shown below in Schemes 6-10.
  • Scheme 1 illustrates a general synthesis of a lincosamine intermediate lc wherein P is an N-protecting group, preferably either Cbz or Boc, and R 1 is as defined for formula (I).
  • Suitable N- protecting groups can be formed by the addition of di-t-butyldicarbonate, N- (benzyloxycarbonyloxy) succinimide, and the like.
  • the hydroxy groups can be protected as silyl ethers.
  • the hydroxyl group can be converted to trimethylsilyl (TMS) ethers by reaction with N, O-bis-(trimethylsilyl)trifluoroacetamide in the presence of an appropriate organic base such as triethylamine (TEA) or trimethylsilyl chloride in the presence of an organic base such as triethylamine.
  • TMS trimethylsilyl
  • TAS trimethylsilyl
  • the N-protection is typically accomplished before the O-protection. Chromatography of the crude product on silica after evaporation of the solvent provides the protected product lb.
  • the 7-O-trimethylsilyl group of lb is chemoselectively deprotected and oxidized to provide the 7-keto-lincosamine derivative lc.
  • This selective transformation is performed by addition of the protected product lb to dimethylsulfoxide and oxalyl chloride in an infert organic solvent such as dichloromethane followed by an appropriate organic base such as triethylamine.
  • the transformation may be performed by addition of lb to dimethyl sulfoxide and an appropriate activating agent such as trifluoroacetic anhydride in an inert organic solvent.
  • the reaction is typically conducted at temperatures in the range of approximately -70°C to 80°C.
  • the resulting reaction mixture is stirred at the low temperature and is then allowed to warm to approximately -50°C.
  • the reaction is maintained at this second temperature for approximately 1 h to 3 h.
  • a suitable organic base such as TEA, pyridine, and the like.
  • the reaction mixture is appropriately worked up to provide the product le.
  • the general class of conditions used in the transformation of lb to lc is known in the art as Swern oxidation conditions.
  • Scheme 2 illustrates a general synthesis of a lincosamine intermediate 2b wherein P is an N-protecting group, preferably either Cbz or Boc, R 1 is as defined for formula (I), and one of R 2 and R 3 is hydrogen and the other is as defined for formula (I).
  • P is an N-protecting group, preferably either Cbz or Boc
  • R 1 is as defined for formula (I)
  • R 2 and R 3 is hydrogen and the other is as defined for formula (I).
  • a keto-lincosamine intermediate lc is reacted to form an alkene using the Wittig or Homer- Wadsworth-Emmons reaction.
  • a suitable phosphonium salt or phosphonate is deprotonated using a strong base to form a phosphorus ylide.
  • Suitable phosphonium salts which can be used are alkyltriphenylphosphonium halides, which can be prepared by the reaction of triphenylphosphine and an alkyl halide.
  • Suitable phosphorous compounds include, for example, methyltriphenylphosphonium bromide, diethyl(cyanomethyl)phosphonate and the like.
  • Suitable strong bases which can be used to form the ylide include organolithium reagents, potassium tert-butoxide, and the like.
  • the formation of the phosphorus ylide is typically conducted under an inert atmosphere, such as N 2 , in an inert organic solvent such as toluene, THF, and the like, at low temperatures.
  • the product lc is added to the reaction.
  • the reaction conveniently can be performed at temperatures between -40°C and room temperature and is stirred until completion, typically 1 to 4 h.
  • the resulting organic solution is worked-up and chromatography of the crude product on silica provides the alkene product 2a.
  • the product of 2a may be purified using conventional techniques, such as chromatography and said purified product may be used in the subsequent coupling reaction to yield vinyl lincosamine derivatives of the present invention.
  • the product 2a is then hydrogenated to provide the saturated product 2b.
  • the hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle.
  • the bottle is purged, and charged with H 2 to approximately 50 to 70 psi and shaken until completion, typically approximately 12 to 24 h.
  • the resulting reaction mixture is filtered, e.g., through celite, and rinsed with a polar organic solvent such as methanol.
  • the organic solution is worked up by transferring to a resin funnel containing dry, washed Dowex 50w-400x H* " form and shaken.
  • Scheme 3 illustrates a general synthesis of a lincosamine intermediate 3b wherein P is an N-protecting group, preferably either Cbz or Boc, R 1 is as defined for formula (I), and one of R 2 or R 3 is alkyl and the other is -OH.
  • P is an N-protecting group, preferably either Cbz or Boc
  • R 1 is as defined for formula (I)
  • one of R 2 or R 3 is alkyl and the other is -OH.
  • suitable carbon nucleophiles add to 7-ketolincosamine intermediate lc in suitable inert organic solvents to provide a 7-hydroxy lincosamine intermediate 3b.
  • suitable carbon nucleophiles include methylmagnesium chloride, diethyl zinc, sodium acetylide and the like and suitable inert organic solvents which can be used include THF, diethyl ether, toluene, and the like.
  • the reaction is typically conducted at reduced temperatures, approximately at 0°C, for about 3 to 5 h.
  • the lincosamine intermediate lc is converted to the oxime by stirring in the presence of a suitable reagent such as O-trimethylsilylhydroxylamine, O- alkylhydroxylamine hydrochloride (for example, O-methylhydroxylamine hydrochloride), and the like.
  • a suitable reagent such as O-trimethylsilylhydroxylamine, O- alkylhydroxylamine hydrochloride (for example, O-methylhydroxylamine hydrochloride), and the like.
  • the reaction is typically conducted in a polar organic solvent such as methanol.
  • the reaction conveniently can be conducted at rt in approximately 8 to 24 h.
  • the solvent is removed to provide the N-protected product 4a.
  • Removal of the protecting group can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like.
  • acids such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like
  • THF trifluoroacetic acid
  • the removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product 4b.
  • Scheme 5 illustrates a general synthesis of a lincosamine intermediate 5b wherein R 2 and R 3 are both fluorine, P is an N-protecting group, preferably Cbz or Boc, and R 1 is as defined for formula (I).
  • the lincosamine intermediate lc is contacted with a suitable fluoride in an inert organic solvent.
  • suitable fluorides which can be used include tetrabutylammonium fluoride, Amberlite resin A-26 F form, HF •pyridine and the like.
  • Suitable inert organic solvents include THF, acetonitrile, dichloromethane, dioxane, and the like.
  • the reaction conveniently can be conducted at rt in about 1 to 2 h.
  • the product (not shown) can be purified on a silica gel column.
  • the O-protecting groups on the product obtained from the column are converted by contact with acetic anhydride and dimethylaminopyridine (DMAP) in a suitable mixture of an inert organic solvent and an organic base, such as, for example, dichloromethane and pyridine.
  • DMAP dimethylaminopyridine
  • the reaction conveniently can be conducted at rt in approximately 6 to 12 h.
  • the product can be purified on silica gel column to provide product 5a.
  • the product 5a is contacted with a suitable fluorinating reagent and then the N- protecting group is removed to provide the product 5b.
  • Suitable fluorinating reagents which can be used include, for example, dimethylaminosmfurtrifluoride, [bis(2-me oxyethyl)amino]sul---urtrifluoride, and the like.
  • the reaction is typically conducted in an inert organic solvent such as dichloromethane, ethylacetate, THF, and the like at room temperature in approximately 6 to 12 h.
  • Removal of the protecting group can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like.
  • acids such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like
  • THF trifluoroacetic acid
  • the removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product 5b.
  • R 9 is as defined for formula (I).
  • the product 6c is prepared as described in Birkenmeyer, et al., Journal of Medicinal Chemistry 1972, 15, 1255-1259.
  • Compound 6a is commercially available from vendors such as RSP (Scientific Research Consortium, Inc.).
  • 6a can be prepared from commercially available protected hydroxy prolines by methods well known in the art. See, e.g., Demange, et al., Tetrahedron Letters 1998, 39,1169-1172.
  • Scheme 7 illustrates a general synthesis of trans-R 9 -proline intermediates 7d, wherein R 9 is alkyl or substituted alkyl.
  • a protected 4-oxoproline, 7a is enolated with a suitable enolation agent and then alkylated with a suitable alkylating agent in an inert organic solvent to provide a lactam 7b (wherein R 9 is alkenyl), as described in the literature procedure by Zhang, et al., J.A.C.S. 1998, 120 3894-3902.
  • Compound 7a is commercially available from vendors such as Bachem.
  • 7a can be prepared by methods well known in the art.
  • Suitable enolating agents include LiHMDS, LiN(iPr) 2 , and the like, and suitable alkylating agents include allylic and benzylic bromides, for example, 4-bromo-2-methyl-2-butene and cis-l-bromo-2- pentene, allylbromide, and the like.
  • the lactam 7b is reduced using a suitable reducing agent to provide a pyrrolidine 7c, wherein R 9 is alkenyl.
  • the reduction is preformed by a two-step sequence involving superhydride reduction of the lactam to the hemiaminal and the subsequent reduction of the hemiaminal.
  • Suitable reducing agents which can be used include Et 3 SiH/BF 3 » OEt 2 , Et 3 SiH/TiCl 4 , and the like.
  • the pyrrolidine 7c is then hydrogenated to simultaneously remove the unsaturation in the R substituent and remove the benzyl protecting group from the carboxylic acid to provide the product 7 .
  • the hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle.
  • the bottle is; purged, and charged with H 2 to approximately 50 to 70 psi and shaken until completion, typically approximately 5 to 24 h.
  • the reaction mixture is filtered, e.g., through a celite pad, and washed with a polar organic solvent, such as methanol.
  • the product 7d is ozonolized to provide the aldehyde 8a.
  • the ozonolysis reaction is typically conducted in an anhydrous inert organic solvent, such as dichloromethane, dioxane, THF, and the like, at low temperatures, e.g., -78°C followed by quenching of the reaction with a reducing agent such as DMS, Ph 3 P.
  • a suitable phosphonium salt in the presence of a strong base in an inert organic solvent.
  • Suitable phosphonium salts which can be used include, for example, fluorobenzyl phosphonium chloride, 4-chlorobenzyl phosphonium chloride, dibromofluoromethane and triphenylphosphine, and the like.
  • Suitable bases which can be used include potassium t-butoxide, organolithium reagents, and activated zinc.
  • Suitable organic solvents which can be used include toluene, THF, dimethylacetamide, and the like.
  • the reaction is typically conducted in an inert atmosphere, such as under nitrogen, with vigorous stirring.
  • the reaction is typically conducted at rt to approximately 110°C for 1 to 2 h.
  • the resulting reaction mixture is appropriately worked-up and can be purified by chromatography to provide 8b (wherein R 9 is alkenyl).
  • the product 8b is then hydrogenated to provide the product 8c.
  • the hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% Palladium on carbon in a Parr bottle.
  • the bottle is purged, and charged with H to approximately 40 to 70 psi and shaken until completion, typically approximately 4 to 24 h.
  • the reaction mixture is filtered, e.g., through a celite pad and washed several times with a polar organic solvent, such as methanol. Evaporation of the combined washings and filtrate affords the product 8c, wherein R 9 is an alkyl or substituted alkyl.
  • Scheme 9 illustrates a general synthesis of trans-R 9 -proline intermediates 9d, wherein R 9 is substituted alkyl wherein X is halo.
  • Suitable reagents to reduce and alkylate the aldehyde include tetraallyltin/boron trifluoride etherate, allylTMS/boron trifluoride etherate and suitable inert organic solvents which can be used include THF, dichloromethane, and the like.
  • the reaction is typically conducted at low temperatures, e.g., 0°C, for approximately 1 to 2 h.
  • a suitable fluoride salt in water for example potassium fluoride in water, followed by the addition of methanol.
  • the reaction mixture is filtered, for example, over celite.
  • the product can be purified by chromatography to provide 9a.
  • the hydroxyalkenyl substituted proline, 9a is oxidized to the ketone by contact with a suitable oxidizing agent in an inert organic solvent.
  • suitable oxidizing agents include oxalyl chloride/DMSO, Dess Martin periodinane, and the like.
  • Suitable inert organic solvents include dichloromethane, and the like.
  • the reaction is typically conducted at reduced temperatures, e.g., -72°C to -50°C, for approximately 30 min to 2 h.
  • a suitable organic base such as triethylamine.
  • the reaction mixture is worked up to provide product 9b.
  • the keto-substituted product 9b is halogenated by contact with a suitable halogenating agent in an inert organic solvent.
  • suitable halogenating agents which can be used include diethylaminosulfur trifluoride, [bis(2-methoxyethyl)amino] sulfur trifluoride, and the like.
  • Suitable inert organic solvents which can be used include dichloromethane, ethyl acetate, THF, and the like.
  • the reaction is typically conducted at low temperatures in the range of approximately -30°C to -78°C.
  • the reaction mixture is gradually allowed to warm to rt and stirred at rt until completion, typically in 6 to 12 h.
  • the reaction mixture is worked up and can be purified by chromatography to provide 9c.
  • the product 9c is then hydrogenated to provide the product 9d.
  • the hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle.
  • the bottle is purged, and charged with H 2 to approximately 40 to 70 psi and shaken until completion, typically approximately 4 to 24 h.
  • the reaction mixture is filtered, e.g., through a celite pad and washed several times with a polar organic solvent, such as methanol. Evaporation of the combined washings and filtrate affords the product 9d.
  • Scheme 10 illustrates a general synthesis, as described in Shuman, Journal of Organic Chemistry. 1990, 55, 741-750, of substituted pyridine carboxylic acid intermediates 10b, wherein R 9 is as defined for formula (I).
  • Scheme 10 General synthesis of substituted pyridin-2-yl carboxylic acids 10b.
  • an appropriately substituted pyridine is contacted with a suitable oxidizing agent in an inert organic solvent.
  • the appropriately substituted pyridine starting materials are commercially available from vendors such as Aldrich and Sigma. Alternatively, these pyridines can be prepared by methods well known in the art.
  • Suitable oxidizing agents which can be used include hydrogen peroxide, MCPBA, and the like.
  • the reaction is typically conducted at reflux for 6 to 12 h.
  • the reaction mixture is then contacted with a suitable cyanide reagent to provide the cyano-substituted pyridine, 10a.
  • Suitable cyanide reagents which can be used include trimethylsilyl cyanide, HCN, and the like.
  • Suitable inert organic solvents include dichloromethane, dioxane, THF, and the like.
  • the reaction conveniently can be conducted at rt in approximately 6 to 12 h. The reaction mixture is worked up to provide the cyano-substituted pyridine, 10a.
  • the cyano-substituted pyridine, 10a is then hydrolyzed to provide the pyridin-2-yl carboxylic acid 10b by contact with a suitable acid.
  • suitable acids for hydrolyzing the cyano group to the carboxylic acid include hydrochloric acid, aqueous sulfuric acid, and the like.
  • the reaction is typically conducted at reflux in 6 to 12 h.
  • Scheme 11 illustrates the coupling reaction of a lincosamine intermediate, prepared as described above in Schemes 1-5, and a pyrrolidinyl or piperidinyl carboxylic acid, prepared as described above in Schemes 6-10, wherein R 1 , R 2 , R 3 , R 6 , and R 9 are as defined for formula (I) and P 1 is a suitable O-protecting group and P 2 is a suitable N-protecting group. 11a
  • an appropriately 7-substititued lincosamine intermediate prepared, for example, according to any one of Schemes 1-5) and an appropriately substituted pyrrolidinyl or piperidyl carboxylic acid (prepared, for example, according to any one of Schemes 6-10) are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base.
  • This reaction can be performed with any number of known coupling reagents, such as O-(7-azabenzotriazol-l-yl)- N,N,N',N'tetramethyluronium hexafluorophosphate (HATU), 1-hydroxybenzotriazole hydrate (HOBT) with carbodiimides, diphenylphosphoryl azide (DPP A), isobutyl chloroformate, and the like.
  • Suitable organic bases include diisopropylethylamine (DIEA), triethylamine (TEA), pyridine, N-methyl morpholine, and the like.
  • Suitable inert organic solvents which can be used include, for example, N,N-dimethylformamide, acetonitrile, dichloromethane, and the like. This reaction is typically conducted using an excess of carboxylic acid to lincosamine at temperatures in the range of about 0°C to about 50°C. The reaction is continued until completion, which typically occurs in from about 2 to 12 h.
  • Removal of the protecting groups can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like.
  • acids such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like
  • THF trifluoroacetic acid
  • the removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product.
  • an appropriately 7-substititued lincosamine intermediate prepared, for example, according to any one of Schemes 1-5) and an appropriately substituted pyridin-2-yl carboxylic acid (prepared, for example, according to Scheme 10) are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base, as described above.
  • the pyridine lib is hydrogenated to provide the piperidyl product.
  • the hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using platinum(IN)oxide in the presence of an acid such as HCl, acetic acid, and the like, in a Parr bottle.
  • the bottle- is purged, and charged with H to approximately 40 to 70 psi and shaken until completion, typically approximately 24 h.
  • the reaction mixture is filtered, e.g., through a celite pad, and washed several times with a polar organic solvent such as methanol.
  • Scheme 12 illustrates the alkylation of the nitrogen of the pyrrolidinyl or piperidinyl ring, wherein R 6 is alkyl, hydroxyalkyl, alkylene-substituted heterocycle, or alkylene-heterocycle, and R 1 , R 2 , R 3 , and R 9 are as defined for formula (I).
  • tlie lincosamine 12a can be ⁇ -substituted by contact with an alkylating agent in the presence of a suitable base to provide a product 12b.
  • Suitable alkylating agents which can be used include epoxides, alkyl bromides, and the like.
  • Suitable bases which can be used include potassium carbonate, cesium carbonate triethylamine, and the like.
  • the alkylation reaction is typically conducted in a polar organic solvent such as methanol or DMF.
  • the alkylation reaction is typically conducted at low temperatures in the range of 0°C to -10°C for 10 to 20 h.
  • Scheme 13 illustrates the acylation of the nitrogen of the pyrrolidinyl or piperidinyl ring, wherein R 6 is -C(O)O-alkylene-cycloalkyl, -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O]-alkylene-heterocycle, -[C(O)O]-alkylene-substituted heterocycle, and R 1 , R 2 , R 3 , and R 9 are as defined for formula (I).
  • Scheme 13 General synthesis of 1 '-N-substituted lincosamines.
  • the lincosamine 12a can be N-substituted by contact with an acyl chloride 101, such as R 6 -X, wherein X is a suitable leaving group, and is preferably halogen, even more preferably chloride in the presence of a suitable base to provide a product 102.
  • an acyl chloride 101 such as R 6 -X, wherein X is a suitable leaving group, and is preferably halogen, even more preferably chloride in the presence of a suitable base to provide a product 102.
  • Suitable bases which can be used include DCC, TEA, and the like.
  • reaction is typically conducted in a polar organic solvent such as methanol or DMF.
  • a polar organic solvent such as methanol or DMF.
  • the reaction is typically conducted at low temperatures in the range of -10°C to 20°C.
  • Scheme 18 General synthesis of 2-substituted esters.
  • the following representative reaction scheme may be used in the synthesis of 2- substituted prodrug compounds of the invention: a. (Boc) 2 O, aq. KHCO 3 THF b. j»-anisaldehyde dimethyl acetal, PPTS, c. R ⁇ acylating agent, base d. TFA, DCE, water.
  • Scheme 19 illustrates a general method for the synthesis of 4,5-disubstituted- 2,3,6,7-tetrahydro-azepine-2-carboxylic acid derivatives.
  • Scheme 21 illustrates the synthesis of 4,4 di-substituted pyrrolidine and piperidine carboxylic acids, wherein R 9 is a suitable substituent and the other substituent is fluorine.
  • Scheme 21 Synthesis of intermediate 21d where R >9 is defined herein.
  • the following representative reaction scheme may be used in Scheme 21 : (a) Tetraallyltin, BF 3 » Et 2 O or R 9 M (R 9 or R 9' carbon nucleophile) (b) H 2 /Pd (c) DAST (d) aq. LiOH or appropriate carboxylate ester deprotection conditions.
  • the compounds of the subject invention are usually administered in the form of pharmaceutical compositions. These compounds can be administered by a variety of routes including oral, parenteral, transdermal, topical, rectal, and intranasal. These compounds are effective as both injectable and oral compositions. Such compositions are prepared in a manner well known in the pharmaceutical art and comprise at least one active compound.
  • compositions that contain, as the active ingredient, one or more of the compounds of the subject invention above associated with pharmaceutically acceptable carriers.
  • the active ingredient is usually mixed with an excipient, diluted by an excipient or enclosed within such a carrier which can be in the form of a capsule, sachet, paper or other container.
  • the excipient employed is typically an excipient suitable for administration to human subjects or other mammals.
  • the excipient serves as a diluent, it can be a solid, semi-solid, or liquid material, which acts as a vehicle, carrier or medium for the active ingredient.
  • compositions can be in the form of tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid medium), ointments containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions, and sterile packaged powders.
  • excipients include lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, sterile water, syrup, and methyl cellulose.
  • the formulations can additionally include: lubricating agents such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preserving agents such as methyl- and propylhydroxy-benzoates; sweetening agents; and flavoring agents.
  • the compositions of the invention can be formulated so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient by employing procedures known in the art.
  • the quantity of active component, that is the compound according to the subject invention, in the pharmaceutical composition and unit dosage form thereof may be varied or adjusted widely depending upon the particular application, the potency of the particular ⁇ compound and the desired concentration.
  • compositions are preferably formulated in a unit dosage form, each dosage containing from about 5 to about 100 mg, more usually about 10 to about 30 mg, of the active ingredient.
  • unit dosage forms refers to physically discrete units suitable as unitary dosages for human subjects and other mammals, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient.
  • the compound of the subject invention above is employed at no more than about 20 weight percent of the pharmaceutical composition, more preferably no more than about 15 weight percent, with the balance being pharmaceutically inert carrier(s).
  • the active compound is effective over a wide dosage range and is generally administered in a pharmaceutically or therapeutically effective amount. It will be understood, however, that the amount of the compound actually administered will be determined by a physician, in the light of the relevant circumstances, including the condition to be treated, the severity of the bacterial infection being treated, the chosen route of administration, the actual compound administered, the age, weight, and response of the individual patient, the severity of the patient's symptoms, and the like.
  • the compounds or pharmaceutical compositions thereof will be administered orally, topically, transdermally, and/or parenterally at a dosage to obtain and maintain a concentration, that is, an amount, or blood-level of active component in the animal undergoing treatment which will be antibacterially effective.
  • a dosage that is, an amount, or blood-level of active component in the animal undergoing treatment which will be antibacterially effective.
  • such antibacterially or therapeutically effective amount of dosage of active component i.e., an effective dosage
  • an effective dosage will be in the range of about 0.1 to about 100, more preferably about 1.0 to about 50 mg/kg of body weight/day.
  • the principal active ingredient is mixed with a pharmaceutical excipient to form a solid preformulation composition containing a homogeneous mixture of a compound of the present invention.
  • a solid preformulation composition containing a homogeneous mixture of a compound of the present invention.
  • the active ingredient is dispersed evenly throughout the composition so that the composition may be readily subdivided into equally effective unit dosage forms such as tablets, pills and capsules.
  • This solid preformulation is then subdivided into unit dosage forms of the type described above containing from, for example, 0.1 to about 500 mg of the active ingredient of the present invention.
  • the tablets or pills of the present invention may be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action.
  • the tablet or pill can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former.
  • the two components can be separated by an enteric layer that serves to resist disintegration in the stomach and permit the inner component to pass intact into the duodenum or to be delayed in release.
  • enteric layers or coatings such materials including a number of polymeric acids and mixtures of polymeric acids with such materials as shellac, cetyl alcohol, and cellulose acetate.
  • liquid forms in which the novel compositions of the present invention may be incorporated for administration orally or by injection include aqueous solutions, suitably flavored syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as corn oil, cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles.
  • compositions for inhalation or insufflation include solutions and suspensions in pharmaceutically acceptable, aqueous or organic solvents, or mixtures thereof, and powders.
  • the liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as described supra.
  • the compositions are administered by the oral or nasal respiratory route for local or systemic effect.
  • Compositions in preferably pharmaceutically acceptable solvents may be nebulized by use of inert gases. Nebulized solutions may be inhaled directly from the nebulizing device or the nebulizing device may be attached to a face mask tent, or intermittent positive pressure breathing machine. Solution, suspension, or powder compositions may be administered, preferably orally or nasally, from devices that deliver the formulation in an appropriate manner.
  • Hard gelatin capsules containing the following ingredients are prepared: Quantity Ingredient (mg/capsule Active Ingredient 30.0 Starch 305.0 Magnesium stearate 5.0
  • Quantity Ingredient (mg/capsule Active Ingredient 25.0 Cellulose, microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0
  • a dry powder inhaler formulation is prepared containing the following components: Ingredient Weight % Active Ingredient 5 Lactose 95
  • Tablets each containing 30 mg of active ingredient, are prepared as follows Quantity Ingredient (mg/capsule) Active Ingredient 30.0 mg Starch 45.0 mg Microcrystalline cellulose 35.0 mg Polyvinylpyrrolidone (as 10% solution in sterile water) 4.0 mg Sodium carboxymethyl starch 4.5 mg Magnesium stearate 0.5 mg Talc 1.0 mg Total 120 mg
  • the active ingredient, starch and cellulose are passed through a No. 20 mesh U.S. sieve and mixed thoroughly.
  • the solution of polyvinylpyrrolidone is mixed with the resultant powders, which are then passed through a 16 mesh U.S. sieve.
  • the granules so produced are dried at 50°C to 60°C and passed through a 16 mesh U.S. sieve.
  • the sodium carboxymethyl starch, magnesium stearate, and talc previously passed through a No. 30 mesh U.S. sieve, are then added to the granules which, after mixing, are compressed on a tablet machine to yield tablets each weighing 120 mg. .
  • Capsules each containing 40 mg of medicament are made as follows: Quantity Ingredient (mg/capsule) Active Ingredient 40.0 mg Starch 109.0 mg Magnesium stearate 1.0 mg Total 150.0 mg
  • the active ingredient, starch and magnesium stearate are blended, passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin capsules in 150 mg quantities.
  • Suppositories each containing 25 mg of active ingredient are made as follows: Ingredient Amount Active Ingredient 25 Saturated fatty acid glycerides to 2,000 mg [0163] The active ingredient is passed through a No. 60 mesh U.S. sieve and suspended in the saturated fatty acid glycerides previously melted using the minimum heat necessary. The mixture is then poured into a suppository mold of nominal 2.0 g capacity and allowed to cool.
  • Formulation Example 7 Suspensions, each containing 50 mg of medicament per 5.0 mL dose are made as follows: Ingredient Amount Active Ingredient 50 mg Xanthan gum 4.0 mg Sodium carboxymethyl cellulose (11%) Microcrystalline cellulose (89%) 50.0 mg Sucrose 1.75 g Sodium benzoate lO.O mg Flavor and Color q.v. Purified water to 5.0 mL
  • the active ingredient, sucrose and xanthan gum are blended, passed through a No. 10 mesh U.S. sieve, and then mixed with a previously made solution of the microcrystalline cellulose and sodium carboxymethyl cellulose in water.
  • the sodium benzoate, flavor, and color are diluted with some of the water and added with stirring. Sufficient water is then added to produce the required volume.
  • Formulation Example 8 Quantity Ingredient (mg/capsule) Active Ingredient 15.0 mg Starch 407.0 mg Magnesium stearate 3.0 mg Total 425.0 mg
  • a subcutaneous formulation may be prepared as follows: Ingredient Quantity Active Ingredient 5.0 mg Corn Oil 1.0 mL Formulation Example 10
  • a topical formulation may be prepared as follows: Ingredient Quantity Active Ingredient 1 - 10 g Emulsifying Wax 30 g Liquid Paraffin 20 g White Soft Paraffin to 100 g
  • the white soft paraffin is heated until molten.
  • the liquid paraffin and emulsifying wax are incorporated and stirred until dissolved.
  • the active ingredient is added and stirring is continued until dispersed.
  • the mixture is then cooled until solid.
  • An intravenous formulation may be prepared as follows: Ingredient Quantity Active Ingredient 250 mg Isotonic saline 1000 mg
  • transdermal delivery devices Such transdermal patches may be used to provide continuous or discontinuous infusion of the compounds of the present invention in controlled amounts.
  • transdermal patches for the delivery of pharmaceutical agents is well known in the art. See, e.g., U.S. Patent 5,023,252, issued June 11, 1991, herein incorporated by reference.
  • patches may be constructed for continuous, pulsatile, or on demand delivery of pharmaceutical agents.
  • Indirect techniques which are generally preferred, usually involve formulating the compositions to provide for drug latentiation by the conversion of hydrophilic drugs into lipid- soluble drugs. Latentiation is generally achieved through blocking of the hydroxy, carbonyl, sulfate, and primary amine groups present on the drug to render the drug more lipid soluble and amenable to transportation across the blood-brain barrier.
  • the delivery of hydrophilic drugs may be enhanced by infra-arterial infusion of hypertonic solutions which can transiently open the blood-brain barrier.
  • the compounds described herein are suitable for use in a variety of drug delivery systems described above. Additionally, in order to enhance the in vivo serum half-life of the administered compound, the compounds may be encapsulated, introduced into the lumen of liposomes, prepared as a colloid, or other conventional techniques may be employed which provide an extended serum half-life of the compounds. A variety of methods are available for preparing liposomes, as described in, e.g., Szoka, et al., U.S. Patent Nos. 4,235,871, 4,501,728 and 4,837,028 each of which is incorporated herein by reference. [0176] As noted above, the compounds administered to a patient are in the form of pharmaceutical compositions described above.
  • compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered.
  • the resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration.
  • the pH of the compound preparations typically will be between 3 and 11 , more preferably from 5 to 9 and most preferably from 7 and 8. It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.
  • the compounds of the subject invention will be administered in a therapeutically effective amount by any of the accepted modes of administration for agents that serve similar utilities. Toxicity and therapeutic efficacy of such compounds can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD 5 o (the dose lethal to 50% of the population) and the ED 5 o (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD 5 o/ ED50. Compounds that exhibit large therapeutic indices are preferred.
  • the data obtained from the cell culture assays and animal studies can be used in formulating a range of dosage for use in humans.
  • the dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity.
  • the dosage may vary within this range depending upon the dosage form employed and the route of administration utilized.
  • the therapeutically effective dose can be estimated initially from cell culture assays.
  • a dose may be formulated in animal models to achieve a circulating plasma concentration range which includes the IC50 (the concentration of the test compound which achieves a half-maximal inhibition of symptoms) as determined in cell culture.
  • IC50 the concentration of the test compound which achieves a half-maximal inhibition of symptoms
  • levels in plasma may be measured, for example, by high performance liquid chromatography.
  • the compounds, prodrugs and pharmaceutically acceptable salts thereof, as defined herein, may have activity against a variety of bacteria, protozoa, fungi, and parasites.
  • the compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against gram positive and gram negative bacteria.
  • the compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against a variety of fungi, including fungi from the genus Mucor and Candida, e.g., Mucor racemosus or Candida albicans.
  • the compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against a variety of parasites, including malaria and cyptosporidium parasite.
  • the compounds of the subject invention exhibit activity against a variety of bacterial infections, including, for example, gram positive infections, gram negative infections, mycobacteria infections, mycoplasma infections, and chlamydia infections. [0181] Since the compounds of the subject invention exhibit potent activities a variety of bacteria, such as gram positive bacteria, the compounds of the present invention are useful antimicrobial agents and may be effective against a number of human and veterinary pathogens, including gram positive bacteria.
  • the Gram positive organisms against which the compounds of tlie present invention are effective include Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidennidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile, and the like.
  • the compounds of the subject invention may be combined with one or more additional antibacterial agents.
  • One or more of the additional antibacterial agents may be active against gram negative bacteria. Additionally, one or more of the additional antibacterial agents may be active against gram positive bacteria.
  • the combination of the compounds of the subject invention and the one or may additional antibacterial agents may be used to treat a gram negative infection. Additionally, the combination of the compounds of the subject invention and the one or more additional antibacterial agents may be used to treat a gram positive infection.
  • the combination of compounds of the subject invention and the one or more additional antibacterial agents may also be used to treat a mycobacteria infection, mycoplasma infection, or chlamydia infection.
  • the in vitro activity of compounds of the subject invention may be assessed by standard testing procedures such as the determination of minimum inhibitory concentration (MIC) by agar dilution as described in "Approved Standard. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically," 3 rd ed., published 1993 by the National Committee for Clinical Laboratory standards, Villanova, Pennsylvania, USA.
  • MIC minimum inhibitory concentration
  • the amount administered to the mammalian patient will vary depending upon what is being administered, the purpose of the administration, such as prophylaxis or therapy, the state of the patient, the manner of administration, and the like.
  • compositions are administered to a patient already suffering from a disease in an amount sufficient to cure or at least partially arrest the symptoms of the disease and its complications.
  • An amount adequate to accomplish this is defined as "therapeutically effective dose.” Amounts effective for this use will depend on the disease condition being treated as well as by the judgment of the attending clinician depending upon factors such as the severity of the inflammation, the age, weight and general condition of the patient, and the like.
  • the compositions administered to a patient are in the form of pharmaceutical compositions described above. These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered.
  • the resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration.
  • the pH of the compound preparations typically will be between 3 and 11, more preferably from 5 to 9 and most preferably from 7 to 8. It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.
  • the therapeutic dosage of the compounds of the present invention will vary according to, for example, the particular use for which the treatment is made, the manner of administration of the compound, the health and condition of the patient, and the judgment of the prescribing physician.
  • the dose will typically be in the range of about 20 mg to about 500 mg per kilogram body weight, preferably about 100 mg to about 300 mg per kilogram body weight.
  • Suitable dosage ranges for intranasal administration are generally about 0.1 mg to 1 mg per kilogram body weight.
  • Effective doses can be extrapolated from dose-response curves derived from in vitro or animal model test systems.
  • D doublet
  • DAST dimethylaminosul-furtrifluoride
  • dd doublet of doublets
  • ddddd doublet of doublets of doublet of doublets
  • dt doublet of triplets
  • DCC dicyclohexylcarbodiimide
  • DCE dicholoroethane
  • DCM dichloromethane
  • DIEA diisopropyethylamine
  • DMSO dimethyl sulfoxide
  • DPPA diphenylphosphoryl azide
  • EDso dose therapeutically effective in 50% of the population
  • ESMS electrospray mass spectrometry
  • Et ethyl EtOAc ethyl acetate Et 2
  • 0 diethyl ether
  • HATU O-(7-azabenzotriazol- 1 -yl)-N,N,N ' ,N' - tetramethyluronium hexafluorophosphate
  • HBTU O-(Benzotriazol-l-yl)-N,N,N',N*- tetramethyluronium hexafluorophosphate
  • HOBT 1-hydroxybenzotriazole hydrate
  • Aldrich indicates that the compound or reagent used in the following procedures is commercially available from Aldrich Chemical Company, Inc., 1001 West Saint Paul Avenue, Milwaukee, Wl 53233 USA; the term “Fluka” indicates that the compound or reagent is commercially available from Fluka Chemical Corp., 980 South 2nd Street, Ronkonkoma NY 11779 USA; the term “Lancaster” indicates that the compound or reagent is commercially available from Lancaster Synthesis, Inc., P.O. Box 100 Windham, NH 03087 USA; the term “Sigma” indicates that the compound or reagent is commercially available from Sigma, P.O. Box 14508, St.
  • RSP indicates that the compound or reagent is commercially available from RSP Amino Acid Analogs, Inc., 106 South St., Hopkinton, MA 01748, USA
  • TCI indicates that the compound or reagent is commercially available from TCI America, 9211 North Harborgate St., Portland, Oregon, 97203, OR, USA
  • Toronto indicates that the compound or reagent is commercially available from Toronto Reasearch Chemicals, Inc., 2 Brisbane Rd., New York, ON, Canada M3 J2J8
  • Alfa indicates that the compound or reagent is commercially available from Johnson Matthey Catalog Company, Inc.
  • the reaction mixture was cooled to 0°C and then bis-N,0-trifluoroacetamide (15.7 mL, 59.0 mmol) was added by syringe over 2 min.
  • the reaction mixture was allowed to warm to rt and stirred for 42 h.
  • Toluene (100 mL) was added and the reaction mixture was evaporated to dryness.
  • the residue was taken up in ethyl acetate (400 mL).
  • the organic solution was washed quickly with 10% citric acid (200 mL), H 2 0 (3 x 100 mL), saturated NaHC0 3 (100 mL), and brine (2 x 100 mL), and dried over Na 2 S0 4 and evaporated to dryness.
  • the reaction mixture was warmed to rt and stirred for 30 minutes, and then was added to an activated zinc (0.25g, 3.82 mmol, 3.3 equiv) with the aid of dimethyl acetamide (0.3 mL).
  • the resulting reaction mixture was stirred at 110°C for 1 h and cooled to rt and filtered with the aid of dimethylacetamide (7 mL).
  • the organic layer was separated, washed with brine, dried over sodium sulfate and evaporated to dryness.
  • the resulting crude fluorinated product was purified by silica gel column chromatography (hexanes-EtO Ac 9:1).
  • the residue obtained by chromatographic purification was dissolved in dioxane (65 mL) and water (26 mL), cooled to 0 ° C, and treated with Os0 4 ⁇ (0.65 mL, 4% aq. solution) and 30% H 2 0 2 (10 mL).
  • the reaction mixture was stirred overnight and concentrated to dryness.
  • the residue was dissolved in DCM and the organic layer washed with water (100 mL), 25% aq.
  • the crude product was eluted from the resin by washing with 5% TEA in MeOH (25 mL x 15 min x 2) and 5% TEA in MeCN (25 mL x 15 min).
  • the combined eluent was evaporated to dryness and purified by silica gel column chromatography using 7% 0.25M NH 3 in methanol in dichloromethane as the eluent (24 mg, 28%).
  • the residual oil obtained was diluted with EtOAc (400 mL), washed sequentially with 10% citric acid (2 x 100 mL), 1:1 saturated aqueous NaHC0 3 , water (2 x 100 mL), and brine (100 mL), dried over Na 2 S0 , and evaporated to dryness.
  • the crude product was purified by column chromatography using 30% ethyl acetate in hexanes as the eluent (200 mg, 57%). Potassium carbonate (450 mg, 3.0 mmol) was added to the product (200 mg, 0.30 mmol) of the above reaction in methanol (3 mL) and water (1 mL), and the mixture was stirred at rt for 2 h. The solvent was removed and the residue obtained was taken in 30%) trifluoroacetic acid in dichloroethane (10 mL) and dimethyl sulfide (0.5 mL) and stirred for 1 h. After removing the solvent, the crude product obtained was purified by column using 10%) methanol in dichloromethane as the eluent (10 mg, 90%).
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate (100 mL), and washed with 10% citric acid (2 x 60 mL), water (60 mL), half sat. NaHCO 3 (60 mL), and brine.
  • the organic layer was dried over Na 2 SO and evaporated to give a yellow syrup.
  • Trifluoroacetic acid (5 mL) and water (0.33 mL) were added to a solution of the above syrup in dichloromethane (15 mL) with methyl sulfide (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice.
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate (100 mL), and washed with 10% citric acid (2 x 60 mL), water (60 mL), half sat. NaHCO 3 (60 mL), and brine.
  • the organic layer was dried over Na 2 SO 4 and evaporated to give a yellow syrup.
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), and washed with 10%) citric acid (2 x 40 mL), water (40 mL), half sat. NaHCO 3 (40 mL), and brine.
  • the organic layer was dried over Na 2 SO 4 and evaporated to give a yellow syrup.
  • Trifluoroacetic acid (3 mL) and water (0.2 mL) were added to a solution of the above syrup in dichloromethane (9 mL) with methyl sulfide (0.2 mL).
  • the reaction mixture was stirred at rt for 1 h.
  • the solvent was removed under vacuum and co-evaporated with toluene twice.
  • the reaction mixture was stirred at rt for 3h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), and washed with 10% citric acid (2 x 40 mL), water * (40 mL), half sat. NaHCO 3 (40 mL), and brine.
  • the organic layer was dried over Na SO 4 and evaporated to give a yellow syrup.
  • Trifluoroacetic acid (3 mL) and water (0.20 mL) were added to a solution of the above syrup in dichloromethane (9 mL) with methyl sulfide (0.20 mL).
  • the reaction mixture was stirred at rt for 1 h.
  • the solvent was removed under vacuum and co-evaporated with toluene twice.
  • the residue was purified by chromatography to provide 4-(2,2-Difluoro-pentyl)-pyrrolidine-2- carboxylic acid [2-met-hyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]- amide (56 mg, 62%) as a white solid.
  • Ethylene oxide (0.6 mL) was added to a solution of 4-(3-methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide (35.1 mg, 0.084 mmol), prepared as in Example 9, in methanol (3 mL), at 0°C. The reaction mixture was stirred at 4°C overnight. Additional ethylene oxide (0.6 mL) was added and stirred at 4°C overnight.
  • reaction mixture was concentrated and purified by chromatography to give a white solid, l-(2-hydroxy-ethyl)-4-(3-methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl-l - (3,4,5-trihydroxy-6-methylsulf-----j ⁇ yl-tetrahydro-pyran-2-yl)-propyl]-amide (24.1 mg, 62%).
  • Ethylene oxide (0.4 mL) was added to a solution of 4-(3,3-difluoro-propyl)- pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propyl]-amide, prepared as in Example 14 (29.7 mg, 0.07 mmol), in methanol (2 mL), at 0°C. The reaction mixture was stirred at 4°C overnight. Additional ethylene oxide (0.4 mL) was added and stirred at 4°C overnight.
  • reaction mixture was concentrated and purified by chromatography to give a white solid, l-[2-(S)-4-(R)-(3-methylbut-l-yl)-N-(2-hydroxyeth-l- yl)pyrrolidin-2-yl]-N- ⁇ l-(R)- [2-(S)-3-(S), 4-(S), 5-(R)-trihydroxy-6-(R)- (methylthio)tetrahydropyran-2-yl]-2-methylprop-l-yl ⁇ acetamide (19.3 mg, 59%).
  • Example 2 To the product of Example 1 (28 mg, 0.07 mmol) in methanol (2 mL), ethylene oxide (0.5 mL) was added and stirred at 4 °C overnight. The solvent was removed and the resulting product was purified by column, chromatography using 20%> MeOH in DCM as eluent to obtain the title compound (16 mg, 51%) as a white powder.
  • Example 17 To the product of Example 17 (50 mg, 0.123 mmol) in water (3 mL) and dioxane (3 mL), ethyl chloroformate (20 mg, 0.147 mmol) and sodium carbonate (25 mg, 0.246 mmol) were added and stirred over night at room temperature. The solvents were removed and the crude material was loaded into a silica gel column and eluted with ethyl acetate to obtain the title compound as a white solid (40 mg, 52%).
  • HATU (96.8 mg, 0.25 mmol, 1.25 equiv).
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid, water, sat. NaHC0 3 and brine.
  • the organic layer was dried over Na S0 and evaporated and used in the next step without additional purification.
  • the reaction mixture was added to the acid 4-(3,3-difluoro-butyl)-pyrrolidine-l,2-diearboxylic acid 1-tert-butyl ester (59.7 mg, 0.20 mmol, 1 equiv) in a 25 mL round bottom flask, followed by the addition of HATU (93.3 mg, 0.25 mmol, 1.25 equiv).
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid, water, sat. NaHC0 3 and brine.
  • the organic layer was dried over Na S0 4 and evaporated to give a syrup.
  • reaction mixture was added to the acid 4-(3,3-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester (76.6 mg, 0.24 mmol, 1.2 equiv) in a 25 mL round bottom flask, followed by the addition of HATU (111.9 mg, 0.29 mmol, 1.5 equiv).
  • HATU 111.9 mg, 0.29 mmol, 1.5 equiv.
  • the reaction mixture was stirred at rt for 3 h.
  • the reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), washed with 10% citric acid (30 mL), water (30 mL), sat. NaHC0 3 (30 mL) and brine.
  • the organic layer was dried over Na 2 S0 4 and evaporated to give a yellow oil.
  • compound 14a 2 g, 7.60 mmol, 1 equiv
  • triphenylphosphine 159.4 mg, 0.61 mmol, 0.08 equiv
  • copper (I) iodide 115.8 mg, 0.61 mmol, 0.08 equiv
  • palladium acetate 68.2 mg, 0.30 mmol, 0.04 equiv
  • triethylamine 28 mL.
  • the mixture was deaerated with nitrogen, followed by addition of 4-pentyn-l-ol (1.28 g, 15.21 mmol, 2 equiv).
  • the reaction mixture was stirred at -72 °C for 25 minutes, then warmed to -50 °C and stirred for an additional 2 h.
  • Triethylamine (1.48 mL, 10.7 mmol, 4.33 equiv) was added and stirred at -50 °C for 25 minutes.
  • the mixture was diluted with ethyl acetate, washed with water (2x), sat. aqueous NaHCO 3 (lx), brine (lx), dried, evaporated and co-evaporated with anhydrous toluene to 4-(5-oxo-pentyl)-pyridine-2-carboxylic acid methyl ester (0.48 mg, 88 %).
  • Triethylamine (0.68 mL, 4.9 mmol, 4.0 equiv), followed by BSTFA (0.58 mL, 2.20 mmol, 1.8 equiv), were added to a stirred suspension of compound prepared above (351 mg, 1.22 mmol, 1 equiv) in anhydrous DMF (5 mL) at 0 °C and under nitrogen. The resulting mixture was stirred at 0 °C for 10 min, and then at rt for 50 min.
  • Example 2 To the product of Example 1 (30 mg, 0.07 mmol) in DMF (1 mL), DIEA (43 mg, 0.35 mmol) was added at room temperature and stirred overnight. Then removed the solvent and the resulting product was purified by column chromatography using 20%) MeOH in DCM to obtain the title compound (20 mg, 66%>) as a white powder.
  • 4-Isobutyl-2-cyanopyridine is prepared as follows. To isobutylfriphenylphosphomum iodide (Aldrich) (8 g, 18.5 mmol) in THF (20 mL) at 0 °C, potassium-tert-butoxide (1.8 g, 16 mmol) was added, stirred at room temperature for 1 hr. pyridine-4-carboxlaldehyde (1 g, 9.3 mmol) was added, stirred at room temperature for overnight. Reaction mixture was then poured to water (100 mL) and extracted with EtOAc (100 mL).
  • reaction mixture was extracted with DCM (50 mL) and washed with NaHC0 3 (30 mL, 10%).
  • the product obtained after removal of solvent was purified by silica gel chromatography using 5% EtOAc in hexanes to obtain N-(tert-butoxycarbonyl)-L-proline-4-fluoro-4-butane (276 mg, 52%).
  • Triethylsilane (0.16 mL), TFA (2 mL) and water (0.16 mL) was added, stirred at room temperature for 1.5 hours.
  • the residue obtained on removal of solvent was purified by column chromatography using 10% MeOH in DCM to obtain the title compound (156 mg, 96%).
  • N-(tert-butoxycarbonyl)-4-fluoro-4-propyl-L-proline was prepared as described in the previous example (except using n-propyl lithium in the place of n-butyl lithium) (164 mg, 0.57 mmol) was suspended in dry acetonitrile (4 mL). Triethylamine (332 ⁇ L, 3.02 mmol) was added and the reaction mixture was cooled to 0°C. Isobutyl chloroformate (78 ⁇ L, 0.57 mmol) was added and after 10 min the reaction was allowed to warm to 4°C.
  • Examples 60 to 62 were prepared using methods and techniques describe herein utilizing commercially available starting materials where appropriate.

Abstract

Novel lincomycin derivatives are disclosed. These lincomycin derivatives exhibit antibacterial activity. As the compounds of the subject invention exhibit potent activities against bacteria, including gram positive organisms, they are useful antimicrobial agents. Methods of synthesis and of use of the compounds are also disclosed.

Description

LINCOMYCIN DERIVATIVES POSSESSING ANTIBACTERIAL ACTIVITY
CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application is a continuation-in-part of U.S Patent Application No. 10/777,455, filed February 11, 2004, which is a continuation-in-part of U.S. Patent Application No. 10/642,807, filed August 15, 2003, which claims the benefit under 35 U.S.C. §119(e) of U.S. Provisional Application No. 60/403,770 filed on August 15, 2002, and which also claims the benefit of U.S. Provisional Application No. 60/479,502, filed June 17, 2003, the disclosures of which are hereby incorporated by reference in their entirety.
BACKGROUND OF THE INVENTION
FIELD OF THE INVENTION [0002] This invention relates to lincomycin derivatives that exhibit antibacterial activity.
STATE OF THE ART [0003] Lincomycin is a biosynthetic product that adversely affects growth of various microorganisms, in particular gram positive bacteria. The characteristics and preparation of lincomycin are disclosed in U.S. Patent 3,086,912. A variety of derivatives of lincomycin, which also have antimicrobial activity, have been prepared. These derivatives include, for example, clindamycin, which is described in U.S. Patent 3,496,163. [0004] Lincomycin derivatives remain attractive targets for antibacterial drug discovery. Accordingly, lincomycin derivatives that possess antimicrobial activity are desired as potential antibacterial agents.
SUMMARY OF THE INVENTION [0005] The present invention provides lincomycin derivatives that possess antibacterial activity.
[0006] In one of its composition aspects, this invention is directed to a compound of formula (I): wherein: R1 is alkyl; R2 and R3 are independently H, alkyl, hydroxy, fluoro, or cyanoalkyl or one of R2 and R3 is =NOR7 and the other is absent, or one of R2 and R3 is =CH2 and the other is absent, with the provisos that both R2 and R3 are not H; when one of R2 and R3 is fluoro, the other is not hydrogen or hydroxy; and when one of R2 and R3 is hydroxy, the other is not fluoro, hydrogen, or hydroxy; R6 is selected from the group consisting of H, alkyl, hydroxyalkyl, -C(O)O-alkylene-cycloalkyl, -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O]p-alkylene-heterocycle, -[C(O)O]p-alkylene- substituted heterocycle, wherein p is 0 or 1 ; R7 is H or alkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH )n-OH, -(CH2)n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R and R5 are H or alkyl; and m is 0, 1, 2 or 3; and prodrugs, tautomers or pharmaceutically acceptable salts thereof; with the proviso that the compound of formula I has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile.
[0007] In one embodiment, m is 0. In another embodiment, m is 1. In another embodiment, m is 2. In another embodiment, m is 3.
[0008] In a preferred embodiment, R1 is methyl.
[0009] In one embodiment, one of R2 and R3 is H, and the other is alkyl. In one embodiment, one of R2 and R3 is H, and the other is methyl. In one embodiment, one of R2 and
R3 is H, and the other is cyanoalkyl. In one embodiment, both of R2 and R3 are F. In one embodiment, one of R2 and R3 is hydroxy, and the other is alkyl. In one embodiment, R2 and R3 together are hydroxyimino or alkoxyimino. In one embodiment, R2 and R3 together are methylene (=CH2). In a preferred embodiment, R is H, alkyl, or hydroxyalkyl. In a preferred embodiment, R6 is selected from the group consisting of H, methyl, ethyl, 2- hydroxyethyl, 2-methyl-2-hydroxyethyl, and 3-hydroxypropyl. In another embodiment, R6 is not one of the following:
[0010] In one embodiment, each R9 is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, cycloalkylalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, - (CH )n-OH, -(CH )n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl.
[0011] In one embodiment, R9 is a substituted alkyl, and wherein the one or more substituents on the alkyl group are selected from the group consisting of halogen, oxygen, hydroxy, amine (primary), amine (secondary-amine substituted by alkyl above), amine (tertiary- amine substituted by alkyl as above), sulfur, -SH, and phenyl. [0012] In a preferred embodiment, each R9 is independently selected from the group consisting of halogen, alkyl, substituted alkyl, alkoxy, alkoxyalkoxy, substituted alkoxy, and cycloalkyl. In another preferred embodiment, each R9 is independently selected from the group consisting of H, alkyl, and substituted alkyl.
[0013] In one embodiment, R9 is selected from the group consisting of 1-ethylpropyl; 2-(4- fluorophenyl)ethyl; 2,2-difluoroethoxymethyl/fluoro; 2,2-difluoropentyl; 2-cyclobutyl-ethyl; 2- cyclopropyl-ethyl; 2-fluoroethoxy; 2-methoxyethoxy; 2-propoxy-ethyl; 3-(2-fluoro-ethoxy)- propyl; 3-(3-fluoro-propoxy)-propyl; 3-(4-chlorophenyl)propyl; 3-(cyclopropyl-methoxy)- propyl; 3,3,3-trifluoropropoxy; 3,3-difluorobutyl; 3,3-difluoropentyl; 3,3-difluoropropyl; 3,3- difluoropropyl/fluoro; 3-cyclobutyl-propyl; 3-cyclopentyl-propyl; 3-cyclopropyl-propyl; 3- ethoxy-propyl; 3-fluoropropoxy; 3-fluoro-propoxy-methyl; 3-fluoropropyl; 3- fluoropropyl/fluoro; 3-hydroxy-3-ethylpentyl; 3-methoxy-propyl; 3-propoxy-propyl; 4,4- difluorobutyl; 4,4-difluoropentyl; 4-fluorobutoxy; 4-fluorobutyl; 4-methoxy-butyl; 5,5- difluoropentyl; 5-fluoropentyl; butoxy; butyl; cyclobutyl-methyl; cyclohexyl; cyclohexyl- methyl; cyclopropyl-methoxy; cyclopropyl-methyl; ethyl; ethyl/fluoro; fluoro/butyl; fluoro/propyl; isobutyl; isopentyl; iso-propyl; methoxy; n-butyl; n-pentyl; pentoxy; pentyl; propoxy-methyl; and propyl. In another embodiment, R9 is propyl.
[0014] In one embodiment, R9 is selected from the group consisting of 1-ethylpropyl; 2-(4- fluorophenyl)ethyl; 2,2-difluoropentyl; 2-cyclobutyl-ethyl; 2-cyclopropyl-ethyl; 2- methoxyethoxy; 3-(2-fluoro-ethoxy)-propyl; 3-(4-chlorophenyl)propyl; 3,3-difluorobutyl; 3,3- difluoropentyl; 3,3-difluoropropyl; 3-cyclobutyl-propyl; 3-cyclopentyl-propyl; 3-cyclopropyl- propyl; 3-ethoxy-propyl; 3-fluoropropyl; 3-hydroxy-3-ethylpentyl; 3-methoxy-propyl; 3- propoxy-propyl; 4,4-difluorobutyl; 4-fluorobutyl; 5,5-difluoropentyl; 5-fluoropentyl; butoxy; butyl; cyclobutyl-methyl; cyclohexj ; cyclohexyl-methyl; cyclopropyl-methoxy; cyclopropylmethyl; ethyl; isobutyl; isopentyl; iso-propyl; methoxy; n-butyl; n-pentyl; pentoxy; pentyl; propoxy-methyl; and propyl.
[0015] In one embodiment, R9 not 2,2-difluoroethoxymethyl/fluoro; 2-fluoroethoxy; 2- propoxy-ethyl; 3-(3-fluoro-propoxy)-propyl; 3-(cyclopropyl-methoxy)-propyl; 3,3,3- trifluoropropoxy; 3,3-difluoropropyl/fluoro; 3-fluoropropoxy; 3-fluoro-propoxy-methyl; 3- fluoropropyl/fluoro; 4,4-difluoropentyl; 4-fluorobutoxy; 4-methoxy-butyl; ethyl/fluoro; fluoro/butyl; or fluoro/propyl. [0016] In one of its composition aspects, this invention is directed to a prodrug compound of the formula (IN):
wherein R1, R2, R3, and R9 are as defined for formula I; wherein R6 is as defined for formula I or is a suitable prodrug group which is cleavable in vivo; and wherein R11 is H or a suitable prodrug group which is cleavable in vivo.
[0017] In one embodiment, the compound is a prodrug wherein R6 is selected from the group consisting of hydrogen; l-(acetyloxy)-ethyl-oxycarbonyl; l-amino-2-methyl-butyl- carbonyl; 1 -amino-2-methyl-butyl-carbonyl-oxy-methyl-oxycarbonyl; 1 -amino-2-methyl- propyl-carbonyl; l-amino-2-phenyl-ethyl-carbonyl; 1-amino-ethyl-carbonyl; 1-methyl-l, 2,3,6 tetrahydro-pyridin-4-yl-oxycarbonyl; 1-methyl-l, 4 dihydro-pyridin-3-yl-carbonyl; l-methyl-3- oxo-but-1-enyl; 5-methyl-[l,3]dioxol-2-one-4-yl-methoxy-carbonyl; 5-methyl-[l,3]dioxol-2- one-4-yl-methyl; ethoxy-carbonyl; ethyl-carbonylamino-methyl; fluorenyl-methylene-oxy- carbonyl; phenoxy-carbonyl; piperidin-4-yl-carbonyl-oxy-methyl-oxycarbonyl; and pyridine-3- yl-carbonylamino-methyl.
[0018] In another embodiment, the compound is a prodrug wherein R6 is selected from the group consisting of hydrogen; l-(acetyloxy)-ethyl-oxycarbonyl; l-amino-2-methyl-butyl- carbonyl; 1 -amino-2 -methyl-butyl-carbonyl-oxy-methyl-oxycarbonyl; 1 -amino-2-methyl- propyl-carbonyl; l-amino-2-phenyl-ethyl-carbonyl; 1-amino-ethyl-carbonyl; 1-methyl-l, 2,3, 6 tetrahydro-pyridin-4-yl-oxycarbonyl; l-methyl-1,4 dilιydro-pyridin-3-yl-carbonyl; l-methyl-3- oxo-but- 1 -enyl; ethyl-carbonylamino-metliyl; piperidin-4-yl-carbonyl-oxy-methyl-oxycarbonyl; and pyridine-3-yl-carbonylamino-methyl.
[0019] In another embodiment, the compound is a prodrug wherem R is not 5-methyl- [1 ,3]dioxol-2-one-4-yl-methoxy-carbonyl; 5-methyl-[l ,3]dioxol-2-one-4-yl-methyl; ethoxy- carbonyl; fluorenyl-methylene-oxy-carbonyl; or phenoxy-carbonyl. [0020] In another embodiment, the compound is a prodrug wherein R6 is not 5-methyl- [1 ,3]dioxol-2-one-4-yl-methoxy-carbonyl; or 5-methyl-[l ,3]dioxol-2-one-4-yl-methyl. [0021] In a preferred embodiment, the compound is a prodrug wherein R11 is selected from the group consisting of hydrogen; 2-(N-(2-morpholin-4-yl-ethyl)-amino-carbonyl)-ethyl- carbonyl; -C(O)CH2CH2COOH; N,N-dimethyl-amino-methyl-carbonyl; pentadecyl- ' carbonyloxy; and -PO3H2.
[0022] In a preferred embodiment, this invention provides compounds of formula (II)
wherein: R1 is alkyl; R2 and R3 are independently H, alkyl, or cyanoalkyl, with the proviso that both R2 and R3 are not H; R6 is H, alkyl, or hydroxyalkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH )n-OH, - (CH2)n-ΝR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 1 or 2; and prodrugs and pharmaceutically acceptable salts thereof; with the proviso that the compound of formula II has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile. [0023] In a particularly preferred embodiment, this invention provides compounds of formula (III):
(HI)
wherein: R1 is alkyl; R2 and R3 are fluoro; R is H, alkyl, or hydroxyalkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH2)n-OH, - (CH2)n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 1 or 2; and prodrugs and pharmaceutically acceptable salts thereof, with tl e proviso that the compound of formula III has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile. In another preferred embodiment, this invention is directed to a compound of formula (IV):
wherein: R1 is alkyl; R and R are independently H, or alkyl, hydroxy, fluoro, or cyanoalkyl or one of R2 and R3 is =NOR7 and the other is absent, or one of R2 and R3 is =CH2 and the other is absent, with the provisos that both R2 and R3 are not H; when one of R2 and R is fluoro, the other is not hydrogen or hydroxy; and when one of R and R is hydroxy, the other is not fluoro, hydrogen, or hydroxy; R6 is selected from the group consisting of -C(O)O-alkylene-cycloalkyl, -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O]p-alkylene-heterocycle, -[C(O)O]p-alkylene-substituted heterocycle, wherein p is 0 or 1 with the proviso that -C(O)O-substituted alkyl does not include the following:
R7 is H or alkyl; R9, which can be singly or multiply substituted in tlie ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH2)n-OH, - (CH )n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 1 or 2; and prodrugs, tautomers or pharmaceutically acceptable salts thereof; with the proviso that the compound of formula I has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile. [0024] Lincomycin derivatives within the scope of this invention include those set forth in Tables I and II as follows:
Table I
Includes R and/or S isomers as either individual isomers or as a mixture
Table II
Includes R and/or S isomers as either individual isomers or as a mixture
[0025] As used below, these compounds are named based on acetamide or amide derivatives but, alternatively, these compounds could have been named based on 1-thio-L-threo-β-D- galacto-octopyranoside derivatives. Specific compounds within the scope of this invention include the following compounds: 4-Ethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 1 -Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [3-cyano-2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Ethyl-piperidine-2 -carboxylic acid [2-hydroxy-2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-hydroxyimino- 1 -(3 ,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-methoxyimino-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 5 -Butyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4, 5 -trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-Pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(354,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2,2-difluoro- 1 -(3 ,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Pentyl-pyrrolidine-2-carboxylic acid [2,2-difluoro- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(4-Fluoro-phenyl)-propyl]-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3,3-Difluoro-propyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(4-Chloro-phenyl)-propyl]-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2,2-Difluoro-pen1yl)-pyιτolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2-Hydroxy-ethyl)-4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5- frihydroxy-6-methylsulfanyl-tet-rahydro-pyran-2-yl)-propyl]-amide; 1 -(2-Hydroxy-propyl)-4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- frihydroxy-6-me ylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; l-(3-Hydroxy-propyl)-4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2-Hydroxy-ethyl)-4-(3 -methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 ,3 -Difluoro-propyl)- 1 -(2-hydroxy-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 ■ (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2-Hydroxy-ethyl)-4-pentyl-pyrrolidine-2-carboxylic acid [2,2-difluoro- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Pentyl-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-Methoxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(l-Ethyl-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Isopropyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Butyl-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclohexyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Ethyl- 1 -(2-hydroxy-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; l-(2-Hydroxy-ethyl)-4-pentyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-methylsulf-myl-tetrahydro-pyr-m-2-yl)-propyl]-arnide; 1 -(2-Hydroxy-ethyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4,4-Difluoro-pentyl)-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3,3-Difluoro-butyl)-pyrrolidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3,3-Difluoro-pentyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3,3-Difluoro-pentyl)-l-(2-hydroxy-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l- " (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3,3-Difluoro-propyl)-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4,4-Difluoro-butyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(5,5-Difluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(5-Fluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4-Fluoro-butyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 -Ethyl-3 -hydroxy-pentyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4, 5- 1rihydroxy-6-memylsulfanyl-tetr-ahydro-pyran-2-yl)-propyl]-amide; 4-Butoxy-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-Pentyloxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4-Fluoro-butoxy)-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5-τrihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Butyl-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-allyl]-amide; l,4-Diethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Fluoro-propoxy)-piperidine-2-carboxy lie acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 ,3 ,3 -Trifluoro-propoxy)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Isobutyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-Propyl-piperidine-2-carboxylic acid [2,2-difluoro- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-propyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Butyl-4-fluoro-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-methoxyethoxy)-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Butyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4,4-Difluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Fluoro-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-anιide; 4-Fluoro-4-propyl-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Fluoroethoxy)-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclobutylmethyl-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 3 -Cyclopropylmethyl-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4, 5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Propyl-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-l-(2-hydroxy-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Pentyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Methyl-butyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Cyclobutyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(2-Cyclobutyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(2-Cyclopropyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Cyclopropyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3 -Butyl- 1 -methyl-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclopropyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 5-Propyl-azepane-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Cyclopentyl-propyl)-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihy droxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Methoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 -Ethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 -Propoxy-propyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Cyclopropylmethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylstιlfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(2-Fluoro-ethoxy)-propyl]-piperidine-2-carboxylic acid [2-methyl- 1 -(3, 4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(3-Fluoro-propoxy)-propyl]-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4-Methoxy-butyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Propoxymethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Fluoro-propoxymethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclohexylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Propyloxyethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethoxy-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-butyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-ethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3 -fluoropropyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4, 5 -trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3,3-difluoropropyl)-piperidine-2-carboxylic acid [2-methyl-l -(3^4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(2,2-difluoroethoxymethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; and prodrugs, tautomers and pharmaceutically acceptable salts thereof. [0026] Specific pro-drug compounds within the scope of this invention include the following compounds: 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid 9H-fluoren-9-ylmethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid ethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid phenyl ester; Phosphoric acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine-2-carbonyl)- amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester; Succinic acid mono-(4,5-dihydroxy-6-{2-methyl-l -[(4-propyl-piperidine-2-carbonyl)- amino] -propyl } -2-methylsulfanyl-tetrahydro-pyran-3 -yl) ester; N-(2-Mo holin-4-yl-ethyl)-succinamic acid 4,5-dihydroxy-6- {2-methyl- 1 - [(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; Dimethylamino-acetic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine-2- carbonyl)-amino]-propyl}-2-meth3 sulfanyl-tetrahydro-pyran-3-yl ester; 1 -(5-Methyl-2-oxo-[l ,3]dioxol-4-ylmethyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-frihydroxy-6-memylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 2- [2-Methyl- 1 -(3 ,4,5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1 -carboxylic acid 5-methyl-2-oxo-[l ,3]dioxol-4-ylmethyl ester; Hexadecanoic acid 4,5 -dihydroxy-6- { 2-methyl- 1 - [(4-propyl-piperidine-2-carbonyl)- amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; 1 -( 1 -Methyl-3 -oxo-but- 1 -enyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trmydroxy-6-memylsulf-anyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2- [2-Methyl- 1 -(3 ,4,5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1-carboxylic acid 1-acetoxy-ethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarb-amoyl]-4-propyl-piρeridine-l-carboxylic acid 2-ammo-3-methyl-pentanoyloxymethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid piperidine-4-carbonyloxymethyl ester; l-(Propionylamino-methyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; N-{2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidin-l-ylmethyl}-nicotinamide; l-(2-Amino-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- > trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -phenyl-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -methyl-pentanoyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; l-(2-Amino-3-methyl-butyryl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(1 -Methyl- 1 ,4-dihydro-pyridine-3-carbonyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid 1 -methyl- 1 ,2,3 ,6-tetrahydro-pyridin-4- yl ester; and tautomers and pharmaceutically acceptable salts thereof. [0027] In one embodiment is a compound selected from the group consisting of: 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclobutylmethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 - trihydroxy-6-me ylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Cyclopropylmethyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Propyl-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-l-(2-hydroxy-ethyl)-azetidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Pentyl-azetidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Methyl-butyl)-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3 -(3 -Cyclobutyl-propyl)-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 - trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3 -(2-Cyclobutyl-ethyl)-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 - trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(2-Cyclopropyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Cyclopropyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-l-methyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; Phosphoric acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester; Succinic acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine- 2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester; N-(2-Morpholin-4-yl-ethyl)-succinamic acid 4,5-dihydroxy-6-{2-methyl- l-[(4-propyl-piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro- pyran-3-yl ester; Dimethylamino-acetic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; 4-Cyclopropylmethyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclopropyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 5-Propyl-azepane-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Cyclopentyl-propyl)-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroχy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Methoxy-propyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3, 4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Ethoxy-propyl)-piperidine-2-carboxy lie acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Propoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-memylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-a.tnide; 4-(3 -Cyclopropylmethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl- l-(3,4,5-trihydiOxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(2-Fluoro-ethoxy)-propyl]-piperidine-2-carboxylic acid [2-methyl-l - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(3-Fluoro-propoxy)-propyl]-piperidine-2-carboxylic acid [2-methyl- l-(3,4,5-frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4-Methoxy-butyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Propoxymethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3, 4,5- 1ιihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3 -Fluoro-propoxymethyl)-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(5-Methyl-2-oxo- [ 1 ,3]dioxol-4-ylmethyl)-4-propyl-piperidme-2- carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2- yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piρeridine-l-carboxylic acid 5-methyl-2-oxo-[l,3]dioxol- 4-ylmethyl ester; Hexadecanoic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine-2- carbonyl)-amino] -propyl } -2-methylsulfanyl-tetrahydro-pyran-3 -yl ester; 1 -( 1 -Methyl-3 -oxo-but- 1 -enyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihy(-UOxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid 1-acetoxy-ethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid 2-amino-3 -methyl- pentanoyloxymethyl ester; 2- [2-Methyl- 1 -(3 ,4,5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid piperidine-4- carbonyloxymethyl ester; 1 -(Propionylamino-methyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl- 1 -(3 ,4, 5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide; N-{2-[2-Memyl-l-(3,4,5-frihydroxy-6-methylsulf-myl-tetrahydro-pyran-2- yl)-propylcarbamoyl] -4-propyl-piperidin- 1 -ylmethyl } -nicotinamide; 1 -(2-Amino-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -phenyl-propionyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-aιnide; l-(2-Amino-3-methyl-pentanoyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -methyl-butyryl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -( 1 -Methyl- 1 ,4-dihydro-pyridine-3-carbonyl)-4-propyl-piperidine-2- carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2- yl)-propyl]-amide; 2-[2-Memyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid 1 -methyl- 1 ,2,3 ,6-tetrahydro- pyridin-4-yl ester; 4-Cyclohexylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Propyloxyethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethoxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-butyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-ethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3-fluoropropyl)-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tetralιydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3,3-difluoropropyl)-piperidine-2 -carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; and 4-Fluoro-4-(2,2-difluoroethoxymethyl)-piperidine-2-carboxylic acid [2- methyl- 1 -(3 ,4,5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide; or a prodrug and/or a pharmaceutically acceptable salt thereof. [0020] The compounds, tautomers, prodrugs and pharmaceutically acceptable salts thereof, as defined herein, may have activity against bacteria, protozoa, fungi, and parasites. [0029] In another aspect, this invention provides pharmaceutical compositions comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound defined herein. The pharmaceutical compositions of the present invention may further comprise one or more additional antibacterial agents. One or more of the antibacterial agents may be active against gram negative bacteria. One or more of the antibacterial agents may be active against gram positive bacteria.
[0030] In one of its method aspects, this invention is directed to a method for tlie treatment of a microbial infection in a mammal comprising administering to the mammal a therapeutically effective amount of a compound of this invention. The compound of this invention may be administered to the mammal orally, parenterally, transdermally, topically, rectally, or intranasaUy.
[0031] In another of its method aspects, this invention is directed to a method for the treatment of a microbial infection in a mammal comprising administering to the mammal a pharmaceutical composition comprising a therapeutically effective amount of a compound of this invention. The pharmaceutical compositions of the present invention may further comprise one or more additional antibacterial agents, one of which may be active against gram negative bacteria and/or one of which may be active against gram positive bacteria. The pharmaceutical composition may be administered to the mammal orally, parenterally, transdermally, topically, rectally, or intranasaUy.
[0032] In a preferred embodiment, the microbial infection being treated is a gram positive bacterial infection. In an additional embodiment, the infection may be a gram negative bacterial infection. In a further embodiment, the infection may be a mycobacteria infection, a mycoplasma infection, or a chlamydia infection.
[0033] In yet another aspect, the present invention provides novel intermediates and processes for preparing' compounds of formula (I), (II), (III) and (IN).
DETAILED DESCRIPTION OF THE INVENTION [0034] As described above, this invention relates to lincomycin derivatives that exhibit antibacterial activity, in particular gram positive antibacterial activity. However, prior to describing this invention in further detail, the following terms will first be defined.
Definitions
[0035] Unless otherwise stated, the following terms used in the specification and claims have the meanings given below.
[0036] "Acyl" means the group -C(O)R' wherein R' is alkyl, alkenyl, alkynyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
[0037] "Acyloxy" means the group -C(O)OR', wherein R' is alkyl, alkenyl, alkynyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
[0038] "Alkenyl" means a linear unsaturated monovalent hydrocarbon radical of two to eight carbon atoms or a branched monovalent hydrocarbon radical of three to eight carbon atoms containing at least one double bond, (-C=C-). Examples of alkenyl groups include, but are not limited to, allyl, vinyl, 2-butenyl, and the like. [0039] "Alkoxy" refers to the group "alkyl-O-" which includes, by way of example, methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n- hexoxy, 1,2-dimethylbutoxy, and the like.
[0040] "Alkoxyalkoxy" refers to the group alkyl-O-alkylene-O-, wherein alkyl is as defined herein.
[0041] "Alkyl" means a linear saturated monovalent hydrocarbon radical of one to eight carbon atoms or a branched saturated monovalent hydrocarbon radical of three to eight carbon atoms. Examples of alkyl groups include, but are not limited to, groups such as methyl, ethyl, n- propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, and the like.
[0042] "Alkylene" means a linear divalent hydrocarbon radical of one to eight carbon atoms or a branched divalent hydrocarbon group of three to eight carbon atoms. Examples of alkylene groups include, but are not limited to, methylene, ethylene, 2-methylpropylene, and the like.
[0043] "Alkylthio" refers to the group "alkyl-S-" which includes, by way of example, methylthio, butylthio, and the like.
[0044] "Alkynyl" means a linear monovalent hydrocarbon radical of two to eight carbon atoms or a branched monovalent hydrocarbon radical of three to eight carbon atoms containing at least one triple bond, (-C≡C-). Examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, 2-butynyl, and the like.
[0045] "Amino" or "substituted nitrogen" refers to the group "-NRaRb" wherein Ra and Rb are independently H, alkyl, haloalkyl, alkenyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl.
[0046] "Aminocarboxyalkyl" means a group "-RcC(O)NRaRb" where Rc is an alkylene, as defined above, and Ra and Rb are as defined above.
[0047] "Aryl" means a monovalent monocyclic or bicyclic aromatic carbocyclic group of six to fourteen ring atoms. Examples include, but are not limited to, phenyl, naphthyl, and anthryl.
The aryl ring may be optionally fused to a 5-, 6-, or 7-membered monocyclic non-aromatic ring optionally containing 1 or 2 heteroatoms independently selected from oxygen, nitrogen, or sulfur, the remaining ring atoms being C where one or two C atoms are optionally replaced by a carbonyl. Representative aryl groups with fused rings include, but are not limited to, 2,5- dihydro-benzo[b]oxepinyl, 2,3-dihydrobenzo[l,4]dioxanyl, chromanyl, isochromanyl, 2,3- dihydrobenzofuranyl, 1,3-dihydroisobenzofuranyl, benzo[l,3]dioxolyl, 1,2,3,4- tetrahydroisoquinolinyl, 1,2,3,4-tetrahydroquinolinyl, 2,3-dihydro-lH-indolyl, 2,3dihydro-lH- isoindolyl, benzimidazole-2-onyl, 2-H-benzoxazol-2-onyl, and the like. [0048] "Carbonyl" means the group "C(O)."
[0049] "Carboxy" means the group "C(O)O."
[0050] "Cyanoalkyl" refers to an alkyl, wherein alkyl is as defined above, substituted with one or more cyano (-CN) groups provided that if two cyano groups are present they are not both on the same carbon atom. Examples of cyanoalkyl groups include, for example, cyanomethyl,
2-cyanoethyl, 2-cyanopropyl, and the like.
[0051] "Cycloalkyl" refers to cyclic alkyl groups of from 3 to 20 carbon atoms having a single or multiple cyclic rings including, by way of example, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, adamantanyl, and the like. Cycloalkyl groups of the present invention also include fused multicyclic rings wherein one or more of the rings within the multicyclic ring system are aromatic, as long as the point of attachment to the core or backbone of the structure is on the non-aromatic ring, e.g., fluorenyl.
[0052] "Cycloalkylalkyl" means a group -RcR-j where Rc, is an alkylene group and Rd is a cycloalkyl group, as defined above. Examples include, but are not limited to, cyclopropylmethylene, cyclohexylethylene, and the like.
[0053] "Halo" or "Halogen" means fluoro, chloro, bromo, or iodo.
[0054] "Haloalkyl" means an alkyl, wherein alkyl is as defined above, substituted with one or more, preferably one to 6, of the same or different halo atoms. Examples of haloalkyl groups include, for example, trifluoromethyl, 3-fluoropropyl, 2,2-dichloroethyl, and the like.
[0055] "Heteroaryl" means a monovalent monocyclic or bicyclic aromatic radical of 5 to 10 ring atoms containing one, two, or three ring heteroatoms selected from N, O, or S, the remaining ring atoms being C. Representative examples include, but are not limited to, thienyl, benzothienyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinolinyl, quinoxalinyl, imidazolyl, furanyl, benzofuranyl, thiazolyl, isoxazolyl, benzisoxazolyl, benzimidazolyl, triazolyl, pyrazolyl, pyrrolyl, indolyl, 2-pyridonyl, 4-pyridonyl, N-alkyl-2-pyridonyl, pyrazinonyl, pyridazinonyl, pyrimidinonyl, oxazolonyl, and the like.
[0056] "Heterocycle" or "heterocyclic" refers to a saturated or unsaturated group having a single ring or multiple condensed rings, from 1 to 10 carbon atoms and from 1 to 4 heteroatoms selected from the group consisting of nitrogen, sulfur, or oxygen within the ring, wherein, in fused ring systems one or more of the rings can be aryl or heteroaryl as defined herein.
Examples of heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine, imidazolidine, imidazoline, piperidine, piperazine, indoline, phthalimide, 1,2,3,4- tetrahydro-isoquinoline, 4,5,6,7-tetrahydrobenzo[b]thiophene, thiazole, thiazolidine, thiophene, benzo [b]thiophene, morpholinyl, thiomorpholinyl (also referred to as thiamorpholinyl), piperidinyl, pyrrolidine, tetrahydrofuranyl, and the like.
[0057] Heterocycles may be optionally substituted with from one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxyl, carboxy, cyano, oxo, nitro, and alkylthio as these terms are defined herein.
[0058] "Hydroxy" or "hydroxyl" means the group -OH.
[0059] "Hydroxyalkyl" refers to an alkyl, wherein alkyl is as defined above substituted with one or more -OH groups provided that if two hydroxy groups are present they are not both on the same carbon atom. Examples of hydroxyalkyl groups include, for example, hydroxymethyl,
2-hydroxyethyl, 2-hydroxypropyl, and the like.
[0060] "Mammal" refers to all mammals including humans, livestock, and companion animals. - <
[0061] "Optional" or "optionally" means that the subsequently described event or circumstance may, but need not, occur, and that the description includes instances where the event or circumstance occurs and instances in which it does not. For example, "aryl group optionally mono- or di- substituted with an alkyl group" means that the alkyl may but need not be present, and the description includes situations where the aryl group is mono- or disubstituted with an alkyl group and situations where the aryl group is not substituted with the alkyl group.
[0062] "Pharmaceutically acceptable carrier" means a carrier that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic and neither biologically nor otherwise undesirable, and includes a carrier that is acceptable for veterinary use as well as human pharmaceutical use. "A pharmaceutically acceptable carrier" as used in the specification and claims includes both one and more than one such carrier.
[0063] "Pharmaceutically acceptable salt" of a compound means a salt that is pharmaceutically acceptable and that possesses the desired pharmacological activity of the parent compound. Such salts include, but are not limited to, (1) acid addition salts, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; or formed with organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2- hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2- naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4- methylbicyclo[2.2.2]oct-2-ene-l-carboxylic acid, glucoheptonic acid, 4,4'methylenebis-(3-hydroxy-2-ene-l-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid, and the like; or (2) salts formed when an acidic proton present in the parent compound either is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth metal ion, or an aluminum ion; or coordinates with an organic base such as ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like. [0064] "Prodrugs" mean any compound which releases an active parent drug according to a compound of the subject invention in.vivo when such prodrug is administered to a mammalian subject. Prodrugs of a compound of the subject invention are prepared by modifying functional groups present in a compound of the subject invention in such a way that the modifications may be cleaved in vivo to release the parent compound. Prodrugs include compounds of the subject invention wherein a hydroxy, sulfhydryl or amino group in the compound is bonded to any group that may be cleaved in vivo to regenerate the free hydroxyl, amino, or sulfhydryl group, respectively. Examples of prodrugs include, but are not limited to esters (e.g., acetate, formate, and benzoate derivatives), carbamates (e.g., N,N-dimethylaminocarbonyl) of hydroxy functional groups in compounds of the subject invention, and the like. Specific examples include -C(O)O-alkylene-cycloalkyl, -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O]p-alkylene-heterocycle, -[C(O)O]p-alkylene-substituted heterocycle, wherein p is 0 or 1. In one embodiment, the -C(O)O-substituted alkyl does not include the following:
[0065] "Substituted alkyl" means an alkyl group, as defined above, in which one or more of the hydrogen atoms has been replaced by a halogen (i.e., CI, Br, F, or I), oxygen, hydroxy, amine (primary), amine (secondary-amine substituted by alkyl above), amine (tertiary-amine substituted by alkyl as above), sulfur, -SH, phenyl, substituted phenyl, cycloalkyl, alkoxy, substituted alkoxy. Examples of substituted alkyl groups include, but are not limited to, 1- fluoroethyl, 1-chloroethyl, 2-fluoroethyl, 2-chloroethyl, 1-bromopropyl, 2-iodopropyl, 1- chlorobutyl, 4-flurobutyl, 4-chlorobutyl, 2-cyclopropyl-ethyl, 3-cyclobutyl-propyl, 4- cyclopentyl-butyl, and 4-cyclohexyl-butyl.
[0066] "Substituted alkoxy" means substituted alkyl-O-, wherein substituted alkyl is as defined herein.
[0067] "Substituted aryl" means an aryl ring substituted with one or more substituents, preferably one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxy, carboxy, cyano, nitro, alkylthio, and thioalkyl. The aryl ring may be optionally fused to a 5-, 6-, or 7-membered monocyclic non-aromatic ring optionally containing 1 or 2 heteroatoms independently selected from oxygen, nitrogen, or sulfur, the remaining ring atoms being carbon where one or two carbon atoms are optionally replaced by a carbonyl.
[0063] "Substituted cycloalkyl" means a cycloalkyl substituted with an alkyl group, wherein alkyl is as defined above or a group as defined above for substituted alkyl. [0069] "Substituted heteroaryl" means a heteroaryl ring, wherein heteroaryl is as defined above, substituted with one or more substituents, preferably one to three substituents selected from the group consisting of alkyl, alkenyl, alkynyl, halo, alkoxy, acyloxy, amino, hydroxy, carboxy, cyano, nitro, alkylthio, and thioalkyl, wherein said substituents are as defined herein. [0070] "Substituted oxygen" refers to the group "-O-Rd" wherein Rd is alkyl, haloalkyl, alkenyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, or substituted heteroaryl, wherein said substituents are as defined herein. [0071] "Substituted phenyl" means a phenyl ring wherein one or more of the hydrogen atoms has been replaced by a halogen, hydroxy, alkyl, amine (primary, secondary, and tertiary with the latter two alkyl substituted), -SH, and phenyl. Representative examples include, but are not limited to, p-bromophenyl, m-iodophenyl, o-chlorophenyl, p-ethylphenyl, m-propylphenyl, o-methylphenyl, and p-octylphenyl.
[0072] "Thioalkyl" refers to an alkyl, wherein alkyl is as defined above, substituted with one or more -SH groups provided that if two hydroxy groups are present they are not both on the same carbon atom. Examples of thioalkyl groups include, for example, thiomethyl, 2-thioethyl, 2-thiopropyl, and the like.
[0073] "Therapeutically effective amount" means the amount of a compound or composition that, when administered to a mammal for treating a disease, is sufficient to effect such treatment for the disease. The "therapeutically effective amount" will vary depending on the compound or composition, the disease and its severity and the age, weight, etc., of the mammal to be treated. [0074] "Treating" or "treatment" of a disease includes: (1) preventing the disease, i.e. causing the clinical symptoms of the disease not to develop in a mammal that may be exposed to or predisposed to the disease but does not yet experience or display symptoms of the disease, (2) inhibiting the disease, i.e., arresting or reducing the development of the disease or its clinical symptoms, or (3) relieving the disease, i.e., causing regression of the disease or its clinical symptoms. "Tautomer" refers to an isomer in which migration of a hydrogen atom results in two or more structures.
[0075] The compounds of the present invention are generally named according to the IUPAC or CAS nomenclature system. Abbreviations that are well known to one of ordinary skill in the art may be used (e.g. "Ph" for phenyl, "Me" for methyl, "Et" for ethyl, "Bn" for benzyl, "h" for hour and "rt" for room temperature).
General Synthetic Schemes
[0076] Compounds of this invention can be made by the methods depicted in the reaction schemes shown below.
[0077] The starting materials and reagents used in preparing these compounds are either available from commercial suppliers such as Toronto Research Chemicals (North York, ON Canada), Aldrich Chemical Co. (Milwaukee, Wisconsin, USA), Bachem (Torrance, California, USA), Emka-Chemie, or Sigma (St. Louis, Missouri, USA) or are prepared by methods known to those skilled in the art following procedures set forth in references such as Fieser and Fieser' s Reagents for Organic Synthesis, Volumes 1-15 (John Wiley and Sons, 1991), Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplemental (Elsevier Science Publishers, 1989), Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), March's Advanced Organic Chemistry, (John Wiley and Sons, 4th Edition), and Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989). These schemes are merely illustrative of some methods by which the compounds of this invention can be synthesized, and various modifications to these schemes can be made and will be suggested to one skilled in the art having referred to this disclosure.
[0078] As it will be apparent to those skilled in the art, conventional protecting groups may be necessary to prevent certain functional groups from undergoing undesired reactions. Suitable protecting groups for various functional groups, as well as suitable conditions for protecting and deprotecting particular function groups are well known in the art. For example, numerous protecting groups are described in T.W. Greene and G.M. Wuts, Protecting Groups in Organic Synthesis, Second Edition, Wiley, New York, 1991, and references cited therein. [0079] The starting materials and the intermediates of the reaction may be isolated and purified if desired using conventional techniques, including but not limited to filtration, distillation, crystallization, chromatography, and the like. Such materials may be characterized using conventional means, including physical constants and spectral data. [0080] The compounds of this invention will typically contain one or more chiral centers. Accordingly, if desired, such compounds can be prepared or isolated as pure stereoisomers. All such stereoisomers (and enriched mixtures) are included within the scope of this invention, unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents well-known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chiral column chromatography, chiral resolving agents, and the like.
Preparation of Compounds of formula (I)
[0081] In general, to prepare the compounds of formula (I) of the present invention, an appropriately 7-substititued lincosamine intermediate and an appropriately substituted pyrrolidinyl or piperidyl carboxylic acid are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base. This reaction can be performed with any number of known coupling reagents, such as O-(7-azabenzotriazol-l- yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), 1-hydroxybenzotriazole hydrate (HOBT) with carbodiimides, isobutyl chloroformate, diphenylphosphoryl azide (DPP A), and the like. Suitable organic bases include diisopropylethylamine (DIEA), triethylamine (TEA), pyridine, N-methyl morpholine, and the like. Suitable inert organic solvents which can be used include, for example, N,N-dimethylformamide, acetonitrile, dichloromethane, and the like. This reaction is typically conducted using an excess of carboxylic acid to lincosamine at temperatures in the range of about 0°C to about 50°C. The reaction is continued until completion, which typically occurs in from about 2 to 12 h.
[0082] Appropriately 7-substititued lincosamine intermediates, as defined in the present invention (i.e., R2/R3), are synthesized by methods well known to those of skill in the art from methyl 6-amino-6,8-dideoxy-l-mio-e-rythro-β-D-galacto-octopyranoside, which can be prepared as described by Hoeksema, et al., Journal of the American Chemical Society, 1967, 89 2448- 2452. Illustrative syntheses for 7-substituted lincosamine intermediates are shown below in Schemes 1-5.
[0083] Appropriately substituted pyrrolidinyl or piperidyl carboxylic acid intermediates, as defined in the present invention (i.e., R9), are also synthesized by methods well known to those of skill in the art from prolines and pyridines. The prolines and pyridines that can be used in the synthesis of the carboxylic acid intermediates of the present invention include, for example, 4- oxoproline and 4-substituted pyridines. The prolines and pyridines used in the synthesis are commercially available from vendors such as Aldrich and Sigma. Alternatively, these prolines and pyridines can be prepared by methods well known in the art. Illustrative syntheses for appropriately substituted pyrrolidinyl or piperidyl carboxylic acid intermediates are shown below in Schemes 6-10.
[0084] Scheme 1 below illustrates a general synthesis of a lincosamine intermediate lc wherein P is an N-protecting group, preferably either Cbz or Boc, and R1 is as defined for formula (I).
Scheme 1. General synthesis of lincosamine intermediate lc. (a) N-Protection (Boc, Cbz); (b) O-silyl protection (TMS); (c) Swern oxidation. [0085] As shown in Scheme 1 , methyl 6-amino-6,8-dideoxy- 1 -thio-erythro-β-D-galacto- octopyranoside, la, is prepared as described by Hoeksema, et al, Journal of the -American Chemical Society, 1967, 89 2448-2452. The amino functional group and the hydroxy functional groups of the product la are then protected with suitable protecting groups. Suitable N- protecting groups can be formed by the addition of di-t-butyldicarbonate, N- (benzyloxycarbonyloxy) succinimide, and the like. The hydroxy groups can be protected as silyl ethers. The hydroxyl group can be converted to trimethylsilyl (TMS) ethers by reaction with N, O-bis-(trimethylsilyl)trifluoroacetamide in the presence of an appropriate organic base such as triethylamine (TEA) or trimethylsilyl chloride in the presence of an organic base such as triethylamine. The N-protection is typically accomplished before the O-protection. Chromatography of the crude product on silica after evaporation of the solvent provides the protected product lb.
[0086] The 7-O-trimethylsilyl group of lb is chemoselectively deprotected and oxidized to provide the 7-keto-lincosamine derivative lc. This selective transformation is performed by addition of the protected product lb to dimethylsulfoxide and oxalyl chloride in an infert organic solvent such as dichloromethane followed by an appropriate organic base such as triethylamine. Alternatively, the transformation may be performed by addition of lb to dimethyl sulfoxide and an appropriate activating agent such as trifluoroacetic anhydride in an inert organic solvent. The reaction is typically conducted at temperatures in the range of approximately -70°C to 80°C. The resulting reaction mixture is stirred at the low temperature and is then allowed to warm to approximately -50°C. The reaction is maintained at this second temperature for approximately 1 h to 3 h. To the reaction mixture is added a suitable organic base, such as TEA, pyridine, and the like. The reaction mixture is appropriately worked up to provide the product le. The general class of conditions used in the transformation of lb to lc is known in the art as Swern oxidation conditions.
[0087] Scheme 2 below illustrates a general synthesis of a lincosamine intermediate 2b wherein P is an N-protecting group, preferably either Cbz or Boc, R1 is as defined for formula (I), and one of R2 and R3 is hydrogen and the other is as defined for formula (I).
Scheme 2. General synthesis of lincosamine intermediate 2b.
(a) Wittig olefination (R2PFh3 +X\ R2PO(OEt)2, base, solvent); (b) and (c) H2/Pd, Global de- protection
[0088] As shown in Scheme 2, a keto-lincosamine intermediate lc is reacted to form an alkene using the Wittig or Homer- Wadsworth-Emmons reaction. In this reaction, a suitable phosphonium salt or phosphonate is deprotonated using a strong base to form a phosphorus ylide. Suitable phosphonium salts which can be used are alkyltriphenylphosphonium halides, which can be prepared by the reaction of triphenylphosphine and an alkyl halide. Suitable phosphorous compounds include, for example, methyltriphenylphosphonium bromide, diethyl(cyanomethyl)phosphonate and the like. Suitable strong bases which can be used to form the ylide include organolithium reagents, potassium tert-butoxide, and the like. The formation of the phosphorus ylide is typically conducted under an inert atmosphere, such as N2, in an inert organic solvent such as toluene, THF, and the like, at low temperatures.
[0089] After formation of the phosphorus ylide, the product lc is added to the reaction. The reaction conveniently can be performed at temperatures between -40°C and room temperature and is stirred until completion, typically 1 to 4 h. The resulting organic solution is worked-up and chromatography of the crude product on silica provides the alkene product 2a. [0090] Optionally, the product of 2a may be purified using conventional techniques, such as chromatography and said purified product may be used in the subsequent coupling reaction to yield vinyl lincosamine derivatives of the present invention.
[0091] The product 2a is then hydrogenated to provide the saturated product 2b. The hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle. The bottle is purged, and charged with H2 to approximately 50 to 70 psi and shaken until completion, typically approximately 12 to 24 h. The resulting reaction mixture is filtered, e.g., through celite, and rinsed with a polar organic solvent such as methanol. The organic solution is worked up by transferring to a resin funnel containing dry, washed Dowex 50w-400x H*" form and shaken. After washing the resin with methanol and water, the product 2b is eluted from the resin by washing with 5% TEA in MeOH. The product can also be purified by silica gel column chromatography. [0092] Scheme 3 illustrates a general synthesis of a lincosamine intermediate 3b wherein P is an N-protecting group, preferably either Cbz or Boc, R1 is as defined for formula (I), and one of R2 or R3 is alkyl and the other is -OH.
Scheme 3. General synthesis of lincosamine intermediate 3b.
(a) R2M (carbon nucleophile); (b) (i) TMS de-protection (H* or F-) and (ii) N-deprotection [0093] As shown in Scheme 3, suitable carbon nucleophiles add to 7-ketolincosamine intermediate lc in suitable inert organic solvents to provide a 7-hydroxy lincosamine intermediate 3b. Suitable carbon nucleophiles include methylmagnesium chloride, diethyl zinc, sodium acetylide and the like and suitable inert organic solvents which can be used include THF, diethyl ether, toluene, and the like. The reaction is typically conducted at reduced temperatures, approximately at 0°C, for about 3 to 5 h. The reaction is then quenched with a saturated aqueous acidic solution, such as saturated aqueous The quenched mixture is then worked up and can be purified by chromatography to provide the product 3b. [0094] Scheme 4 below illustrates a general synthesis of a lincosamine intermediate 4b wherein P is a N-protecting group, preferably Boc, R is as defined for formula (I), and R R is an oxime (=NOR7), wherein R7 is as defined for formula (I).
Scheme 4. General synthesis of 7-oxime-lincosamines 4b.
[0095] As shown in Scheme 4, the lincosamine intermediate lc is converted to the oxime by stirring in the presence of a suitable reagent such as O-trimethylsilylhydroxylamine, O- alkylhydroxylamine hydrochloride (for example, O-methylhydroxylamine hydrochloride), and the like. The reaction is typically conducted in a polar organic solvent such as methanol. The reaction conveniently can be conducted at rt in approximately 8 to 24 h. The solvent is removed to provide the N-protected product 4a.
[0096] Removal of the protecting group can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like. The removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product 4b.
[0097] Scheme 5 below illustrates a general synthesis of a lincosamine intermediate 5b wherein R2 and R3 are both fluorine, P is an N-protecting group, preferably Cbz or Boc, and R1 is as defined for formula (I).
Scheme 5. General synthesis of 7-deoxy-7,7-difluorolincosamines 5b.
(a) F "; (b) Ac2O, pyridine, DMAP; (c) DAST; (d) TFA
[0098] As shown in Scheme 5, the lincosamine intermediate lc is contacted with a suitable fluoride in an inert organic solvent. Suitable fluorides which can be used include tetrabutylammonium fluoride, Amberlite resin A-26 F form, HF •pyridine and the like. Suitable inert organic solvents include THF, acetonitrile, dichloromethane, dioxane, and the like. The reaction conveniently can be conducted at rt in about 1 to 2 h. The product (not shown) can be purified on a silica gel column.
[0099] The O-protecting groups on the product obtained from the column are converted by contact with acetic anhydride and dimethylaminopyridine (DMAP) in a suitable mixture of an inert organic solvent and an organic base, such as, for example, dichloromethane and pyridine. The reaction conveniently can be conducted at rt in approximately 6 to 12 h. The product can be purified on silica gel column to provide product 5a.
[00100] The product 5a is contacted with a suitable fluorinating reagent and then the N- protecting group is removed to provide the product 5b. Suitable fluorinating reagents which can be used include, for example, dimethylaminosmfurtrifluoride, [bis(2-me oxyethyl)amino]sul---urtrifluoride, and the like. The reaction is typically conducted in an inert organic solvent such as dichloromethane, ethylacetate, THF, and the like at room temperature in approximately 6 to 12 h.
[0100] Removal of the protecting group can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like. The removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product 5b.
[0101] Scheme 6 below illustrates a general synthesis of a proline intermediate 6c wherein
R9 is as defined for formula (I).
Scheme 6. General synthesis of cis/trans R -proline intermediate mixtures 6c.
(a) R9CH2Br+Ph3P , NaH, DMSO; (b) H2/Pt
[0102] As shown in Scheme 6, the product 6c is prepared as described in Birkenmeyer, et al., Journal of Medicinal Chemistry 1972, 15, 1255-1259. Compound 6a is commercially available from vendors such as RSP (Scientific Research Consortium, Inc.). Alternatively, 6a can be prepared from commercially available protected hydroxy prolines by methods well known in the art. See, e.g., Demange, et al., Tetrahedron Letters 1998, 39,1169-1172.
[0103] Scheme 7 below illustrates a general synthesis of trans-R9-proline intermediates 7d, wherein R9 is alkyl or substituted alkyl.
0 V∞2Bn UI . 0Jr ∞2Bn b.l,ιι -.C02Bn I I I Boc Boc Boo 7a 7b 7c
Scheme 7. General synthesis of trans-alkylprolines 7d.
(a) (i) LiHMDS, THF -78°C, (ii) bromoalkene; (b) (i) LiBHEt3, THF -78°C, (ii) BF3OEt2,
Et3SiH; (c) H2 Pd/C. [0104] As shown in Scheme 7, a protected 4-oxoproline, 7a, is enolated with a suitable enolation agent and then alkylated with a suitable alkylating agent in an inert organic solvent to provide a lactam 7b (wherein R9 is alkenyl), as described in the literature procedure by Zhang, et al., J.A.C.S. 1998, 120 3894-3902. Compound 7a is commercially available from vendors such as Bachem. Alternatively, 7a can be prepared by methods well known in the art. Suitable enolating agents include LiHMDS, LiN(iPr)2, and the like, and suitable alkylating agents include allylic and benzylic bromides, for example, 4-bromo-2-methyl-2-butene and cis-l-bromo-2- pentene, allylbromide, and the like.
[0105] The lactam 7b is reduced using a suitable reducing agent to provide a pyrrolidine 7c, wherein R9 is alkenyl. The reduction is preformed by a two-step sequence involving superhydride reduction of the lactam to the hemiaminal and the subsequent reduction of the hemiaminal. Suitable reducing agents which can be used include Et3SiH/BF3 »OEt2, Et3SiH/TiCl4, and the like.
[0106] The pyrrolidine 7c is then hydrogenated to simultaneously remove the unsaturation in the R substituent and remove the benzyl protecting group from the carboxylic acid to provide the product 7 . The hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle. The bottle is; purged, and charged with H2 to approximately 50 to 70 psi and shaken until completion, typically approximately 5 to 24 h. The reaction mixture is filtered, e.g., through a celite pad, and washed with a polar organic solvent, such as methanol. Evaporation of the combined washings and filtrate affords the product 7d, wherein R9 is an alkyl or substituted alkyl. [0107] Scheme 8 below illustrates a general synthesis of trans-R9-proline intermediates 8c, wherein R9 is alkyl or substituted alkyl.
Scheme 8. General synthesis of trans-R9-substituted prolines 8c, wherein R9 is alkyl or substituted alkyl. (a) O3, DCM, -78°C, DMS; (b) P+Ph3 salt, Base; (c) H2, Pd/C.
[0108] As shown in Scheme 8, the product 7d is ozonolized to provide the aldehyde 8a. The ozonolysis reaction is typically conducted in an anhydrous inert organic solvent, such as dichloromethane, dioxane, THF, and the like, at low temperatures, e.g., -78°C followed by quenching of the reaction with a reducing agent such as DMS, Ph3P. [0109] The aldehyde, 8a, is reacted with a suitable phosphonium salt in the presence of a strong base in an inert organic solvent. Suitable phosphonium salts which can be used include, for example, fluorobenzyl phosphonium chloride, 4-chlorobenzyl phosphonium chloride, dibromofluoromethane and triphenylphosphine, and the like. Suitable bases which can be used include potassium t-butoxide, organolithium reagents, and activated zinc. Suitable organic solvents which can be used include toluene, THF, dimethylacetamide, and the like. The reaction is typically conducted in an inert atmosphere, such as under nitrogen, with vigorous stirring. The reaction is typically conducted at rt to approximately 110°C for 1 to 2 h. The resulting reaction mixture is appropriately worked-up and can be purified by chromatography to provide 8b (wherein R9 is alkenyl).
[0110] The product 8b is then hydrogenated to provide the product 8c. The hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% Palladium on carbon in a Parr bottle. The bottle is purged, and charged with H to approximately 40 to 70 psi and shaken until completion, typically approximately 4 to 24 h. The reaction mixture is filtered, e.g., through a celite pad and washed several times with a polar organic solvent, such as methanol. Evaporation of the combined washings and filtrate affords the product 8c, wherein R9 is an alkyl or substituted alkyl.
[0111] Scheme 9 below illustrates a general synthesis of trans-R9-proline intermediates 9d, wherein R9 is substituted alkyl wherein X is halo.
Scheme 9. Example synthesis of trans-halosubstituted alkyl prolines 9d. (a) Tetraallyltin, BF3 «Et2O; (b) DMSO, (COCl)2, TEA; (c) DAST (d)10% Pd/C, H2 [0112] As shown in Scheme 9, aldehyde, 8a, is reduced and alkylated using a suitable reagent in an inert organic solvent to provide a hydroxyalkenyl substituted proline, 9a. Suitable reagents to reduce and alkylate the aldehyde include tetraallyltin/boron trifluoride etherate, allylTMS/boron trifluoride etherate and suitable inert organic solvents which can be used include THF, dichloromethane, and the like. The reaction is typically conducted at low temperatures, e.g., 0°C, for approximately 1 to 2 h. To the reaction mixture is added a solution of a suitable fluoride salt in water, for example potassium fluoride in water, followed by the addition of methanol. The reaction mixture is filtered, for example, over celite. The product can be purified by chromatography to provide 9a.
[0113] The hydroxyalkenyl substituted proline, 9a, is oxidized to the ketone by contact with a suitable oxidizing agent in an inert organic solvent. Suitable oxidizing agents include oxalyl chloride/DMSO, Dess Martin periodinane, and the like. Suitable inert organic solvents include dichloromethane, and the like. The reaction is typically conducted at reduced temperatures, e.g., -72°C to -50°C, for approximately 30 min to 2 h. To the reaction mixture is added a suitable organic base, such as triethylamine. The reaction mixture is worked up to provide product 9b. [0114] , The keto-substituted product 9b is halogenated by contact with a suitable halogenating agent in an inert organic solvent. Suitable halogenating agents which can be used include diethylaminosulfur trifluoride, [bis(2-methoxyethyl)amino] sulfur trifluoride, and the like. Suitable inert organic solvents which can be used include dichloromethane, ethyl acetate, THF, and the like. The reaction is typically conducted at low temperatures in the range of approximately -30°C to -78°C. The reaction mixture is gradually allowed to warm to rt and stirred at rt until completion, typically in 6 to 12 h. The reaction mixture is worked up and can be purified by chromatography to provide 9c.
[0115] The product 9c is then hydrogenated to provide the product 9d. The hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using 10% palladium on carbon in a Parr bottle. The bottle is purged, and charged with H2 to approximately 40 to 70 psi and shaken until completion, typically approximately 4 to 24 h. The reaction mixture is filtered, e.g., through a celite pad and washed several times with a polar organic solvent, such as methanol. Evaporation of the combined washings and filtrate affords the product 9d. [0116] Scheme 10 below illustrates a general synthesis, as described in Shuman, Journal of Organic Chemistry. 1990, 55, 741-750, of substituted pyridine carboxylic acid intermediates 10b, wherein R9 is as defined for formula (I).
10a 10b
Scheme 10. General synthesis of substituted pyridin-2-yl carboxylic acids 10b. [0117] As shown in Scheme 10, an appropriately substituted pyridine, is contacted with a suitable oxidizing agent in an inert organic solvent. The appropriately substituted pyridine starting materials are commercially available from vendors such as Aldrich and Sigma. Alternatively, these pyridines can be prepared by methods well known in the art. Suitable oxidizing agents which can be used include hydrogen peroxide, MCPBA, and the like. The reaction is typically conducted at reflux for 6 to 12 h. The reaction mixture is then contacted with a suitable cyanide reagent to provide the cyano-substituted pyridine, 10a. Suitable cyanide reagents which can be used include trimethylsilyl cyanide, HCN, and the like. Suitable inert organic solvents include dichloromethane, dioxane, THF, and the like. The reaction conveniently can be conducted at rt in approximately 6 to 12 h. The reaction mixture is worked up to provide the cyano-substituted pyridine, 10a.
[0118] The cyano-substituted pyridine, 10a, is then hydrolyzed to provide the pyridin-2-yl carboxylic acid 10b by contact with a suitable acid. Suitable acids for hydrolyzing the cyano group to the carboxylic acid include hydrochloric acid, aqueous sulfuric acid, and the like. The reaction is typically conducted at reflux in 6 to 12 h.
[0119] Scheme 11 below illustrates the coupling reaction of a lincosamine intermediate, prepared as described above in Schemes 1-5, and a pyrrolidinyl or piperidinyl carboxylic acid, prepared as described above in Schemes 6-10, wherein R1, R2, R3, R6, and R9 are as defined for formula (I) and P1 is a suitable O-protecting group and P2 is a suitable N-protecting group. 11a
Scheme 11. General coupling and deprotection methods.
[0120] As shown in Scheme 11, an appropriately 7-substititued lincosamine intermediate (prepared, for example, according to any one of Schemes 1-5) and an appropriately substituted pyrrolidinyl or piperidyl carboxylic acid (prepared, for example, according to any one of Schemes 6-10) are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base. This reaction can be performed with any number of known coupling reagents, such as O-(7-azabenzotriazol-l-yl)- N,N,N',N'tetramethyluronium hexafluorophosphate (HATU), 1-hydroxybenzotriazole hydrate (HOBT) with carbodiimides, diphenylphosphoryl azide (DPP A), isobutyl chloroformate, and the like. Suitable organic bases include diisopropylethylamine (DIEA), triethylamine (TEA), pyridine, N-methyl morpholine, and the like. Suitable inert organic solvents which can be used include, for example, N,N-dimethylformamide, acetonitrile, dichloromethane, and the like. This reaction is typically conducted using an excess of carboxylic acid to lincosamine at temperatures in the range of about 0°C to about 50°C. The reaction is continued until completion, which typically occurs in from about 2 to 12 h.
[0121] Removal of the protecting groups can be carried out with acids, such as trifluoroacetic acid (TFA), hydrochloric acid, p-toluenesulfonic acid, and the like, in an inert organic solvent such as dichloromethane, dichloroethane, dioxane, THF, and the like. The removal is typically conducted at low temperatures, e.g., 0°C, and then gradually allowed to warm to room temperature to provide the product.
[0122] Also as shown in Scheme 11, an appropriately 7-substititued lincosamine intermediate (prepared, for example, according to any one of Schemes 1-5) and an appropriately substituted pyridin-2-yl carboxylic acid (prepared, for example, according to Scheme 10) are condensed under reactive conditions, preferably in an inert organic solvent, in the presence of a coupling reagent and an organic base, as described above.
[0123] The pyridine lib is hydrogenated to provide the piperidyl product. The hydrogenation is typically performed in a polar organic solvent such as methanol, ethanol, and the like, using platinum(IN)oxide in the presence of an acid such as HCl, acetic acid, and the like, in a Parr bottle. The bottle- is purged, and charged with H to approximately 40 to 70 psi and shaken until completion, typically approximately 24 h. The reaction mixture is filtered, e.g., through a celite pad, and washed several times with a polar organic solvent such as methanol.
Evaporation of the combined washings and filtrate affords the piperidyl product.
[0124] The coupling of pyridine carboxylic acids and lincosamines to pyridine lib followed by reduction to the piperidyl product may also be conducted as described in Birkenmeyer, et al.,
Journal of Medicinal Chemistry 1984, 27, 216-223.
[0125] Scheme 12 below illustrates the alkylation of the nitrogen of the pyrrolidinyl or piperidinyl ring, wherein R6 is alkyl, hydroxyalkyl, alkylene-substituted heterocycle, or alkylene-heterocycle, and R1, R2, R3, and R9 are as defined for formula (I).
Scheme 12. General synthesis of P-Ν-substituted lincosamines. a. alkylating agents [0126] As shown in Scheme 12, tlie lincosamine 12a can be Ν-substituted by contact with an alkylating agent in the presence of a suitable base to provide a product 12b. Suitable alkylating agents which can be used include epoxides, alkyl bromides, and the like. Suitable bases which can be used include potassium carbonate, cesium carbonate triethylamine, and the like. The alkylation reaction is typically conducted in a polar organic solvent such as methanol or DMF. The alkylation reaction is typically conducted at low temperatures in the range of 0°C to -10°C for 10 to 20 h.
[0127] Scheme 13 below illustrates the acylation of the nitrogen of the pyrrolidinyl or piperidinyl ring, wherein R6 is -C(O)O-alkylene-cycloalkyl, -C(O)O-alkylene-substituted cycloalkyl, -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -[C(O)O]-alkylene-heterocycle, -[C(O)O]-alkylene-substituted heterocycle, and R1, R2, R3, and R9 are as defined for formula (I).
Scheme 13. General synthesis of 1 '-N-substituted lincosamines. [0128] As shown in Scheme 13, the lincosamine 12a can be N-substituted by contact with an acyl chloride 101, such as R6-X, wherein X is a suitable leaving group, and is preferably halogen, even more preferably chloride in the presence of a suitable base to provide a product 102. Examples of compound 101, include bromofluorenyl, Cl-C(O)O-alkyl, Cl-C(O)O-aryl, and the like. Suitable bases which can be used include DCC, TEA, and the like. The reaction is typically conducted in a polar organic solvent such as methanol or DMF. The reaction is typically conducted at low temperatures in the range of -10°C to 20°C. [0129] Scheme 16 below illustrates the general synthesis of trø«5'-alkylazetidine carboxylic acids.
16a 16b 16c
16d 16e
16f 16g Scheme 16. General synthesis of trα«-s-alkylazetidine carboxylic acids.
[0130] The following reaction scheme may be used in the synthesis of trαrø-alkylazetidine carboxylic acids, wherein R9 is as defined herein: (a) (i) LDA, THF, 0 °C, (ii) bromoalkane or bromoalkene; (b) TMSCHN2, MeOH, 23 °C; (c) (i) TMSCHN2, MeOH, 23 °C, (ii) H2, Pd/C,
EtOAc, 23 °C; (d) Et3N.3HF, THF, 23 °C; (e) LiAlH4, THF, 68 °C; (f) Boc2O, CH2C12, 23°C;
(g) RuCl3.xH2O, NaIO4, acetone, H20, 23 °C.
[0131] Scheme 17 below illustrates the general synthesis of frans-alkylazetidine carboxylic acids via the aldehyde.
17a 17b
17c 17d
17e 17f Scheme 17. General synthesis of tr rø-s-alkylazetidine carboxylic acids via aldehyde. [0132] The following reaction scheme may be used in the synthesis of trαrø-alkylazetidine carboxylic acids: (a) TBSCl, imidazole, DMF, 23 °C; (b) (i) ozone, CH2C12, -78 °C, (ii) PPh3; (c) olefination: P+Phs salt, base, solvent; (d) H2, Pd/C, EtOAc, 23 °C or KO2CN=NCO2K, AcOH, dioxane, 23 °C; (e) TBAF, THF, 23 °C; (f) RuCl3.xH2O, NaIO4, acetone, H20, 23 °C. [0133] Scheme 18 below illustrates a general synthesis of 2-substituted ester prodrug compounds.
Scheme 18. General synthesis of 2-substituted esters. [0134] The following representative reaction scheme may be used in the synthesis of 2- substituted prodrug compounds of the invention: a. (Boc)2O, aq. KHCO3 THF b. j»-anisaldehyde dimethyl acetal, PPTS, c. Rπacylating agent, base d. TFA, DCE, water. [0135] Scheme 19 below illustrates a general method for the synthesis of 4,5-disubstituted- 2,3,6,7-tetrahydro-azepine-2-carboxylic acid derivatives.
Scheme 19.
[0136] The following general reaction scheme may be used in the preparation of 4,5- disubstituted-253,6,7-tetrahydro-azepine-2-carboxylic acid derivatives: a. 2,2- dimethoxypropane, Catalytic HCl, MeOH b. 2-nitrobenzenesulfonyl chloride, 2,4,6-collidine, dichloroethane c. Cs2C03, TBABr, DMF, Homoallylic tosylates or halides d. Benzylidene[l,3- bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene] dichloro-(tricyclohexylphosphine)ruthenium e. 7-methyl,l,5,7-triazabicyclo[4.4.0]dec-5-ene, thiophenol f. (Boc) O, TEA g. Aq. LiOH, Dioxane.
[0137] Scheme 21 below illustrates the synthesis of 4,4 di-substituted pyrrolidine and piperidine carboxylic acids, wherein R9 is a suitable substituent and the other substituent is fluorine.
Scheme 21. Synthesis of intermediate 21d where R >9 is defined herein. [0138] The following representative reaction scheme may be used in Scheme 21 : (a) Tetraallyltin, BF3 »Et2O or R9M (R9 or R9' carbon nucleophile) (b) H2/Pd (c) DAST (d) aq. LiOH or appropriate carboxylate ester deprotection conditions.
Pharmaceutical Formulations
[0139] When employed as pharmaceuticals, the compounds of the subject invention are usually administered in the form of pharmaceutical compositions. These compounds can be administered by a variety of routes including oral, parenteral, transdermal, topical, rectal, and intranasal. These compounds are effective as both injectable and oral compositions. Such compositions are prepared in a manner well known in the pharmaceutical art and comprise at least one active compound.
[0140] This invention also includes pharmaceutical compositions that contain, as the active ingredient, one or more of the compounds of the subject invention above associated with pharmaceutically acceptable carriers. In making the compositions of this invention, the active ingredient is usually mixed with an excipient, diluted by an excipient or enclosed within such a carrier which can be in the form of a capsule, sachet, paper or other container. The excipient employed is typically an excipient suitable for administration to human subjects or other mammals. When the excipient serves as a diluent, it can be a solid, semi-solid, or liquid material, which acts as a vehicle, carrier or medium for the active ingredient. Thus, the compositions can be in the form of tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid medium), ointments containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions, and sterile packaged powders. [0141] In preparing a formulation, it may be necessary to mill the active compound to provide the appropriate particle size prior to combining with the other ingredients. If the active compound is substantially insoluble, it ordinarily is milled to a particle size of less than 200 mesh. If the active compound is substantially water soluble, the particle size is normally adjusted by milling to provide a substantially uniform distribution in the formulation, e.g., about 40 mesh.
[0142] Some examples of suitable excipients include lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, sterile water, syrup, and methyl cellulose. The formulations can additionally include: lubricating agents such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preserving agents such as methyl- and propylhydroxy-benzoates; sweetening agents; and flavoring agents. The compositions of the invention can be formulated so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient by employing procedures known in the art.
[0143] The quantity of active component, that is the compound according to the subject invention, in the pharmaceutical composition and unit dosage form thereof may be varied or adjusted widely depending upon the particular application, the potency of the particular < compound and the desired concentration.
[0144] The compositions are preferably formulated in a unit dosage form, each dosage containing from about 5 to about 100 mg, more usually about 10 to about 30 mg, of the active ingredient. The term "unit dosage forms" refers to physically discrete units suitable as unitary dosages for human subjects and other mammals, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient. Preferably, the compound of the subject invention above is employed at no more than about 20 weight percent of the pharmaceutical composition, more preferably no more than about 15 weight percent, with the balance being pharmaceutically inert carrier(s).
[0145] The active compound is effective over a wide dosage range and is generally administered in a pharmaceutically or therapeutically effective amount. It will be understood, however, that the amount of the compound actually administered will be determined by a physician, in the light of the relevant circumstances, including the condition to be treated, the severity of the bacterial infection being treated, the chosen route of administration, the actual compound administered, the age, weight, and response of the individual patient, the severity of the patient's symptoms, and the like. [0146] In therapeutic use for treating, or combating, bacterial infections in warm-blooded animals, the compounds or pharmaceutical compositions thereof will be administered orally, topically, transdermally, and/or parenterally at a dosage to obtain and maintain a concentration, that is, an amount, or blood-level of active component in the animal undergoing treatment which will be antibacterially effective. Generally, such antibacterially or therapeutically effective amount of dosage of active component (i.e., an effective dosage) will be in the range of about 0.1 to about 100, more preferably about 1.0 to about 50 mg/kg of body weight/day. [0147] For preparing solid compositions such as tablets, the principal active ingredient is mixed with a pharmaceutical excipient to form a solid preformulation composition containing a homogeneous mixture of a compound of the present invention. When referring to these preformulation compositions as homogeneous, it is meant that the active ingredient is dispersed evenly throughout the composition so that the composition may be readily subdivided into equally effective unit dosage forms such as tablets, pills and capsules. This solid preformulation is then subdivided into unit dosage forms of the type described above containing from, for example, 0.1 to about 500 mg of the active ingredient of the present invention. [0148] The tablets or pills of the present invention may be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action. For example, the tablet or pill can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former. The two components can be separated by an enteric layer that serves to resist disintegration in the stomach and permit the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymeric acids and mixtures of polymeric acids with such materials as shellac, cetyl alcohol, and cellulose acetate. [0149] The liquid forms in which the novel compositions of the present invention may be incorporated for administration orally or by injection include aqueous solutions, suitably flavored syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as corn oil, cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles.
[0150] Compositions for inhalation or insufflation include solutions and suspensions in pharmaceutically acceptable, aqueous or organic solvents, or mixtures thereof, and powders. The liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as described supra. Preferably the compositions are administered by the oral or nasal respiratory route for local or systemic effect. Compositions in preferably pharmaceutically acceptable solvents may be nebulized by use of inert gases. Nebulized solutions may be inhaled directly from the nebulizing device or the nebulizing device may be attached to a face mask tent, or intermittent positive pressure breathing machine. Solution, suspension, or powder compositions may be administered, preferably orally or nasally, from devices that deliver the formulation in an appropriate manner.
[0151] The following formulation examples illustrate representative pharmaceutical compositions of the present invention.
Formulation Example 1
[0152] Hard gelatin capsules containing the following ingredients are prepared: Quantity Ingredient (mg/capsule Active Ingredient 30.0 Starch 305.0 Magnesium stearate 5.0
[0153] The above ingredients are mixed and filled into hard gelatin capsules in 340 mg quantities.
Formulation Example 2 [0154] A tablet formula is prepared using the ingredients below:
Quantity Ingredient (mg/capsule Active Ingredient 25.0 Cellulose, microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0
[0155] The components are blended and compressed to form tablets, each weighing 240 mg.
For ulatioii Example 3
[0156] A dry powder inhaler formulation is prepared containing the following components: Ingredient Weight % Active Ingredient 5 Lactose 95
[0157] The active ingredient is mixed with the lactose and the mixture is added to a dry powder inhaling appliance. Formulation Example 4
[0158] Tablets, each containing 30 mg of active ingredient, are prepared as follows Quantity Ingredient (mg/capsule) Active Ingredient 30.0 mg Starch 45.0 mg Microcrystalline cellulose 35.0 mg Polyvinylpyrrolidone (as 10% solution in sterile water) 4.0 mg Sodium carboxymethyl starch 4.5 mg Magnesium stearate 0.5 mg Talc 1.0 mg Total 120 mg
[0159] The active ingredient, starch and cellulose are passed through a No. 20 mesh U.S. sieve and mixed thoroughly. The solution of polyvinylpyrrolidone is mixed with the resultant powders, which are then passed through a 16 mesh U.S. sieve. The granules so produced are dried at 50°C to 60°C and passed through a 16 mesh U.S. sieve. The sodium carboxymethyl starch, magnesium stearate, and talc, previously passed through a No. 30 mesh U.S. sieve, are then added to the granules which, after mixing, are compressed on a tablet machine to yield tablets each weighing 120 mg. .
Formulation Example 5
[0160] Capsules, each containing 40 mg of medicament are made as follows: Quantity Ingredient (mg/capsule) Active Ingredient 40.0 mg Starch 109.0 mg Magnesium stearate 1.0 mg Total 150.0 mg
[0161] The active ingredient, starch and magnesium stearate are blended, passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin capsules in 150 mg quantities.
Formulation Example 6
[0162] Suppositories, each containing 25 mg of active ingredient are made as follows: Ingredient Amount Active Ingredient 25 Saturated fatty acid glycerides to 2,000 mg [0163] The active ingredient is passed through a No. 60 mesh U.S. sieve and suspended in the saturated fatty acid glycerides previously melted using the minimum heat necessary. The mixture is then poured into a suppository mold of nominal 2.0 g capacity and allowed to cool.
Formulation Example 7 [0164] Suspensions, each containing 50 mg of medicament per 5.0 mL dose are made as follows: Ingredient Amount Active Ingredient 50 mg Xanthan gum 4.0 mg Sodium carboxymethyl cellulose (11%) Microcrystalline cellulose (89%) 50.0 mg Sucrose 1.75 g Sodium benzoate lO.O mg Flavor and Color q.v. Purified water to 5.0 mL
[0165] The active ingredient, sucrose and xanthan gum are blended, passed through a No. 10 mesh U.S. sieve, and then mixed with a previously made solution of the microcrystalline cellulose and sodium carboxymethyl cellulose in water. The sodium benzoate, flavor, and color are diluted with some of the water and added with stirring. Sufficient water is then added to produce the required volume.
Formulation Example 8 Quantity Ingredient (mg/capsule) Active Ingredient 15.0 mg Starch 407.0 mg Magnesium stearate 3.0 mg Total 425.0 mg
[0166] The active ingredient, starch, and magnesium stearate are blended, passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin capsules in 425.0 mg quantities.
Formulation Example 9
[0167] A subcutaneous formulation may be prepared as follows: Ingredient Quantity Active Ingredient 5.0 mg Corn Oil 1.0 mL Formulation Example 10
[0168] A topical formulation may be prepared as follows: Ingredient Quantity Active Ingredient 1 - 10 g Emulsifying Wax 30 g Liquid Paraffin 20 g White Soft Paraffin to 100 g
[0169] The white soft paraffin is heated until molten. The liquid paraffin and emulsifying wax are incorporated and stirred until dissolved. The active ingredient is added and stirring is continued until dispersed. The mixture is then cooled until solid.
Formulation Example 11
[0170] An intravenous formulation may be prepared as follows: Ingredient Quantity Active Ingredient 250 mg Isotonic saline 1000 mg
[0171] Another preferred formulation employed in the methods of the present invention employs transdermal delivery devices ("patches"). Such transdermal patches may be used to provide continuous or discontinuous infusion of the compounds of the present invention in controlled amounts. The construction and use of transdermal patches for the delivery of pharmaceutical agents is well known in the art. See, e.g., U.S. Patent 5,023,252, issued June 11, 1991, herein incorporated by reference. Such patches may be constructed for continuous, pulsatile, or on demand delivery of pharmaceutical agents.
[0172] Frequently, it will be desirable or necessary to introduce the pharmaceutical composition to the brain, either directly or indirectly. Direct techniques usually involve placement of a drug delivery catheter into the host's ventricular system to bypass the blood- brain barrier. One such implantable delivery system used for the transport of biological factors to specific anatomical regions of the body is described in U.S. Patent 5,011,472 which is herein incorporated by reference.
[0173] Indirect techniques, which are generally preferred, usually involve formulating the compositions to provide for drug latentiation by the conversion of hydrophilic drugs into lipid- soluble drugs. Latentiation is generally achieved through blocking of the hydroxy, carbonyl, sulfate, and primary amine groups present on the drug to render the drug more lipid soluble and amenable to transportation across the blood-brain barrier. Alternatively, the delivery of hydrophilic drugs may be enhanced by infra-arterial infusion of hypertonic solutions which can transiently open the blood-brain barrier.
[0174] Other suitable formulations for use in the present invention can be found in Remington's Pharmaceutical Sciences, Mace Publishing Company, Philadelphia, PA, 17th ed. (1985).
[0175] As noted above, the compounds described herein are suitable for use in a variety of drug delivery systems described above. Additionally, in order to enhance the in vivo serum half-life of the administered compound, the compounds may be encapsulated, introduced into the lumen of liposomes, prepared as a colloid, or other conventional techniques may be employed which provide an extended serum half-life of the compounds. A variety of methods are available for preparing liposomes, as described in, e.g., Szoka, et al., U.S. Patent Nos. 4,235,871, 4,501,728 and 4,837,028 each of which is incorporated herein by reference. [0176] As noted above, the compounds administered to a patient are in the form of pharmaceutical compositions described above. These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered. The resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration. The pH of the compound preparations typically will be between 3 and 11 , more preferably from 5 to 9 and most preferably from 7 and 8. It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.
[0177] In general, the compounds of the subject invention will be administered in a therapeutically effective amount by any of the accepted modes of administration for agents that serve similar utilities. Toxicity and therapeutic efficacy of such compounds can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD5o (the dose lethal to 50% of the population) and the ED5o (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD5o/ ED50. Compounds that exhibit large therapeutic indices are preferred.
[0178] The data obtained from the cell culture assays and animal studies can be used in formulating a range of dosage for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. For any compound used in the method of the invention, the therapeutically effective dose can be estimated initially from cell culture assays. A dose may be formulated in animal models to achieve a circulating plasma concentration range which includes the IC50 (the concentration of the test compound which achieves a half-maximal inhibition of symptoms) as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Levels in plasma may be measured, for example, by high performance liquid chromatography.
Utility
[0179] The compounds, prodrugs and pharmaceutically acceptable salts thereof, as defined herein, may have activity against a variety of bacteria, protozoa, fungi, and parasites. By way of example, the compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against gram positive and gram negative bacteria. The compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against a variety of fungi, including fungi from the genus Mucor and Candida, e.g., Mucor racemosus or Candida albicans. The compounds, prodrugs and pharmaceutically acceptable salts thereof may be active against a variety of parasites, including malaria and cyptosporidium parasite.
[0180] The compounds of the subject invention exhibit activity against a variety of bacterial infections, including, for example, gram positive infections, gram negative infections, mycobacteria infections, mycoplasma infections, and chlamydia infections. [0181] Since the compounds of the subject invention exhibit potent activities a variety of bacteria, such as gram positive bacteria, the compounds of the present invention are useful antimicrobial agents and may be effective against a number of human and veterinary pathogens, including gram positive bacteria. The Gram positive organisms against which the compounds of tlie present invention are effective include Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidennidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile, and the like.
[01S2] The compounds of the subject invention may be combined with one or more additional antibacterial agents. One or more of the additional antibacterial agents may be active against gram negative bacteria. Additionally, one or more of the additional antibacterial agents may be active against gram positive bacteria. The combination of the compounds of the subject invention and the one or may additional antibacterial agents may be used to treat a gram negative infection. Additionally, the combination of the compounds of the subject invention and the one or more additional antibacterial agents may be used to treat a gram positive infection. The combination of compounds of the subject invention and the one or more additional antibacterial agents may also be used to treat a mycobacteria infection, mycoplasma infection, or chlamydia infection.
[0183] The in vitro activity of compounds of the subject invention may be assessed by standard testing procedures such as the determination of minimum inhibitory concentration (MIC) by agar dilution as described in "Approved Standard. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically," 3rd ed., published 1993 by the National Committee for Clinical Laboratory standards, Villanova, Pennsylvania, USA. [0184] The amount administered to the mammalian patient will vary depending upon what is being administered, the purpose of the administration, such as prophylaxis or therapy, the state of the patient, the manner of administration, and the like. In therapeutic applications, compositions are administered to a patient already suffering from a disease in an amount sufficient to cure or at least partially arrest the symptoms of the disease and its complications. An amount adequate to accomplish this is defined as "therapeutically effective dose." Amounts effective for this use will depend on the disease condition being treated as well as by the judgment of the attending clinician depending upon factors such as the severity of the inflammation, the age, weight and general condition of the patient, and the like. [0185] The compositions administered to a patient are in the form of pharmaceutical compositions described above. These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered. The resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration. The pH of the compound preparations typically will be between 3 and 11, more preferably from 5 to 9 and most preferably from 7 to 8. It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.
[0186] The therapeutic dosage of the compounds of the present invention will vary according to, for example, the particular use for which the treatment is made, the manner of administration of the compound, the health and condition of the patient, and the judgment of the prescribing physician. For example, for intravenous administration, the dose will typically be in the range of about 20 mg to about 500 mg per kilogram body weight, preferably about 100 mg to about 300 mg per kilogram body weight. Suitable dosage ranges for intranasal administration are generally about 0.1 mg to 1 mg per kilogram body weight. Effective doses can be extrapolated from dose-response curves derived from in vitro or animal model test systems.
[0187] The following synthetic and biological examples are offered to illustrate this invention and are not to be construed in any way as limiting the scope of this invention.
EXAMPLES
[0188] In the discussion above and in the examples below, the following abbreviations have the following meanings. If an abbreviation is not defined, it has its generally accepted meaning.
ιethyl/MTL = l-methylthio-7-deoxy-7- methyllincosamine apt = apparent triplet atm = atmospheres Bn = benzyl Boc = tert-butoxycarbonyl protecting group br s = broad singlet
BSTFA N,O- bis(trimethylsilyl)trifluoroacetamide Cbz = carbonyloxybenzyloxy protecting group CDC13 = deuterated chloroform
CD3OD = deuterated methanol cfu = colony forming units D = doublet DAST = dimethylaminosul-furtrifluoride dd = doublet of doublets dddd = doublet of doublets of doublet of doublets dt = doublet of triplets DCC = dicyclohexylcarbodiimide DCE = dicholoroethane DCM = dichloromethane DIEA = diisopropyethylamine
DMAP = dimethylaminopyridine DMF = dimethylformamide
DMSO = dimethyl sulfoxide DPPA = diphenylphosphoryl azide EDso = dose therapeutically effective in 50% of the population EDC = 1 -(3 -dimethylaminopropyl)-3 - ethylcarbodiamide HCl Equiv = equivalents ESMS = electrospray mass spectrometry Et ethyl EtOAc = ethyl acetate Et20 = diethyl ether g = grams h = hours HATU = O-(7-azabenzotriazol- 1 -yl)-N,N,N ' ,N' - tetramethyluronium hexafluorophosphate HBTU O-(Benzotriazol-l-yl)-N,N,N',N*- tetramethyluronium hexafluorophosphate HOBT = 1-hydroxybenzotriazole hydrate
1H NMR = Hydrogen Nuclear Magnetic Resonance spectroscopy HPLC = high pressure liquid chromatography Hz = hertz IC50 ------ concentration of the test compound which achieves a half-maximal inhibition of symptoms J = coupling constant in hertz L = liters LD50 = dose lethal to 50% of the population LDA = lithium diisopropylamide LiHMDS = lithium hexamethyldisilazide LiN(iPr)2 = lithium diisopropylnitride m = multiplet M = molar MCPBA -= 2-(4-chloro-o-tolyloxy) acetic acid Me = methyl MeCN -= acetonitrile MeOH ------ methanol mg =--= milligrams MHz -= megahertz Min ------ minutes mL = milliliters Mm ------ millimeter mmol -= millimol
MS(ESPOS) ~ mass specfrometry by positive mode electrospray ionization
MS(ESNEG) ' — Mass Specfrometry by negative mode electrospray ionization MTL 1-methylthiolincosamine (methyl 6- amino-6,8-dideoxy-l-thio-erythro-β-D- galacto-octopyranoside) N = normal NMR ------ nuclear magnetic resonance OBz = benzyloxy protecting group OtBu = tert-butoxy Pd/C = palladium/carbon pg = picograms Ph = phenyl PPTS = pyridinium p-toluenesulfonate Pro = L-proline psi -= pounds per square inch PTFE = polytetrafluoroethylene q -= quartet q.v. = quantitative Rf = Retention factor rt = room temperature s = singlet sat. = saturated t -= triplet TBAF -= tetrabutylammonium fluoride TBS = tert-butyldimethylsilyl TEA = triethylamine TFA = trifluoroacetic acid THF = tetrahydrofuran TLC = thin layer chromatography TMS = trimethylsilyl μg = micrograms μL = microliters μm = micromolar v/v = volume by volume w/w = weight by weight
[0189] Additionally, the term "Aldrich" indicates that the compound or reagent used in the following procedures is commercially available from Aldrich Chemical Company, Inc., 1001 West Saint Paul Avenue, Milwaukee, Wl 53233 USA; the term "Fluka" indicates that the compound or reagent is commercially available from Fluka Chemical Corp., 980 South 2nd Street, Ronkonkoma NY 11779 USA; the term "Lancaster" indicates that the compound or reagent is commercially available from Lancaster Synthesis, Inc., P.O. Box 100 Windham, NH 03087 USA; the term "Sigma" indicates that the compound or reagent is commercially available from Sigma, P.O. Box 14508, St. Louis MO 63178 USA; the term "Chemservice" indicates that the compound or reagent is commercially available from Chemservice Inc., Westchester, PA, USA; the term "Bachem" indicates that the compound or reagent is commercially available from Bachem Bioscience Inc., 3700 Horizon Drive, Renaissance at Gulph Mills, King of Prussia, PA 19406 USA; the term "Maybridge" indicates that the compound or reagent is commercially available from Maybridge Chemical Co.
[0190] Trevillett, Tintagel, Cornwall PL34 OHW United Kingdom; the term "RSP" indicates that the compound or reagent is commercially available from RSP Amino Acid Analogs, Inc., 106 South St., Hopkinton, MA 01748, USA, and the term "TCI" indicates that the compound or reagent is commercially available from TCI America, 9211 North Harborgate St., Portland, Oregon, 97203, OR, USA; the term "Toronto" indicates that the compound or reagent is commercially available from Toronto Reasearch Chemicals, Inc., 2 Brisbane Rd., New York, ON, Canada M3 J2J8; the term "Alfa" indicates that the compound or reagent is commercially available from Johnson Matthey Catalog Company, Inc. 30 Bond Street, Ward Hill, MA 018350747; and the term "Nova Biochem" indicates that the compound or reagent is commercially available from NovaBiochem USA, 10933 North Torrey Pines Road, P.O. Box 12087, La Jolla CA 92039-2087.
[0191] In the examples below, all temperatures are in degrees Celsius (unless otherwise indicated) and the following general procedures are used to prepared the compounds as indicated. It will be appreciated by one of skill in the art that the following general procedures are meant to be illustrative only and that the methods may be broadened to synthesize other compounds of the subject invention.
General Procedures Method A
[0192] Methyl 6-amino-6,8-dideoxy-l-thio-erythro-β-D-galacto-octopyranoside la (R1 = Me) (MTL) was prepared as described by Hoeksema, H. et al., J. Am. Chem. Soc, 1967, 89, 2448-2452. N-(Benzyloxycarbonyloxy)succinimide (5.8 g 23.1 mmol) and la (5.0 g, 19.7 mmol) were suspended in pyridine (40mL) and stirred under N2 atmosphere 36 h. The reaction mixture was cooled to 0°C and then bis-N,0-trifluoroacetamide (15.7 mL, 59.0 mmol) was added by syringe over 2 min. The reaction mixture was allowed to warm to rt and stirred for 42 h. Toluene (100 mL) was added and the reaction mixture was evaporated to dryness. The residue was taken up in ethyl acetate (400 mL). The organic solution was washed quickly with 10% citric acid (200 mL), H20 (3 x 100 mL), saturated NaHC03 (100 mL), and brine (2 x 100 mL), and dried over Na2S04 and evaporated to dryness. Chromatography of the crude product on silica 10% EtOAc/Hexanes containing 0.2% TEA after co-evaporation from toluene (100 mL) and cyclohexane (2 x 100 mL) provided the protected product lb (P=Cbz, R1=Me) (7.2 g, 54%) as a colorless oil.
[0193] 1H NMR (300 MHz, CD3SOCD3) δ 7.34-7.31 (m, 5), 7.05 (d, J= 8.2, 1), 5.19 (d, J= 5.8, 1), 5.01 (d, J= 1.6, 2), 3.99 (apt dt, J= 5.5, 9.3, 9.3, 2), 3.93-3.86 (m, 3), 3.49 (dd, J= 2.5, 9.6, 1), 2.01 (s, 3), 1.03 (d, J= 6.3, 3), 0.10 (s, 9), 0.09 (s, 9), 0.04 (m, 18). [0194] To dimethylsulfoxide (413 μL, 5.82 mmol) in DCM (1.5 mL) cooled to -72°C was added oxalyl chloride 2 M in DCM (1.49 mL, 2.98 mmol) over 1 min. After 25 min the protected product lb (1.92 g, 2.84 mmol) in DCM (4.0 mL) was added by cannula. The resulting reaction mixture was stirred for 25 min and then allowed to warm to -50°C (dry ice acetonitrile) and maintained at this temperature for 2 h. To the reaction mixture was added TEA (1.29 mL, 3.30 mmol). After 25 min the reaction mixture was diluted with EtOAc (300 mL). The resulting organic solution was washed quickly with 5% citric acid (300 mL), H 0 (2 x 300 mL), saturated NaHC03 (100 L), brine (100 mL) dried over Na2S0 and evaporated to dryness with the aid of toluene (100 mL) to provide the product lc. The product lc (P=Cbz, R1=Me) was obtained as a colorless crystalline solid (1.60 g, 94%) after co-evaporation with n-pentane and removal of residual solvent under high vacuum.
[0195] !HNMR (300 MHz CDC13) δ 7.37-7.33 (m, 5), 5.60 (m, 1), 5.21 (d, J = 5.2, 1), 5.17 (d, J= 12.4, 1), 5.08 (d, J= 12.4, 1), 4.74 (m, 1), 4.16-4.12 (m, 2), 3.87 (d, J = 2.2, 1), 3.69 (dd, J= 2.5, 9.3, 1), 2.01 (br s, 3), 1.90 (s, 3), 0.19 (s, 9), 0.16 (s, 9), 0.15 (s, 9).
Method B
[0196] The Boc-protected product lc (P=Boc, R1 =Me) may be prepared in general as outlined above, la (R1 = Me) (MTL) (Dried at 50°C high vacuum) (21.8 g, 86 mmol) was suspended in methanol (200 mL) and TEA (26 mL) and was cooled to 0°C on ice. To this mixture di-t-butyldicarbonate (57.0 g, 0.26 mol) was added. The reaction mixture was then stirred overnight at rt. To the reaction mixture was added toluene (100 mL). The solvents were removed to a total volume of 100 mL, leaving a thick suspension to which cyclohexane (300 mL) was added. The resulting solid precipitate was triturated, then filtered and washed with cyclohexane, ether, and pentane and dried to constant weight. The crude Boc-protected product was used without further purification (87%).
[0197] TLC Rp 0.75 (10% MeOH/DCM); MS(ESPOS): 354 [M+H]+; 2H NMR (300 MHz, CD3OD) δ 0.14 (d, J = 6.3, 3),1.43 (s, 9), 2.07 (s, 3), 3.55 (dd, J = 3.3, 10.43, 1), 3.84 -4.08 (m, 3), 4.10-4.15 (m, 2), 5.25 (d, J = 5.5, 1).
[0198] To N-Boc-1-methylthiolincosamide (240 mg, 0.68 mmol) in DMF (5 mL, BSTFA (0.52 mL, 2.0 mmol) and triethylamine (0.14 mL, 1.42 mmol) were added at 0°C and then , stirred at rt overnight. DMF was removed and the crude product was quickly passed through a silica gel column (pretreated with 2% TEA in ethyl acetate) eluting with 10% ethyl acetate in hexanes lb (P=Boc, R1==Me) (350 mg, 95%). To oxalyl chloride (0.16 mL, 0.78 mmol) in dichloromethane (5 mL) at -60°C, dimethylsulfoxide (0.22 mL, 0.78 mmol) was added slowly and then stirred for 15 min. After which, lb (370 mg, 0.65 mmol) in DCM (5 mL) was added slowly. The reaction mixture was stirred for 45 min, during which the reaction temperature was raised to -40°C. Triethylamine (0.70 mL, 3.25 mmol) was then added and the stirring continued for an additional 15 min at -40°C. It was then extracted with DCM (100 mL) and washed with 10% citric acid (50 mL). The residue obtained on removal of solvent was then purified on silica gel column using 10% ethyl acetate in hexanes as eluent lc (P=Boc, R!=Me) as a colorless oil (289 mg, 78%). [0199] TLC: Rf = 0.60 (10% EtOAc/Hexanes); MS(ESPOS): 590 [M+Na]+; 1H NMR (300 MHz, CDC13) δ 0.11 (s, 18), 0.17 (s, 18), 1.40 (s, 9), 1.84 (s, 3), 2.26 (s, 3), 3.63 (dd, J = 2.7, 9.34, 1), 3.82 (d, J = 1.9, 1), 4.01-4.12 (m, 2), 5.15 (d, J = 5.5, 1).
Method C
[0200] Triphenylphosphonium bromide (3.29 g, 9.2 mmol) and potassium tert-butoxide (715 mg, 6.4 mmol) under N2 atmosphere were suspended in toluene (31 mL) with vigorous stirring. After 4 h protected product lc (P=Cbz, R1=Me) (1.4 g, 2.36 mmol) in toluene (20 mL) was added by cannula. The resulting reaction mixture was stirred 2 h and then diluted with EtOAc (250 mL). The resulting organic solution was washed quickly with H20 (2 x 100 mL), brine (1 x 100 mL) dried over Na S04 and evaporated to dryness. Chromatography of the crude product on silica 6% EtOAc/Hexanes containing 0.2% TEA gave the alkene product 2a (P=Cbz, R1=Me, R2=CH ) as a colorless oil that crystallized after co-evaporation from toluene and cyclohexane (0.65 g, 46%).
[0201] IH NMR (300 MHz CDC13) δ 7.35-7.27 (m, 5), 6.36 (d, J = 7.1, 1), 5.24 (d, J = 5.5, 1), 5.08 (m, 4), 4.34 (m, 1), 4.16 (m, 2), 3.88 (d, J = 2.2, 1), 3.61 (dd, J = 2.2, 9.3, 1), 2.20 (s, 3), 1.79 (s, 3), 0.17-0.13 (m, 27).
[0202] The product 2a (P=Cbz, R1=Me, R2=CH2) (490 mg, 0.82 mmol)s in ethanol (50 mL) was added to 10% palladium on carbon (Degussa wet form 50% w/w water) (700 mg) in a par bottle. The bottle was purged, and charged with H2 to 65 psi and shaken 24 h. The reaction mixture was filtered through celite, rinsed with methanol. The organic solution was transferred to a resin funnel containing dry, washed Dowex 50w-400x H"1" form (0.8 g) and shaken for 10 min. After washing the resin with methanol three times and water two times, the saturated product 2b (R =Me, R -Me) was eluted from the resin by washing with 5% TEA in MeOH (35 mL, x 10 min x 5). The combined filtrate was evaporated to dryness, co-evaporated from EtOH twice and lyophilized from 1 : 1 MeCN/H20 to give the product as a colorless powder (198.4 mg 96%).
[0203] 'HNMR (300 MHz, D20) δ 5.17 (d, J = 5.8, 1), 3.97-3.84 (m, 3), 3.52 (dd, J = 3.0, 10.0, 1), 2.82 (dd, J = 4.4, 8.5, 1), 1.94 (s, 3),1.89-1.81 (m, 1), 0.82 (d, J = 6.9, 3), 0.72 (d, J = 6.9, 3). MS(ESPOS): 252.2 [M+H]+, (ESNEG): 250.4 [M-H]\ Method D
[0204] In the alternative when a Boc-protecting group is used, methyltriphenylphosphonium bromide (12 g, 33.6 mmol) and potassium t-butoxide (3g, 26.7 mmol) were taken in THF (70 mL) at 0°C, and stirred at rt for 4 h. Then Boc-protected product lc (P=Boc, R1=Me) (4.7 g, 8.2 mmol) in THF (30 mL) was added and stirred at rt for 2 h. After which it was extracted with EtOAc (300 mL), washed with brine (100 mL) and dried over sodium sulfate. The crude alkene 1 9 product 2a (P=Boc, R =Me, R =CH2) was purified on silica gel column chromatography using
10% EtOAc in Hexane as eluent (4.1 g, 87.6%).
[0205] TLC: Rf = 0.5 (10% of EtOAc in Hexane): 1H NMR (300 MHz, CD3OD) δ 7.24 (m,
2), 5.22 (d, J = 5.7, 1), 4.21 (m, 1), 4.09 (m, 2), 3.87 (d, J = 2.4, 1), 3.60 (dd, J = 2.7, 9.3, 1),
1.99 (s, 3), 1.76 (s, 3); 1.43 (s, 9); MS(ESPOS): 444 (M-2TMS+Na).
[0206] To the product 2a (P=Boc, R*=Me, R2=CH2) in methanol (30 mL), Dowex H1" resin
(1 g) was added and stirred at rt for 1 h. The resin was filtered and the product obtained on removal of solvent (2.4 g, 6.8 mmol,) was taken in MeOH (30 mL). Pd/C (2.5 g) was added and hydrogenated at 55 psi overnight. The crude product obtained on filtering and removal of solvent was purified on silica gel column chromatography using 10% MeOH in DCM to provide
Boc-protected 7-Methyl MTL as a white solid (2.06 g, 86%). TLC Rf= 0.5 (10% of MeOH' in
DCM).
[0207] 1H NMR (300 MHz, CD3OD) δ 5.23 (d, J = 5.4, 1), 4.11 (m, 1), 3.97 (d, J = 10.2, 1),
3.84 (m, 1), 3.52 (m, 1), 2.08 (s, 3), 1.44 (s, 9), 1.14 (m, 1), 0.93 (d, J = 6.9, 3), 0.85 (d, J = 6.9,
3); MS(ESPOS): 351[M+H]+.
[0208] To the Boc-protected 7-Methyl MTL (150 mg, 0.43 mmol) in dichloroethane (6 mL), dimethylsulfide (0.16 mL, 2.5 mmol) was added, followed by TFA (2 mL), water (0.16 mL) and stirred at rt for 1 h. The solvent was removed to obtain the crude product 2b (R1==Me, R2=Me).
After purification on silica gel column chromatography using 30% MeOH in DCM as eluent, the 1 9 product 2b (R =Me, R =Me) was obtained identical in all respects to the material obtained from method C.
Method E
[0209] Sodium hydride (80 mg, 3.3 mmol) under N2 atmosphere was suspended in THF (4 mL) with vigorous stirring. The suspension was cooled to -30°C and diethyl(cyanomethyl)phosphonate (805 μL, 5.0 mmol) was added. After 30 min protected product lc (P=Cbz, RJ=Me) (1.0 g, 1.7 -mmol) in THF (3 mL) was added by cannula. The resulting reaction mixture was stirred 4 h and then diluted with EtOAc (250 mL). The resulting organic solution was washed quickly saturated aqueous NaHC03 (1 x 100 mL), brine (1 x 50 mL) dried over Na2S0 and evaporated to dryness. Chromatography of the crude product on silica 6% EtOAc/Hexanes to 10% EtOAc/Hexanes containing 0.2% TEA gave the protected alkene product 2a (P=Cbz, R^Me, R2=CHCN) as a colorless oil (0.38 g, 37%). MS(ESPOS): 625.5.2 [M+H]+, ES(NEG): 659.5 [M+Cl].
[0210] The product 2a (P=Cbz, R1=Me, R2=CHCN) (180 mg, 0.29 mmol) in ethanol (15 mL) was added to 10% palladium on carbon (Degussa wet form 50% w/w water) (300 mg) in a Parr bottle and concentrated HCl (29 μL) was added. The bottle was purged, and charged with H2 to 65 psi and shaken for 24 h. The reaction mixture was filtered through celite, rinsed with methanol. The organic solution was transferred to a resin funnel containing dry, washed Dowex 50w-400x ϊm form (1 g) and shaken 10 min. After washing the resin with methanol twice and water, the saturated product 2b (R =Me, R =CH CN) was eluted from the resin by washing with 5% TEA in MeOH (20 mL x 20 min x 3) and MeCN (20 mL x 20 min). The combined organic filtrate was evaporated to dryness lyophilized from 1 : 1 MeCN/H20 to give the product 2b (R1=Me, R2=CH2CN) as a colorless solid (70 mg, 91%). ES(NEG): 275.3 [M-H]".
Method F
[0211] To the protected product lc (P=Cbz, R1==Me) (0.75g, 1.3 mmol) in THF (7.3 mL) was added MeMgCl (3M) in THF (7.0 mL 2.1 mmol) at 0°C. Over 30 min the reaction mixture was warmed to 4°C and after 4 h the reaction mixture was quenched with 1 :3 saturated aqueous NH4C1/H20 (lOmL). The quenched mixture was diluted to 100 mL with water and extracted with DCM (4 x 50 mL). The combined organic phase was dried and evaporated. The residue was dissolved in 1 :2:4 H20/HOAc/THF (100 mL) and stirred for 20 h, and then evaporated with the aid of toluene (2 x 100 mL). Chromatography 10:1 to 10:2 DCM / MeOH gave product 3a (P=Cbz, R^Me, R2=Me) (153 mg, 31%). [0212] (ESNEG): 399.5 [M-H]".
[0213] 3a (P=€bz, R*=Me, R2=Me) (79 mg, 0.2 mmol) in ethanol (10 mL) was added to 10% palladium on carbon (Degussa wet form 50% w/w water) (400 mg) in a Parr bottle. The bottle was purged, and charged with H2 to 65 psi and shaken 6 h. The reaction mixture was filtered through celite, rinsed with methanol. The combined filtrate was evaporated to dryness and lyophilized from 1:1 MeCN/H20 to give the product 3b (R1==Me, R2=Me) as a colorless powder (42 mg, 80%).
[0214] 1H NMR (300 MHz, D2O) δ 5.33 (d, J = 5.8,-1), 4.83-4.06 (m, 3), 3.65-3.60 (m, 1), 3.06-3.03 (m, 1), 2.18 (s, 3), 1.30 (s, 3), 1.23 (s, 3). MS(ESPOS): 268.4 [M+H]+, MS(ESNEG): 266.2 [M-H]".
Method G
[0215] To the Boc-protected product lc (P=Boc, R1=Me) (100 mg, 0.18 mmol) in methanol (3 mL), O-trimethylsilylhydroxylamine (0.10 mL, 0.88 mmol) was added and stirred at rt overnight. The solvent was removed to obtain the crude Boc-protected product 4a (P=Boc, R1=Me, R7=H). To the crude product 4a (95 mg, 0.15 mmol), 30% trifluoroacetic acid in dichloroethane (10 mL) and dimethyl sulfide (0.5 mL) were added and stirred for 1 h. The solvent was removed and the product 4b (R1=Me, R7=H) was taken as such for the next step. [0216] TLC: R_= 0.35 (10% MeOH/DCM); MS(ESPOS): 267 [M+H]+; 1H NMR (300 MHz, CD3OD) δ 1.96 (s, 3), 2.09 (s, 3), 3.58 (dd, J - 3.3, 10.2, 1), 3.90 (s, 1), 4.11 (dd, J - 5.7, 10.20, 1), 4.19 (d, J = 5.4, 1), 4.50 (d, J = 5.1, 1), 5.36 (d, J = 5.7, 1).
Method H
[0217] To the Boc-protected product lc (P=Boc, R1=Me) (100 mg, 0.176 mmol) in methanol (4 mL) and water (1 mL), O-alkylhydroxylamine hydrochloride (for example, O- methylhydroxylamine hydrochloride) (60 mg, 0.70 mmol) and sodium acetate (57 mg, 0.70 mmol) were added and heated at 80°C for 3 h and then stirred at rt overnight. The solvent was removed under high vacuum to obtain the crude Boc-protected product 4a (P=Boc, R1=H, R7=Me). The crude product 4a was taken in 30% trifluoroacetic acid in dichloroethane (10 mL), dimethylsulfide (0.5 mL) and stirred for 1 h at rt. The solvent was removed and the residue was kept under high vacuum for 1 h and the product 4b (R =Me, R -Me) was taken as such for the next step.
[0218] TLC: Rf= 0.63 (10% MeOH/DCM); MS(ESPOS): 281 [M+H]+. 1H NMR (300 MHz, CD3OD) δ 1.95 (s, 3), 2.08 (s, 3), 3.60 (dd, J = 3.3, 10.20, 1), 3.92 (s, 3), 4.13 (dd, J = 4.8, 10.20, 1), 4.49 (d, J = 1.2, 1), 5.38 (d, J = 5.4, 1). Method I
[0219] To the Boc-protected product lc (P=Boc, R*=Me) (500 mg, 0.88 mmol) in THF (10 mL), tefrabutylammonium fluoride (2.5 mmol, 1 M in THF) was added and the reaction mixture was stirred at rt for 1 h. The solvent was removed and the residue was purified on silica gel column using 5% methanol in dichloromethane as eluent. The product (111 mg, 0.31 mmol) obtained from the column was then taken in a mixture of dichloromethane (3 mL) and pyridine (3 mL) to which acetic anhydride (0.5 mL, 10.6 mmol) and dimethylaminopyridine (80 mg, 1.7 mmol) were added and stirred at rt overnight. The solvent was removed and the crude product was purified on silica gel column using 30% ethyl acetate in hexanes as eluent to provide 5a (P=Boc, R]=Me) (58 mg, 38%).
[0220] TLC: Rf = 0.73 (50% EtOAc/Hexanes); MS(ESPOS): 500 [M+Na]+. 1H NMR (300 MHz, CDC13) δ 1.38 (s, 9), 1.91 (s, 3), 1.98 (s, 3), 2.07 (s, 3), 2.18 (s, 3), 4.33 (m, 1), 4.72 (m, 1), 4.94 (m, 1), 5.21 (m, 2), 5.45 (s, 1), 5.57 (m, 1).
[0221] To product 5a (P=Boc, R1=Me) (158 mg, 0.331 mmol) in DCM (5 mL), dimethylaminosulfurtrifluoride (732 μL, 3.31 mmol) was added and stirred overnight. More DCM was added and the organic portion was washed with sodium bicarbonate. The residue obtained on removal of solvent was purified on silica gel column chromatography using 20% ethyl acetate in hexanes as eluent (100 mg, 60%) to provide the protected product (P=Boc, R1=Me). The Boc-protected product was taken up in 30%> trifluoroacetic acid in dichloroethane and dimethylsulfide and stirred for 1 h at rt. The solvent was removed to provide the product 5b (R1=Me).
[0222] TLC: Rf = 0.63 (40% MeOH/Hexanes); MS(ESPOS): 522 [M+Naf. 1H NMR (300 MHz, CDC13) δ 1.40 (s, 9), 1.69 (t, J = 18.9, 3), 1.98 (s, 3), 2.08 (s, 6), 2.13 (s, 3), 4.22-4.30 (m, 1), 4.53 (dd, J = 10.9, 25.3, 1), 5.16-5.28 (m, 2), 5.52 (s, 1), 5.63 (d, J = 5.2, 1).
Method J
[0223] Enolization (LiHMDS) and alkylation of 7a with 4-bromo-2-methyl-2-butenyl afforded a mixture of diastereomers of the lactam 7b (R9 =2-methyl-2-butenyl) (61%) according to the literature procedure by Zhang, R.; et al., Journal of the American Chemical Society. 1998, 120, 3894-3902. Compound 7a is commercially available from vendors such as Bachem. Alternatively, 7a can be prepared by methods well known in the art for an example see Baldwin, et al.; Tetrahedron, 1989, 45, 7449-7468. [0224] The lactam 7b was reduced to the pyrrolidine 7c (R9'=2-methyl-2-butenyl) (70%) by the two-step sequence involving superhydride reduction of the lactam to the hemiaminal and the subsequent reduction of the hemiaminal with Et3SiH/BF3»OEt . The pyrrolidine 7c (R9 =2- methyl-2-butenyl) (778 mg, 2.08 mmol), 10% palladium on carbon (230 mg), in anhydrous methanol (25 mL) was subjected to Parr hydrogenolysis at 50 psi for 5 h. The reaction mixture was filtered through a celite pad and washed several times with methanol. The combined washings and filtrate were evaporated to dryness, affording, without further purification, a colorless oil 7d (R9=2-methylbutyl).
[0225] TLC: R = 0.3 [Solvent system: DCM:hexanes:MeOH(6:5:l)]. MS(NEGATINE): 284.5 [M-H]".
Method K
[0226] Enolization (LiHMDS, 33 mmol, 33 mL, 1.1 equiv) and alkylation of 7a (9.47 g, 29.7 mmol, 1 equiv) with cis-l-bromo-2-pentene (4.21 mL, 35.6 mmol, 1.2 equiv), in anhydrous THF at -78°C under nitrogen, afforded a mixture of diastereomers of the lactam 7b (R9 =2- pentenyl) (43.2%) after silica gel purification. The lactam 7b (3.96g, 10.22mmol) was reduced to the pyrrolidine 7c (R9 =2-pentenyl) by the two-step sequence involving superhydride reduction of the lactam to the hemiaminal, at -78°C in anhydrous THF, and the subsequent - reduction of the hemiaminal with Et3SiH/BF3OEt2 in anhydrous DCM at -78°C affording 7c (R9'=2-pentenyl) (71%>) after silica gel purification. The pyrrolidine 7c (2.71 g, 7.26 mmol), 10% palladium on carbon (560mg), in anhydrous methanol (30 mL) was subjected to Parr hydrogenolysis at 50 psi for 5 h. The reaction mixture was filtered through a celite pad and washed several times with methanol. The combined washings and filtrate were evaporated to dryness, affording, without further purification, a colorless oil 7d (R9=pentyl) (1.68g, 80%). [0227] TLC: Rf= 0.3 [Solvent system:DCM:hexanes:MeQH(6:5:l)]. MS(ESΝEG): 284.5 [M-H]".
Method L
[0228] Ozonolysis of 7d (R9=2-methylbutyl) in anhydrous dichloromethane followed by treatment with DMS at -78°C afforded aldehyde 8a (77%). 4-Fluorobenzyl phosphonium chloride (0.87 g, 2.13 mmol) and potassium t-butoxide (0.17 g, 1.48 mmol) were suspended in toluene under nitrogen with vigorous stirring. After 4 h, a solution of aldehyde -Sa (204 mg, 0.59 mmol) in toluene (4.6 mL) was added drop-wise. The reaction mixture was stirred at rt for 2 h and diluted with ethyl acetate (50 mL). The organic layer was washed with water (2 x 20 mL), brine, dried and concentrated. The residue was purified by chromatography to give a clear syrup 8b (R9'=3-(4-fluorophenyl)prop-2-enyl) (171 mg).
[0229] To a solution of 8b (R9'=3-(4-fluorophenyl)prop-2-enyl) (171 mg, 0.39 mmol) in MeOH (25 mL) in a Parr bottle was added 10% palladium on carbon (Degussa wet form 50%> w/w water) (200 mg). The bottle was purged and charged with H2 to 40 psi, and shaken for 4 h. The reaction mixture was filtered through celite and rinsed with MeOH. The filtrate was concentrated to give a yellow oil 8c (R9=3-(4-fluorophenyl)propyl) (120 mg).
[0230] MS(ESPOS): 374.5 [M +Na]+, MS(ESNEG): 350.3 [M-H]". Method M
[0231] 4-Chlorobenzyl phosphonium chloride (0.95g, 2.24 mmol, 3.9 equiv) and potassium t-butoxide (0.17 g, 1.55 mmol, 2.7 equiv) were suspended in toluene (7.5 mL) under nitrogen with vigorous stirring. After 4 h, a solution of aldehyde 8a (200 mg, 0.58 mmol, 1 equiv) in toluene (4.9 mL) was added dropwise. The reaction mixture was stirred at rt for 2 h and diluted with ethyl acetate (50 mL). The organic layer was washed with water (2 x 20 mL), brine, dried and concentrated. The residue was purified by chromatography to give a clear syrup 8b (R9 =3- (4-chlorophenyl)prop-2-enyl) (216 mg, 82%).
[0232] MS(ESPOS): 478.5 [M +Na]+, MS(ESNEG): 454.4 [M-H]". [0233] To a solution of 8b (R9'=3-(4-chlorophenyl)prop-2-enyl) (147 mg, 0.32 mmol) in cyclohexane (50 mL) was added 10% palladium on carbon (Degussa wet form 50%» w/w water) (80 mg). The reaction mixture was stirred at rt under 1 atm H2 overnight. The reaction mixture was filtered through celite and rinsed with MeOH. The filtrate was concentrated to give the alkane product 8e (R =3-(4-chlorophenyl)propyl) as a clear oil (131 mg, 89%). To a solution of the alkane (131 mg, 0.29 mmol, 1 equiv) in THF (3 mL) and water (1 mL) was added lithium hydroxide monohydrate (60 mg, 1.43 mmol, 5 equiv). The reaction mixture was stirred at rt overnight. The THF was removed under vacuum. The residue was diluted with water (5 mL) and washed with ether (10 mL). The aqueous layer was taken up in ethyl acetate (60 mL) and partitioned with 10% citric acid (30 mL). The organic layer was washed with water and brine, dried and concentrated to give a clear syrup 8c (R9=3-(4-chlorophenyl)propyl) (105 mg, 100%). [0234] MS(ESPOS): 390.4 [M + Na]+, 268.4 [M - Boc + H]+. Method N
[0235] To a solution of aldehyde 8a (406.5 mg, 1.17 mmol, 1 equiv) in dimethyl acetamide (0.25 mL) at 0 °C was added dibromodifluoromethane (0.21 mL, 2.34 mmol, 2 equiv). To the stirred mixture was added a solution of triphenylphosphine (0.61 g, 2.34 mmol, 2 equiv) in dimethyl acetamide (0.5 mL) over a period of 20 minutes under nitrogen. The reaction mixture was warmed to rt and stirred for 30 minutes, and then was added to an activated zinc (0.25g, 3.82 mmol, 3.3 equiv) with the aid of dimethyl acetamide (0.3 mL). The resulting reaction mixture was stirred at 110°C for 1 h and cooled to rt and filtered with the aid of dimethylacetamide (7 mL). The filtrate was poured into ice water (100 mL) and extracted with ether (150 mL). The ether layer was washed with brine, dried and concentrated. The residue was purified by chromatography to give a clear oil 8b (R9 =3,3-difluoroprop-2-enyl) (182 mg, 41 %).
[0236] MS(ESPOS): 282.4 [M - Boc + H]+.
[0237] To a solution of 8a (R9'=3,3-difluoroprop-2-enyl) (126 mg, 0.33 mmol) in MeOH (35 mL) was added 10% palladium on carbon (Degussa wet form 50% w/w water) (120 mg). The reaction mixture was stirred at rt under hydrogen (1 atm) overnight and was filtered through celite with the aid of MeOH. The filtrate was concentrated to give a clear syrup 8c (R9=3,3- difluoropropyl) (97 mg, 100%).
[0238] MS(ESPOS): 194.4 [M - Boc + H]+, MS(ESNEG): 292.4 [M-H]".
Method O
[0239] To a solution of aldehyde 8b (258 mg, 0.74 mmol, 1 equiv) in THF (3 mL) at 0°C was added tetraallyltin (178 μL, 0.74 mmol, 1 equiv), followed by the drop-wise addition of boron trifluoride etherate (94.3 μL, 0.74 mmol, 1 equiv) over a period of 15 min. The reaction mixture was stirred at 0°C for 1.5 h. Then a solution of potassium fluoride (125 mg) in water (1.25 mL) was added. The resulting mixture was warmed to rt and stirred at rt for 20 min. This was followed by the addition of methanol (10 mL) and the resulting mixture was stirred at rt for another 20 min. The reaction mixture was filtered over celite. The filtrate was evaporated to dryness. The residue was diluted with dichloromethane (100 mL), washed with water (50 mL), dried, concentrated and purified by chromatography to give a clear oil 9a (R9=2-hydroxypent-4- enyl) (261 mg, 90%): MS(ESPOS): 412.5 [M + Naf, 290.4 [M - Boc + H]+. [0240] To a solution of dimethylsulfoxide (0.17 mL, 2.42 mmol, 3 equiv) in dichloromethane (0.5 mL) at -72°C was added a 2 M solution of oxalyl chloride in dichloromethane (0.61 mL, 1.21 mmol, 1.5 equiv) over a period of 1 min. The mixture was stirred at -72°C for 25 min, followed by the drop- wise addition of a solution of the alcohol 9a (314 mg, 0.81 mmol, 1 equiv) in dichloromethane (1.4 mL) over a period of 2 min. The reaction mixture was stirred at -72°C for 25 min, then warmed to -50°C and stirred for an additional 2 h. Triethylamine (0.56 mL, 4.04 mmol, 5 equiv) was added and stirred at -50°C for 25 min. The mixture was diluted with ethyl acetate (100 mL), washed with 5% citric acid (100 mL), water, saturated aqueous NaHC03 and brine, dried, evaporated and coevaporated with anhydrous toluene to give a clear syrup 9b (R9=2-propenylcarboxymethyl) (287 mg, 92%). MS(ESPOS):
288.5 [M - Boc + H]+; MS(ESNEG): 386.2 [M-H]".
[0241] To a solution of ketone 9b (225.1 mg, 0.58 mmol, 1 equiv) in dichloromethane (2 mL) at -78°C was added diethylaminosulfur trifluoride (0.46 mL, 3.49 mL, 6 equiv). The reaction mixture was warmed to rt and stirred at rt for 3 h, followed by an addition of additional (diethylamino)sulfur trifluoride (0.46 mL, 3.49 mL, 6 equiv) at -78°C. The mixture was warmed to rt and stirred overnight. Then the mixture was diluted with dichloromethane (60 mL), washed with sat. aqueous NaHC03 (1 x), brine (1 x), dried, and evaporated. The residue was purified by chromatography to give a yellow oil 9c (X, X=fluoro, flouro) (75 mg, 32%). [0242] MS(ESPOS): 310.5 [M - Boc + H]+.
[0243] To a solution of 9c (R9=2,2-difluoropent-4-enyl) (85 mg, 0.21 mmol) in MeOH (20 mL) was added 10%» palladium on carbon (Degussa wet form 50% w/w water) (100 mg). The reaction mixture was stirred at rt under hydrogen (1 atm) overnight, was filtered through celite with the aid of MeOH (10 mL). To the filtrate was added 10% palladium on carbon (Degussa wet form 50% w/w water) (130 mg). The reaction mixture was stirred at rt under hydrogen (1 atm) overnight, was filtered through celite with the aid of MeOH (10 mL). The filtrate was concentrated to give haloalkyl N-Boc-amino acid 9d (X,X=fluoro, fluoro) (67.7 mg, 100%) as a clear syrup.
[0244] MS(ESPOS): 344.4 [M + Na]+, 222.4 [M - Boc + H]+ MS(ESNEG): 320.2 [M-H]".
Method P
[0245] To 4-propylpyridine (2.5 g, 20 mmol), 30% hydrogen peroxide (2.4 g) was added and refluxed overnight. The solvent was removed and the resulting residue was taken in DCM (30 mL). Trimethylsilyl cyanide (2.6 g, 26 mmol) was added to the above solution followed by dimethylcarbamyl chloride (2.8 g, 26 mmol), and the reaction mixture was stirred at rt overnight. Potassium carbonate (10%, 100 mL) was added. The organic layer was separated, dried over sodium sulfate and then concentrated to obtain 4-propyl-2-cyanopyridine (2.5 g, 93%). It was then refluxed in hydrochloric acid (6N, 60 mL) for overnight. The 4-propyl-2- carboxylic acid pyridine 10b (R9=propyl) was obtained after crystallization from acetonitrile (2g, 71%).
[0246] MS(ESPOS): 166 [M+H]+; 1H NMR (300 MH2, CD3OD) δ 8.75 (dd, J= 9.0, 3.0, 1), 8.42 (s, 1), 8.08 (dd, J = 9.0, 3.0, 1), 3.00 (t, J = 7.5, 2), 1.82 (m, 2), 1.05 (t, J = 7.2, 3).
13c
Method Q
[0247] A mixture of picolinic acid (Aldrich) (20 g, 162 mmol, 1 equiv) and sodium bromide (33.43 g, 325 mmol, 2 equiv) in thionyl chloride (81 mL) was refluxed for 5 h. The solvent was removed under vacuum. Absolute methanol (160 mL) was added and the mixture was stirred at rt for 30 minutes. The solvent was evaporated, and the residue was taken up in 5% sodium bicarbonate and extracted with ethyl acetate (3x). The organic layers were combined and dried over MgS0 and evaporated. The residue was purified by chromatography to give 4- chloropicolinic acid methyl ester as a white solid (19.9 g, 72%): 1H NMR (300 MHz, CDC13) δ 8.63 (d, J = 5.4, 1), 8.13 (d, J = 2.1, 1), 7.48 (dd, J = 2.0, 5.3, 1), 4.00 (s, 3). [0248] A mixture of 4-chloropicolinic acid methyl ester (2.4 g, 14.1 mmol), 57%> hydroiodic acid (13.3 mL) and 50% aqueous hypophosphorous acid (0.66 mL) was stirred at 85 °C for 2 h and then was stirred at 107 °C overnight. The mixture was cooled to 95 °C. At this temperature over 30 minutes 10 M sodium hydroxide aqueous solution (4.2 mL) was added, followed by the addition of water (15.2 mL). The mixture was cooled to rt and stirred at rt for lh. The precipitate was filtered, washed with cold water and dried under high vacuum overnight to give 4-iodopipecolinic acid 13a (3.5 g, 66%): !H NMR (300 MHz, DMSO d6) δ 8.39 (d, J = 5.1, 1), 8.35 (d, J = 1.8, 1), 8.07 (dd, J = 1.7, 5.2, 1); MS (ESPOS): 250.2 [M+H]+. [0249] To a mixture of 7-Me MTL HCl salt 2b (R!=Me, R2=Me) (200 mg, 0.69 mmol, 1 equiv) in dry DMF (1.8 mL) at 0 °C was added triethylamine (0.50 mL, 3.61 mmol, 5.2 equiv), followed by the addition of BSTFA (0.28 mL, 1.04 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. To the reaction mixture was added the acid 13a (341 mg, 0.90 mmol, 1.3 equiv) and HATU (423 mg, 1.11 mmol, 1.6 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with water (1 x), sat. NaHC03 (1 x) and brine. The organic layer was dried over Na2S04 and evaporated to give a yellow residue which was dissolved in methanol (20 mL) to which was added dry Dowex resin (250 mg). The reaction mixture was stirred at rt for 1 h. The resin was removed by filtration and the crude product eluted with 2M ammonia in methanol. The methanolic eluent was evaporated, and the resulting residue was purified by chromatography to provide a white solid 13b (R1=Me, R2= Me, R3=H) (250 mg, 75%): 1H NMR (300 MHz, CD3OD) δ 8.46 (d, J = 1.8, 1), 8.30 (d, J = 5.4, 1), 7.98 (dd, J = 1.8, 5.1, 1), 5.25 (d, J = 6.0, 1), 4.32-4.23(m, 2), 4.09 (dd, J = 5.7, 10.2, 1), 3.87 (d, J = 3.0, 1), 3.54 (dd, J = 3.3, 10.2, 1), 2.24-2.15 (m, 1), 2.11 (s, 3), 0.99-0.96 (m, 6); MS (ESPOS): 483.5 [M+H]+; MS (ESNEG): 481.4 [M-H]".
[0250] To a dry flask was added 13b (Rl=Me, R2= Me, R3=H) (133.9 mg, 0.28 mmol, 1 equiv), triphenylphosphine (46.7 mg, 0.18 mmol, 0.64 equiv), copper (I) iodide (33.9 mg, 0.18 mmol, 0.64 equiv), palladium acetate (20 mg, 0.09 mmol, 0.32 equiv) and triethylamine (1.6 mL). The mixture was deaerated with nitrogen, followed by addition of 3-prop-2-ynyl- cyclopentane (120 mg, 1.11 mmol, 4 equiv). The mixture was stirred at 50 oC overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 13c (Rl=Me, R9'= 3-cyclopentyl-prop-l-ynyl, R2= Me, R3=H) as a yellow solid (106 mg, 83%): IH NMR (300 MHz, CD3OD) δ 8.55 (d, J = 4.8, 1), 7.98 (s, 1), 7.47 (dd, J = 1.7, 5.0, 1), 5.26 (d, J = 5.4, 1), 4.33-4.22 (m, 2), 4.10 (dd, J = 5.5, 10.4, 1), 3.86 (d, J = 3.3, 1), 3.55 (dd, J = 3.3, 10.5, 1), 2.49 (d, J = 6.9, 2), 2.26-2.12 (m, 2), 2.11 (s, 3), 1.93-1.82 (m, 2), 1.73-1.55 (m, 4), 1.43-1.31 (m, 2), 1.00-0.96 (m, 6); MS (ESPOS): 463.6 [M+H]+; MS (ESNEG): 461.5 [M-H]".
14a 14b 14c
Method R
[0251] To a solution of 13a prepared in method Q (5 g, 13.26 mmol) in methanol (500 mL) was added a few drops of cone, sulfuric acid. The reaction mixture was refluxed overnight. The solvent was evaporated and the residue was purified by chromatography to give 4- iodopipecolinic acid methyl ester 14a as a yellow solid (3.0 g, 86%): 1H NMR (300 MHz, CDC13) δ 8.49 (d, J = 1.5, 1), 8.37 (d, J = 5.4, 1), 7.85 (dd, J = 1.6, 5.2, 1), 4.00 (s, 3); MS (ESPOS): 264.3 [M+H]+.
[0252] . To a dry flask were added 14a (1 g, 3.8 mmol, 1 equiv), triphenylphosphine (79.7 > mg, 0.3 mmol, 0.08 equiv), copper (I) iodide (57.9 mg, 0.3 mmol, 0.08 equiv), palladium acetate (34.1 mg, 0.15 mmol, 0.04 equiv) and triethylamine (14 mL). The mixture was deaerated with nitrogen, followed by addition of 3-butyn-l-ol (0.53 g, 7.6 mmol, 2 equiv). The mixture was stirred at rt for 3 h. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 14b (R9 = 3-hydoxy-but-l-ynyl) as a yellow oil (0.78 g, 100%): 1H NMR (300 MHz, CDC13) δ 8.66-8.63 (m, 1), 8.09-8.08 (m, 1), 7.43-7.40 (m, 1), 3.99 (s, 3), 3.88-3.82 (m, 2), 2.72 (t, J = 6.3, 2). MS (ESPOS): 206.4 [M+Hj+. [0253] To a solution of the above 14b (R9 = 3-hydoxy-but-l-ynyl) (0.78 g, 3.8 mmol) in methanol (40 mL) was added 10 % palladium on carbon (0.4 g). The flask containing the reaction mixture was purged and charged with hydrogen (1 atm) and stirred at rt overnight. The palladium was removed by filtration and the filtrate was concentrated to give 14c (R9=3- hydroxybutyl) as an oil (0.77 g, 97%): 1HNMR (300 MHz, CDC13) δ 8.60 (d, J = 4.5, 1), 7.97 (d, J = 1.2, 1), 7.29 (dd, J = 1.6, 5.0, 1), 3.99 (s, 3), 3.67 (t, J = 6.3, 2), 2.72 (t, J = 7.7, 2), 1.81- 1.69 (m, 2), 1.62-1.54 (m, 2); MS (ESPOS): 210.4 [M+H]+.
Method S
[0254] To 4-hydroxypyridine-2-carboxylic acid 10b (R9 = hydroxy) (200 mg, 1.4 mmol) in DMF (2 mL), potassium carbonate (397 mg, 2.8 mmol) was added followed by rø-bromobutane (197 mg, 1.4 mmol), warmed at 60°C for overnight. The solvent was removed to obtain the crude ester product. The crude ester (360 mg, 1.4 mmol) was dissolved in THF (4 mL), lithium hydroxide (72 mg, 1.7 mmol) was added, and the reaction mixture stirred at room temperature for 2 hr. The residue obtained on removal of solvent was purified by silica gel chromatography using 10%) MeOH in DCM to provide 4-butoxypyridine-2-carboxylic acid 15a (R9 = butyl) (100 mg, 43%). 1H NMR (300 MHz, CD3OD) δ 8.37 (d, J=6.0, 1), 7.63 (d, J=2.7, 1), 7.07 (dd, J=2.7, 6.0, 1), 4.15 (t, J=6.6, 2), 1.82 (m, 2), 1.54 (m, 2), 1.01 (t, J=7.5, 3). MS (ESNEG):194 [M-H]". [0255] To 4-butoxypyridine-2-carboxylic acid 15a (R9 =Butyl) (100 mg, 0.5 mmol) in DMF (2 mL), 7-methyl -thiolincosaminide 2b (R1=Me, R2=Me) (147 mg, 0.5 mmol) was added, followed by HBTU (214 mg, 0.55 mmol) and DIEA (132 mg, 1 mmol). The reaction mixture was stirred at room temperature for 2 hr. The solvent was removed, and purification of the crude material was carried out by silica gel column chromatography to obtain compound 15b (R1=Me, R2=Me, R9'= butyl) (201 mg, 91%): 1H NMR (300 MHz, CD3OD) δ 8.42 (m, 1), 7.96 (s, 1), 7.09 (m, 1), 5.27 (d, J=5.4, 1), 4.10-4.87 (m, 3), 3.85 (d, J=3.3, l), 3.76 (m, 1), 2.11 (m, 4), 1.81 (m, 2), 1.49 (m, 4), 0.99 (m, 9). MS (METHOD ES+): 428 [M+H]+. [0256] To a solution of the pyridine 15b (R1=Me, R2=Me, R9'= butyl) (200 mg, 0.46 mmol) in water (10 mL), AcOH (3 mL) and MeOH (2 mL), was added Pt02 (200 mg) and the resulting reaction mixture shaken under 55 psi hydrogen overnight. Residual catalyst was removed by filtration through celite, and the solvent was removed to obtain the crude product. Purification was carried out by silica gel column chromatography using 20% MeOH in DCM to obtain lincosamide analog 1 (R*=Me, R2=Me, R3=H, R9 = butoxy) (12 mg, 6%). 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.4, 1), 4.22 (dd, J=10.2; 3.3, 1), 4.08 (m, 2), 3.81 (d, J=3.0, 1), 3.70 (m, 1), 3.54 (m, 4), 3.43 (m, 2), 2.90 (m, 1), 2.41 (m, 1), 2.19 (m, 1), 2.10 (s, 3) 1.45 (m, 6), 0.92 (m, 9); MS (ESPOS): 435 [M+H]+.
Method T
[0257] Following the general method found in scheme 16, to a solution of β-lactam 16a
(2.92 g, 12.8 mmol) 1 equiv; prepared from benzyl (S)-(-)-4-oxo-2-azetidine-carboxylate (Aldrich) as described by Baldwin et al, Tetrahedron, 1990, 46, 4733 in THF (30 mL) at 0 °C was added a solution of LDA (2.0 M, 14.0 mL, 28.1 mmol, 2.2 equiv) via syringe pump over 20 min. The reaction was stirred at 0 °C for 30 min., crotyl bromide (85%, 2.89 mL, 28.1 mmol, 2.2 equiv) was added dropwise over ca. 1.5 min, and the mixture was stirred for -2 h at 0 °C, and then partitioned between 1.0 M aqueous KHS04 (100 mL) and EtOAc (100 mL). The organic layer was separated and washed with 1.0 M aqueous KHS0 (100 mL), brine (100 mL), dried (MgS04), filtered and concentrated to give 16b (R9 =2-butenyl) 3.65 g (100%) of greenish yellow solid. This material was used without further purification. [0258] MS (ESNEG): 282.2 [M - H]\
[0259] (Trimethylsilyl)diazomethane (2.0 M in Et20, 25.0 mL, 50 mmol, 3.9 equiv) was slowly added to a solution of acid 16b (R9 =2-butenyl) (3.65 g, 12.9 mmol, 1 equiv) in methanol (70 mL) at 0 °C. Solvent was removed under vacuum to give 3.53 g (11.9 mmol, 92%) of the desired ester product as a yellow oil. This material was used in the subsequent reaction without further purification.
[0260] To a solution of alkene 16c (R9 =2-butenyl) (3.53 g, 11.9 mmol, 1 equiv) in EtOAc (40 mL) at 23 °C was added Pd/C (10 wt. %, 482 mg). The reaction vessel was charged with hydrogen (balloon), and the mixture stirred vigorously. After 2.5 h, the reaction mixture was filtered tlirough a pad of Celite. Celite was washed, with EtOAc (200 mL) and the filtrate was concentrated to provide 3.51 g (11.7 mmol, 99%) of 16c (R9=butyl) as a yellow oil. This material was used without further purification. [0261] MS (ESPOS): 300.4 [M+H]+.
[0262] To a solution of N-TBS β-lactam 16c (R9=butyl) (3.51 g, 11.7 mmol, 1 equiv) in THF (50 mL) at 23 °C was added Et3Ν*3HF (0.95 mL, 5.85 mmol, 0.5 equiv). After stirring for 60 min at 23 °C, the reaction mixture was partitioned between 90% saturated brine (150 mL) and EtOAc (200 mL). The organic layer was separated and washed with brine (150 mL), dried (MgS04), filtered and concentrated. The product was purified via flash column chromatography on silica gel using 50% EtOAc in hexane as eluent to give 1.48 g (8.0 mmol, 68%) of 16d (R9=butyl) as a clear oil. [0263] MS (ESPOS): 578.3 [3M+H]+.
[0264] To a solution of β-lactam 16d (R9=butyl) (2.06 g, 11.1 mmol, 1 equiv) in THF (150 mL) at 23 °C was added a solution of LiAlH4 (1.0 M in THF, 22.9 mL, 22.9 mmol, 2.06 equiv) via syringe over the course of 2 min. After stirring for 10 min at 0 °C, the reaction was warmed to 23 °C, stirred for 15 min, and then refluxed for 3 h. The mixture was then cooled to 0 °C and quenched via careful addition of H 0 (1.0 mL), followed by 15% aqueous NaOH (1.0 mL), and then H20 (2.5 mL). The resulting suspension was stirred at 23 °C for 1.5 h, diluted with Et20 (250 mL), and filtered through Celite, washing with Et20 (250 mL). The filtrate was concentrated to furnish 1.42 g of the desired product 16e (R9=butyl) (9.93 mmol, 89%) as a clear oil. The product was used without further purification. [0265] MS (ESPOS): 287.4 [2M+H]+.
[0266] To a solution of amino alcohol 16e (R9=butyl) (1.41 g, 9.86 mmol, 1 equiv) in dichloromethane (50 mL) at 23 °C was added Boc 0 (2.59 g, 11.9 mmol, 1.2 equiv). After stirring for 2 h at 23 °C, the reaction mixture was concentrated. The product was purified via flash column chromatography on silica gel using 33% EtOAc in hexane as eluent to give 1.53 g (6.31 mmol, 64%) 16f (R9=butyl) as a clear oil. [0267] MS (ESPOS): 266.0 [M + Na]+.
[026§] To a solution of NaI04 (8.81 g, 41.2 mmol, 10 equiv) in H20 (60 mL) at 23 °C was added RuCl3°xH20 (350 mg, catalytic amount) followed by a solution of alcohol 16f (R9=butyl) (1.00 g, 4.12 mmol, 1 equiv) in acetone (60 mL). The biphasic mixture was stirred for 30 min at 23 °C, then extracted with EtOAc (250 mL), decanting the organic layer. The aqueous residue was extracted with two further portions of EtOAc (2 x 150 mL). The combined organic extracts were treated with 2-propanol (75 mL) and stirred at 23 °C. After stirring for 2 h tlie mixture was filtered through Celite, washing with EtOAc (300 mL). The filtrate was concentrated to furnish 0.78 g of the desired product 16g (R9=butyl) (3.04 mmol, 74%) as a dark oil. The product was used without further purification. MS (ESPOS): 280.0 [M + Na]+.
Method U [0269] Following the method shown in general scheme 17, to a solution of alcohol 16f
(R9'=2-methyl-2-butenyl) (3.31 g, 13.0 mmol, 1 equiv) in DMF (100 mL) at 23 °C was added imidazole (2.21 g, 32.5 mmol, 2.5 equiv) followed by TBSCl (2.93 g, 19.5 mmol, 1.5 equiv).
The reaction was stirred for 35 min and then quenched with MeOH (2.0 mL). After stirring for
5 min, the resulting mixture was partitioned between Et20 (500 mL) and H20 (400 mL). The organic layer was separated and washed with H20 (400 mL), brine (200 mL), dried (MgS0 ), filtered and concentrated to give 17a (R9"=2-methyl-2-butenyl) 4.13 g (11.2 mmol, 86%) of the desired product as a clear oil.
[0270] MS (ESPOS): 392.4 [M + Na]+.
[0271] A solution of intermediate 17a (R9 =2-methyl-2-butenyl) prepared as described in general method T (2.03 g, 5.50 mmol, 1 equiv) in dichloromethane (80 mL) at -78 °C was treated with ozone (1.2 L/min) introduced via a gas dispersion tube until a blue color was observed (20 min). A stream of oxygen (1.2 L/min) was then passed through the reaction mixture to discharge excess ozone. After 15 min, oxygen flow was ceased and PPh3 (2.16 g,
8.25 mmol, 1.5 equiv) was added. The reaction mixture was stirred at -78 °C for 30 min, then at
0 °C for 15 min, and then warmed to 23 °C. After stirring for 10 min at 23 °C, silica gel was added, and the resulting mixture concentrated to dryness under vacuum to afford a free-flowing powder that was loaded directly onto a silica gel column. Flash column chromatography using
30-33% EtOAc in hexane as eluent gave 1.52 g (4.42 mmol, 80%) of 17b as a clear oil.
[0272] MS (ESPOS): 398.0 [M + MeOH + Na]+.
[0273] To a suspension of Wittig salt (cyclopropylmethyl triphenyl phosphine) (1.216 g,
3.06 mmol, 1.5 equiv) in THF (10 mL) at 0 °C was added a solution of NaHMDS (1.0 M in
THF, 3.06 mL, 3.06 mmol, 1.5 equiv) dropwise via syringe over the course of 1 min. The resulting solution was stirred for 20 min at 0 °C then treated with a solution of aldehyde 17b
(700 mg, 2.04 mmol, 1 equiv) in THF (3.0 mL; 2 x 1.0 mL flush) transferred via canula. After
15 min at 0 °C the reaction was warmed to 23 °C, stirred for a further 10 min then quenched with saturated NH4CI (30 mL). The resulting mixture was partitioned between Et20 (120 mL) and
H20 (50 mL). The organic layer was separated and washed with brine (50 mL), dried (MgSQ4) filtered and concentrated. Flash column chromatography using 10%) EtOAc in hexane as eluent gave 588 mg (1.54 mmol, 76%) of 17c (R9'=2-(3-cyclopropyl)-propen-l-yl) as a clear oil. [0274] MS (ESPOS): 404.3 [M + Na]+.
[0275] To a solution of TBS ether 17c (R9'=2-(3-cycloρropyl)-propen-l-yl) (190 mg, 0.50 mmol, 1 equiv) in THF (10 mL) at 23 °C was added a solution of TBAF (1.0 M in THF, 0.55 mL, 0.55 mmol, 1.1 equiv). The resulting solution was stirred for 40 min at 23 °C then partitioned between Et 0 (50 mL) and H 0 (50 mL). The organic layer was separated and washed with brine (50 mL), dried (MgSO4), filtered and concentrated to give 133 mg (0.50 mmol, 100%)) of 17c (R9 =2-(3-cyclopropyι)-propen-l-yl) as a clear oil. [0276] MS (ESPOS): 290.2 [M + Naf.
[0277] To a solution of alkene 17c (R9'=2-(3-cyclopropyl)-propen-l-yl) (191 mg, 0.50 mmol, 1 equiv) in dioxane (5.0 mL) at 23 °C was added dipotassium azodicarboxylate (973 mg, 5.01 mmol, 10 equiv) followed by slow addition of a solution of AcOH (573 μL, 10.0 mmol, 20 equiv) in dioxane (5.0 mL) over the course of 16 h via syringe pump. Following the completion of the addition the reaction was stirred a further 6 h then filtered through a glass frit with the aid of Et O (150 mL) to remove precipitate. The resulting solution was washed with saturated aqueous NaHCO3 (2 x 100 mL), brine (80 mL), dried (MgS04), filtered and concentrated. The above procedure was repeated three times on the crude material obtained to give complete conversion of the alkene, providing 183 mg (0.48 mmol, 96%) of the saturated product 17d (R =3 -cyclopropyl-propyl) as a clear oil. [0278] MS (ESPOS): 406.0 [M + Na]+. Method V [0279] Following the general method in Scheme 18, to a solution of the compound 1 hydrochloride (wherein R1=R2 =Me, R3=H, R6=H, R9=4-cr-s-n-Pr, and m=2) (4.00 g, 9.90 mmol, 1 equiv) in THF (70 mL) at 23 °C was added H20 (70 mL) followed by KHC03 (1.29 g, 12.9 mmol, 1.3 equiv) followed by (Boc)20 (2.81 g, 12.9 mmol, 1.3 equiv). After stirring for 5 h, the reaction mixture was partitioned between brine (200 mL) and EtOAc (300 mL). The organic layer was separated and washed with brine (150 mL), and dried (MgS04). Solvent was removed under vacuum and the crude product purified using Biotage® column chromatography system (40+M cartridge, 40 mm ID x 150 mm) using a linear gradient (75% EtOAc/hexanes-100% EtOAc) over 1.2 L total eluent at 50 mL/min to give 4.49 g of the pure carbamate 18a (wherein R1=R2 = e, R3=H, R9=4-cw-n-Pr, and m=2) (8.91 mmol, 90%). [0280] To a solution of carbamate 18a (wherein R =R2 =Me, R3=H, R9=4-c/s-n-Pr, and m=2) (7.99 g, 15.9 mmol, 1 equiv) in benzene (300 mL) at 23 °C was added -anisaldehyde dimethyl acetal (4.06 mL, 23.8 mmol, 1.5 equiv), followed by PPTS (199 mg, 0.79 mmol, 0.05 equiv). The reaction mixture was heated to reflux. After 4 h a second portion of j-?-anisaldehyde dimethyl acetal (2.0 mL, 11.7 mmol, 0.74 equiv) was added. After a further 17 h a third portion of p-anisaldehyde dimethyl acetal (2.0 mL, 11.7 mmol, 0.74 equiv) was added. Following the final addition the reaction was refluxed a further 3 h then cooled to 23 °C and partitioned between EtOAc (300 mL) and H20 (300 mL). The organic layer was washed with 50% saturated aqueous NaHC03 (300 mL), brine (150 mL), dried (MgS0 ), filtered and concentrated. The crude product was purified via silica gel flash column chromatography using 40% EtOAc in hexane as eluent to give acetal 18b (wherein R1=R2 =Me, R3=H, R^-cz'-y-n-Pr, and m=2) 7.00 g (11.3 mmol, 71%) of the desired product as a white foam (1:1 ratio of diastereomers). A small portion of the product was purified on Biotage® column chromatography system (40+S cartridge, 40 mm ID x 75 mm) using a linear gradient (5% EtOAc/hexanes-90% EtOAc hexanes) over 1.2 L total eluent at 50 mL/min to separate the two diastereomers for characterization.
[0281] Diastereomer with high Rf: MS (ESPOS): 623.0 [M+H]+. [0282] Diastereomer with low Rf: MS (ESPOS): 623.0 [M+H]+.
[0283] To a solution of alcohol 18b (wherein R1=R2 =Me, R3=H, R9=4-ci.s-n-Pr, and m=2) (3.00 g, 4.82 mmol, 1 equiv) in trimethyl phosphate (60 mL) at 0 °C was added pyridine (3.90 mL, 48.2 mmol, 10 equiv), followed by POCl3 (0.88 mL, 9.65 mmol, 2 equiv) added over the course of 60 sec. Following the addition, the reaction was maintained at 0 °C for 2 h, then triethylammonium bicarbonate buffer (1.0 M, pH 8.5, 40 mL) was added carefully to quench the reaction. H 0 (60 mL) was then added, and the resulting mixture was stirred at 0 °C for 30 min then warmed to 23 °C. After stirring the quenched reaction mixture for 2 h at 23 °C, volatiles were removed in vacuo with aid of gentle heating in water bath (40-45 °C). The resulting crude product was azeotropically dried by co-evaporation with DMF (3 x 100 mL), then toluene (150 mL, bath temperature=40^1-5 °C) to provide 9.4 g of white solid. The crude product 18c (wherein and m=2) was substantially contaminated with triethylammonium salts, but was carried forward without purification. [0284] MS (ESNEG): 701.2 [M - H]\
[0285] To a solution of the protected phosphate 18c (wherein R1=R2 =Me, R3=H, R9=4-cw- n-propyl, R11=PO(OH)2 and m=2) prepared as described above (9.4 g, crude from previous step, approximately 4.8 mmol) in 1,2-dichloroethane (600 mL) at 0 °C was added H20 (25 mL) followed by TFA (200 mL). Following the additions, the reaction was maintained at 0 °C for 5 min then warmed to 23 °C. After stirring for 25 min at 23 °C, volatiles were removed in vacuo to give 16.2 g of oil. The crude product was dissolved in 1:1 H 0/MeOH (70 mL), filtered and the resulting solution was purified by preparative HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 40 mm ID x 200 mm, 5-60% acetonitrile in H20 w/ 0.1% AcOH over 30 min, 75 mL/min flow rate) to give 1.497 g of the desired phosphate 5 (wherein R =R =Me, R3=H, (3.10 mmol, 64% from free alcohol) as a white solid.
Method W
[0286] Following the general method outlined in Scheme 19, to amino acid 19a (R12=H)
(Aldrich) (1.0 g, 8.7 mmol) suspended in MeOH (5 mL) and 2,2-dimethoxypropane (15 mL) was added concentrated HCl (1.0 mL), and the mixture was stirred 12 h at r.t.. Solvents were removed under vacuum, and the residue was triturated with Et20 and co-evaporated from dry toluene to furnish the crude methyl ester as an off-white solid which was used without further purification.
[0287] To a suspension of crude L-allylglycine methylester in dichloroethane (32 mL) at 0 °C was added 2,4,6-collidine (2.3 mL, 19.1 mmol, 2.2 equiv) and solid 2-nitrobenzenesulfonyl chloride. The reaction was stirred for 3 h at r.t. The solvent was removed under vacuum and the residue was distributed between EtOAc (200mL) and sat. aqueous NH4C1. The organic layer was washed with 1.0 M aq. KHS0 , sat. aq. NaHCQ3, brine, and dried (MgS04), and concentrated to give a residue that was purified by column chromatography on silica (gradient 10 to 20% EtOAc/hexanes) to give the desired product 19b (R12=H) 0.70 g (26 %) as a yellow oil.
[0288] 1H NMR (300 MHz, CDC13) δ 8.10-8.06 (m, 1), 7.95-7.92 (m, 1), 7.76-7.73 (m, 2), 6.08 (d, J=8.2, 1), 5.74-5.60 (m, 1), 5.17-5.12 (m, 2), 4.33-4.26 (m, 1), 3.52 (s, 3), 2.58 (dd, J=6.0, 6.0, 1), 3.44-3.30 (m, 2), 2.25-2.10 (m, 2), 2.11 (s, 3), 2.00-1.88 (m, 1), 1.86-1.70 (m, 1), 1.44-1.25 (m, 6), 0.98-0.88 (m, 9 H). MS (ESNEG): 313.0 [M - H]". [0289] To a stirred suspension of sulfonamide 19b (R12=H) (685 mg, 2.18 mmol), Cs2C03 (710 mg, 2.18 mmol), and tefrabutylammonium bromide (702 mg, 2.18 mmol), in DMF (5.0 mL) was added a solution of 3-methylenehex-l-yl-toluenesulfonate (702 mg, 2.61 mmol; prepared as described by Kelvin H. Yong et al. Journal of Organic Chemistry, 2001, 66, 8248) in DMF (1.0 mL), the reaction mixture was heated to 60 °C overnight. The reaction solvent was removed by evaporation, the resulting residue taken up in EtOAc and washed with 10% aqueous citric acid and brine, the organic phase was dried over MgS0 concentrated to give a residue that was purified by column chromatography on silica (17%-20% EtOAc/hexanes) to give the desired product 19c (R12=H, RI3=proρyl), (0.38g, 42 %) as an oil. [0290] MS (ESPOS): 433 [M + Na]+.
[0291] To a solution of 19c (R12=H, R13=propyl) (0.38 g, 0.92 mmol) in anhydrous DCM (40 mL) was added benzylidene[l,3-bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene] dichloro- (tricyclohexylphosphine)ruthenium (23.3 mg, 0.0276 mmol). The resulting reaction mixture was refluxed under N2 for 2.5 hrs, cooled to room temperature and concentrated. The product was purified by flash column chromatography on silica gel (35% ethyl acetate/hexanes) to give the desired compound 19d (R12=H, R13=propyl) (0.29 g, 81 %). [0292] MS (ESPOS): 383 [M + Na .
[0293] To a stirred solution of thiophenol (183 μL, 1.79 mmol) and 7-methyl-l,5,7- triazabicyclo-[4,4,0] dec-5-ene (214 μL, 1.49 mmol) in anhydrous DMF (3 mL) was added a solution of alkene 19d (R12=H, R13=propyl) (228 mg, 0.596 mmol) in anhydrous DMF (3.0 mL) via a canula. A color change to yellow was observed upon the mixing of the solutions, the resulting reaction mixture was stirred under N2 for one hour then concentrated to a residue. The residue was taken up in ether, stirred with IN aqueous HCl (15.0 mL) for 5 min. The aqueous phase was washed with ether then made basic with solid potassium carbonate. The resulting basic aqueous phase was extracted with ether three times. The combined organic layer was washed with brine, dried with anhydrous sodium sulfate, concentrated, cooled to 0 °C, treated with 2M HCl in ether (0.8 mL), and the resulting mixture was stirred for 5 min and then evaporated to dryness to give the desired product 19d (R12=H, R13=propyl) as the hydrochloride salt (144 mg, 103%)). [0294] MS (ESPOS): 198 [M+H]+.
[0295] To a solution of amine 19d (R12=H, R13=propyl) (143 mg, 0.61 mmol) in anhydrous dichrolomethane (2.0 mL) was added triethylamine (170 μL, 1.22 mmol) and di-t- butyldicarbonate (350 mg, 1.6 mmol). The resulting reaction mixture was stirred over night at room temperature under N2 then evaporated to dryness and purified by flash column chromatography on silica gel using 20% ethylacetate in hexanes as an eluent to give the desired compound 19e (R12=H, R13=proρyl) (176 mg, 86%). [0296] MS (ESPOS): 320 [M + Na . [0297] To a solution of ester 19e (R12=H, RI3=propyl) (175 mg, 0.59 mmol) in dioxane/water (6:1) (4 mL) was added 1 M aqueous lithium hydroxide (0.65 mL, 0.648 mmol). The resulting reaction mixture was stirred over night at room temperature under N2 and the solvent was removed under reduced pressure. The residue was taken up in water washed with ether. The aqueous layer was acidified with 10% citric acid and extracted with ether. The organic layer was washed with brine, dried with sodium sulfate, and evaporated to dryness to give the desired protected cyclic amino acid 19f (R12=H, R13=propyl) (175 mg, 105 %). [0298] MS (ESNEG): 292 [M-H]".
Method X
[0299] As shown in Scheme 20 below, suitable amino esters 20g n=l,2 may be appended with psuedoephedrine which serves as a chiral auxiliary, allowing stereospecifϊc alkylation of the α carbon with a suitable ally lie bromide 20e. Protection of the secondary amine followed by olefin metathesis and cleavage of the chiral auxiliary leads to 4,5 unsaturated N-protected cyclic amino acids 201. Reduction of the 2-carboxylic acid to a hydroxymethyl allows stereospecific tnms-reduction of the 4,5 unsaturation in which leads to 20n. Re-oxidation of the hydroxymethyl to a carboxylic acid in which R12 or R 3 may be R9 as defined in formula I.
20a 20b 20c 20d 20e
20f 20g 20h
20m 20n 20o Scheme 20. General stereospecific synthesis of /r -TO-alkylpiperidine-2-carboxylic acids (n=l) and trα?2-s,-alkylazapine-2-carboxylic acids (n=2). [0300] The following general reaction scheme may be used in the preparation of trans- alkylpiperidine-2-carboxylic acids and tr rø-alkylazapine-2-carboxylic acids: (a) NaH, DMF; R-Br, 65 °C; (b) 1 M aqueous KOH, EtOH, 80 °C; (c) CH20, piperidine, EtOH, 80 °C; (d) DIBALH, CH2C12, -50 °C; (e) PBr3, pyridine, Et20, 0 °C; (f) Ethyl bromoacetate, Et20, 23 °C; (g) 4.0 M HCl in dioxane, Et2O, hexane, 23 °C; (h), (lR,2R)-(-)-Pseudoephedrine, t-BuOLi, THF, 23 °C; (i) LiHMDS, LiCl, THF, 0 °C; (j) (Boc)2O, Et3N, CH2C12, 23 °C; (k) Benzylidene[l,3-bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene]dichloro- (tricyclohexylphosphine)ruthenium, CH2C12, 23 °C; (1) 1 M Aq. NaOH, MeOH, 70 °C; (m) (i) Isobutyl chloroformate, 4-methylmorpholine, DME, -15 °C, (ii) NaBHj, H20, 0 °C; (n) H2, IrCODPyPCy3.PF6, CH2C12, 23 °C; (o) RuCl3.xH20, NaI04, acetone, H20, 23 °C. [0301] As shown in Scheme 20, suitable amino esters 20g n=l,2 may be appended with psudoephedrine which serves as a chiral auxiliary, allowing stereospecific alkylation of the α carbon with a suitable allylic bromide 20e. Protection of the secondary amine followed by olefin metathesis and cleavage of the chiral auxiliary leads to 4,5 unsaturated N-protected cyclic amino acids 201. Reduction of the 2-carboxylic acid to a hydroxymethyl allows stereospecific trans- reduction of the 4,5 unsaturation in which leads to 20n. Re-oxidation of the hydroxymethyl to a carboxylic acid in which R12 or R13 may be R9 as defined in general structure.
Method Y
[0302] Following the general reaction scheme 21, a rapidly stirred solution of 4-oxo- piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 21a (m=2, P=H, P2=Boc) (16.0 g, 0.066 mol) (prepared by the method described by Bousquet et al. Tetrahedron, 1997, 53, 15671) in DMF (200 mL) was treated with solid cesium carbonate (10.7 g, 0.033 mol) and methyl iodide (4.5 mL, 0.072 mol). The reaction mixture was stirred 5h, diluted with EtOAc and extracted with sat. aq. sodium bicarbonate, 10% aq. citric acid and brine, the organic layer was separated and dried over sodium sulfate, frittered and evaporated to dryness. The product obtained on removal of solvent was azeotropically dried by evaporation from dry benzene to afford 14.8 g (98%) of the desired product 4-Oxo-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester 21a (m=2, P=Me, P2= Boc) as an oil: TLC Rf 0.53 (Hexanes/EtOAc, 1 : 1); 1H NMR (300 MHz, CDC13) δ 5.33 (broad m, 0.5) rotamer, 5.06 (broad m, 0.5) rotamer, 4.31-4.19 (m, 1), 3.95 (s, 3), 3.95-3.70 (m, 1), 3.16-2.97 (m, 2), 2.71 (m, 2), 1.68 (broad s, 9).
[0303] A O C stirred solution of 4-oxo-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 2- methyl ester 21a (m=2, P=Me, P =Boc) (5.17 g, 0.02 mol) in DCM (60 mL) was treated with tetraallyltin (Aldrich) (5.3 mL, 0.022 mol) followed by dropwise addition of BF3®OEt2 (2.5 mL, 0.02 mol). The reaction mixture was stirred Ih, then aq. IM potassium fluoride (38.0 mL) and celite (5 g) was added and the reaction mixture was stirred 3h. The reaction mixture was filtered and concentrated to dryness, the residue was dissolved in DCM and washed with water and brine, dried over MgS04 and evaporated to dryness. The residue obtained was purified by silica gel column chromatography (DCM 100% to DCM: acetone 9:1) to afford 3.85 g (64%) of the desired product 4-allyl-4-hydroxy-piperidine-l,2-dicarboxylicacid 1-tert-butyl ester 2-methyl ester 21b (m=2, P=Me, P2=Boc, R9'=allyl) as an oil. [0304] 1H NMR (300 MHz, CDC13) δ 6.11-5.97 (m, 1), 5.42-5.32 (m, 2), 5.06 (broad d, J = 6.0, 0.5) rotamer, 4.87 (broad d, J = 6.0, 0.5) rotamer, 4.18-4.03 (m, 1), 3.93 (s, 3), 2.48-2.37 (m, 2), 1.98-1.43 (m, 11); MS (ESPOS): 322.0 [M+Naf.
[0305] A stirred suspension of 21b (m=2, P=Me, P2= Boc, R9'=allyl) (3.80 mL, 1.27 mmol) and 10% Pd/C (degusa wet form 50% w/w) (1.35 g, 1.3 mmol) in MeOH (80 mL) was stirred 6h under 1 atm hydrogen. The reaction mixture was filtered through celite and evaporated to dryness and dried azeotropically by evaporation from toluene the residue obtained (3.15 g) was used in the next step without further purification.
[0306] To a stirred -78°C solution of DAST (1.7 mL, 1.3 mmol) in DCM (50 mL) was added 4-Hydroxy-4-propyl-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester in DCM (30 mL). The reaction mixture was then stirred for lh, then allowed to warm to -40°C for 5h. Additional DAST (0.4 mL) was added and the reaction mixture was stirred an additional 2h, sat. aq. K2C03 (20 mL), and water (60 mL) were added followed by diethyl ether (500mL). The organic layer was separated, washed with brine, dried over sodium sulfate and evaporated to dryness. The resulting crude fluorinated product was purified by silica gel column chromatography (hexanes-EtO Ac 9:1). The residue obtained by chromatographic purification was dissolved in dioxane (65 mL) and water (26 mL), cooled to 0°C, and treated with Os04 ■ (0.65 mL, 4% aq. solution) and 30% H202 (10 mL). The reaction mixture was stirred overnight and concentrated to dryness. The residue was dissolved in DCM and the organic layer washed with water (100 mL), 25% aq. Na2S03 (2x100 mL), and brine (100 mL), dried over Na2S04 and evaporated to dryness. The residue obtained was purified by silica gel column chromatography (hexanes-EtOAc 9:1) to afford (1.08 g, 34 %) of the desired product 4-fluoro-4-propyl- piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester 21c (πrf, P=Me, P2= Boc, R9= -propyl) as an oil.
[0307] 1H NMR (300 MHz, CDC13) δ 4.59 (dd, J=6.0, 6.0, 1), 3.82-3.69 (m, 1), 3.74 (s, 3), 3.28 (m, 1), 3.29-2.04 (m, 2), 1.91-1.71 (m, 3), 1.60-1.31 (m, 6), 1.45 (s, 9), 0.92 (t, J=7.1, 3); MS (ESPOS): 204.1 (M+H-Boc), 326.3 [M+Naf.
[0308] A stirred solution of 21c (m=2, P2= Boc, R9=n-propyl) (1.06 g, 3.47 mmol) in dioxane-water 6:1 (42 mL) was treated with 1.0 M aq. lithium hydroxide (5.3 mL, 5.3 mmol). The reaction mixture was stirred at room temperature overnight, then treated with additional 1.0 M aq. lithium hydroxide (1.5 mL) and the resulting reaction mixture was stirred 5h. The reaction solvent was removed, the residue was taken up in ethyl acetate (500 mL), washed with 10%) Citric acid and brine and dried over MgS0 . Concentration of organic solution to afford the desired product 4-fluoro-4-propyl-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester 21d
(m=2, P2=Boc, R9=«-propyl) (0.88 g, 87 %) as a white solid.
[0309] 1H NMR (300 MHz, CDC13) δ 4.59 (dd, J=6.0, 6.0, 1), 3.82-3.69 (m, 1), 3.74 (s, 3),
3.28 (m, 1), 3.29-2.04 (m, 2), 1.91-1.71 (m, 3), 1.60-1.31 (m, 6), 1.45 (s, 9), 0.92 (t, J=7.1, 3);
MS (ESNEG): 288.4 [M-H]".
[0310] Example 1 Preparation of 4-Ethyl-piperidme-2-carboxylic acid [2-methyI-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0311] 4-Ethyl pyridine-2-carboxylic acid HCl salt (Toronto) (117 mg, 0.64 mmol) was suspended in dry acetonitrile (4 mL). Triethylamine (180 μL, 1.28 mmol) was added and the reaction mixture was cooled to 0°C. Isobutyl chloroformate (129 μL, 0.62 mmol) was added and the reaction mixture was warmed to 4°C. After 1.5 h the activated ester solution was transferred to a solution of 2b (R1=Me, R2=Me), prepared as in Method C, in 1 :1 acetone/water (2 mL) and warmed to 30°C to dissolve. Triethylamine (80 μL, 0.057 mmol) was then added to the reaction mixture. The reaction mixture was stirred for 10 h at rt, then evaporated to dryness and chromatographed on silica 94:5 dichloromethane:0.25% ammoniain methanol to provide lib (R!=Me, R =Me R3=H, R9=ethyl) (167 mg 69.7%). MS (ESPOS): 385.2 [M+H]+. [0312] A solution of pyridine lib (m=2, R =Me, R2=Me R3=H, R9=ethyl) (167 mg, 0.435 mmol) in 3:2 methanol/water (20 mL) was added to platinum(ιN)oxide (339 mg, 0.521 mmol) in a Parr bottle. Concentrated HCl (52 μL, 0.52 mmol) was then added. The bottle was purged, and charged with H to 65 psi and shaken for 24 h. The reaction mixture was filtered through celite and rinsed with methanol. The combined filtrate was evaporated to dryness and chromatographed on silica 88:12 to 80:20 dichloromethane: 0.25% ammoniain methanol to give 43 mg of a high Rf product and 49 mg of a mixed fraction. Chromatography of the low Rf fraction on fluorosil 84:16 to 80:20 dichloromethane: 0.25% ammoniain methanol provided 4- ethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro- pyran-2-yl)-propyl]-amide (21.9 mg, 12.9%), which was taken up in 1:1 acetonitrile: water (50 mL), 0.2μ millipore filtered, and cooled to 0°C. IN HCl (67 μL) in water (20 mL) was added and re-lyophilized to provide the HCl salt (24.0 mg) as a colorless powder. [0313] 1H NMR (300 MHz, D20) δ 5.32 (d, J = 5.8, 1), 4.14-4.06 (m, 1), 4.12 (s, 2), 3.85 (d, J = 3.30, 1), 3.60 (dd, J = 3.3, 10.4, 1), 3.30 (dd, J = 2.5, 11.8, 1), 3.09 (m, 1), 2.56 (ddd, J = 2.8, 12.9, 15.7, 1), 2.14 (s, 3), 2.14-2.05 (m, 1), 1.96-1.90 (m, 1), 1.74-1.69 (m, 1), 1.45-1.35 (m, 1), 1.33-1.23 (m, 2), 1.08-0.98 (m, 2), 0.86 (m, 9); MS(ESPOS): 391.4 [M+H]+, 803.5.4 [2M+Na], (ESNEG): 389.5 [M-H]".
Example 2 Preparation of l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0314] 4-n-Propylhygric acid prepared by the method of Hoeksema, H. et. al. Journal of the American Chemical Society, 1967, 89 2448-2452 (157 mg, 0.76 mmol) was suspended in dry acetonitrile (5 mL). Triethylamine (421 μL, 3.02 mmol) was added and the reaction mixture was cooled to 0°C. Isobutyl chlorofoπnate (98 μL, 0.76 mmol) was added and after 10 min the reaction was allowed to warm to 4°C. After 1.5 h a solution of 2b (R =Me, R =Me), from Method C (190 mg, 0.76 mmol) in 1 :1 acetone:water (5 mL) was added and the reaction mixture was stirred for lOh at rt. The reaction mixture was evaporated to dryness and chromatographed on silica 94:6 dichloromethane:0.25% ammoniain methanol. Fractions 14-18 contained the product as a colorless oil (50.2 mg, 16.5%).
[0315] 1H NMR (300 MHz, D2O) δ 5.33 (d, J = 6.0, 1), 4.27-4.22 (m, 1), 4.18 (s, 1), 4.09 (dd, J = 5.8, 10.2, 1), 3.92-3.81 (m, 1), 3.92-3.81 (m, 1), 3.64-3.59 (m, 1), 2.92 (s, 3), 2.92-2.85 (m, 1), 2.35-2.28 (m, 3), 2.13 (s, 3), 1.46-1.41 (m, 2), 1.40-1.28 (m, 2), 0.89-0.84 (m, 9); MS(ESPOS): 405.5 [M+H]+.
Example 3 > Preparation of l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [3-cyano-2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0316] Lincosamine 2b (R1=Me, R2=CH2CN) (54.2 mg, 0.20 mmol) prepared by Method E was dissolved in DMF (0.7 mL). The reaction mixture was cooled to 0°C and triethylamine (170 μL, 1.2 mmol) and BSTFA (96 μL, 0.36 mmol) was added. The reaction mixture was allowed to warm to rt, and stirred at rt for 1 h. 4-n-Propylhygric acid prepared by the method of Hoeksema, et al, J. Am. Chem. Soc, 1967, 892448-2452 (66.4 mg, 0.32 mmol) and HATU (149 mg, 0.39 mmol) were added, and the mixture was stirred at rt for 3 h. DMF was removed and the residue was dissolved in DCM (100 mL), washed with saturated NaHCO3 (30 mL) and brine (30 mL), and dried over sodium sulfate. The residue obtained by removing the solvent was dissolved in methanol (20 mL) and treated with Dowex resin H " (300 mg) for 15 min. The crude product was eluted from the resin by washing with 5% TEA in MeOH (25 mL x 15 min x 2) and 5% TEA in MeCN (25 mL x 15 min). The combined eluent was evaporated to dryness and purified by silica gel column chromatography using 7% 0.25M NH3 in methanol in dichloromethane as the eluent (24 mg, 28%).
[0317] 1H NMR (300 MHz, D20) δ 5.61 (d, J = 5.8, 1), 4.59 (d, J = 10.2, 1), 4.46 (d, J = 10.2, 1), 4.46 (dd, J = 6.0, 10.4, 1), 4.05 (d, J = 3.0, 1), 3.84 (dd, J = 3.3, 10.4, 1), 3.48 (dd, J = 5.8, 8.0, 1), 3.34 (dd, J = 5.2, 10.2, 1) 2.81-2.61 (m, 2), 2.65 (s, 3), 2.43 (s, 3), 2.31-2.10 (m, 2), 1.32 (d, J = 6.0, 1), 1.18 (t, J =7.1, 3); MS(ESPOS): 430.5 [M+H]+ MS(ESPOS): 428.5 [M-H]". Example 4 Preparation of 4-Ethyl-piperidine-2-carboxylic acid [2-hydroxy-2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0318] Lincosamine 3b (R1=Me, R -Me) prepared by Method F (54.2 mg, 0.20 mmol) was dissolved in DMF (1.0 mL). The reaction mixture was cooled to 0°C and triethylamine (178 μL, 1.3 mmol) and BSTFA (85 μL, 0.32 mmol) were added. The reaction mixture was allowed to warm to rt, and stirred at for 1 h. 4-Ethyl pyridine-2-carboxylic acid HCl salt (Toronto) (55.3 mg, 0.29 mmol) and HATU (122 mg, 0.32 mmol) were added, and the mixture was stirred at rt for 3 h. DMF was removed and the residue was dissolved in THF (10 mL), and treated with (600 mg) Amberlite A-26 F" form resin and catalytic TBAF for 5h. The crude product was obtained by removal of the resin and evaporation of the solvent to dryness and purified by silica gel column chromatography using 10% 0.25M NH3 in methanol in dichloromethane as the eluent to provide the pyridine product lib (m=2, R1==Me, R2=Me R3=OH, R9=ethyl) (26 mg, 33%).
[0319] MS(ESNEG): 399.5 [M-H]".
[0320] A solution of pyridine lib (m=2, R =Me, R2=Me R3=OH, R9=ethyl) (26 mg, 0.065 mmol) in 3:2 methanol: water (10 mL) was added to platinum(-[N)oxide (51 mg) in a Parr bottle. Concentrated HCl (6.0 μL, 0.072 mmol) was then added. The bottle was purged and charged with H to 65 psi and shaken for 24 h. The reaction mixture was filtered through celite and rinsed with methanol. The combined filtrate was evaporated to dryness and chromatographed on silica 80:20 dichloromethane: 0.25% ammoniain methanol to give a high Rf product and the title compound (5.8 mg, 21.8%).
[0321] 1H ΝMR (300 MHz, D2O) δ 5.37 (d, J = 6.0, 1), 4.41 (d, J = 9.6, 1), 4.32 (d, J = 9.3, 1), 4.08 (dd, J = 6.6, 11.0, 1), 3.93-3.90 (m, 2), 3.59 (dd, J = 3.0, 10.7, 1), 3.93-3.90 (m, 2), 3.04 (ap dt, J = 7.1, 14.6, 14.6, 1), 2.24-2.18 (m, 1), 2.20 (s, 3), 1.70-1.60 (m, 1), 1.42-1.13 (m, 1), 0.88 (dd, J =6.0, 7.4, 1); MS(ESPOS): 407.4 [M+H]+. Example 5
Preparation of l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2-hydroxyimino-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0322] Triethylamine (0.041 mL, 0.28 mmol) and BSTFA (0.24 mL, 0.94 mmol) were added to the crude oxime 4b (R1=Me, R7=H) prepared by Method G (50 mg, 0.19 mmol) in DMF (3 mL) at 0°C and the mixture was stirred at rt overnight. Next, 4-n-propylhygric acid (63 mg, 0.37 mmol) and HATU (142 mg, 0.37 mmol) were added and the mixture was stirred at rt for 4 h. DMF was removed and the residue was extracted with dichloromethane (100 mL) and washed with saturated bicarbonate (20 mL) and brine (20 mL). The residue obtained on removal of dichloromethane was then treated with 10% TFA in dichloroethane (10 mL) and dimethyl sulfide (0.5 mL) for 1 h. The solvent was then removed to obtain the crude product, which was purified by silica gel column chromatography using 20% methanol in dichloromethane as the eluent to provide the title compound (20 mg, 25%).
[0323] TLC: Rf = 0.67 (20% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 0.93 (t, J = 6.8, 3), 1.31-1.44 (m, 4), 1.88 (s, 3), 1.99 (s, 3), 2.09 (m, 2), 2.11 (m, 1), 2.62-2.98 (m, 3), 2.76 (s, 3), 3.60 (m, 2), 4.10 (dd, J = 5.7, 10.20, 1), 4.27 (d, J = 9.6, 1), 5.23 (d, J = 5.5, 1), MS(ESPOS): 420 [M+H]+.
Example 6
Preparation of l-Methyl-4-propyl-pyrrolidine-2-carbo-sylic acid [2-metho-j£yimino-l-(394,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide [0324] From crude oxime product 4b prepared by Method H (R1=Me, R7=Me), the title compound was prepared as in Example 5 (10 mg, 47%).
[0325] TLC: Rf = 0.55 (10% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 0.91 (m, 3), 1.32 (m, 4), 1.88 (s, 3), 1.98 (s, 3), 1.78-2.04 (m, 2), 2.34 (s, 3), 2.90 (dd, J = 5.1, 6.30, 8.10 1), 3.21 (dd, J = 6.3, 10.2, 1), 3.57 (dd, J = 3.3, 10.2, 1), 4.23 (dd, J = 5.4, 10.2, 1), 5.25 (d, J = 5.7, 1); MS(ESPOS): 434 [M+H]+.
Example 7 Preparation of 5-Bu yl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0326] Lincosamine intermediate 2b (R1=Me, R2=Me), prepared by Method C, was dissolved in DMF (2 mL). Triethylamine (80 mg, 1 mmol) and BSTFA (307 mg, 1.1 mmol) were added, and the mixture was stirred at rt for 1.5 h. Next, fusarie acid (143 mg, 0.7 mmol) and HATU (184 mg, 0.5 mmol) were added, and the mixture was stirred at rt for 3 h. DMF was removed and the residue was dissolved in EtOAc (50 mL), washed with sodium bicarbonate (10%», 30 mL) and brine (30 mL), and dried over sodium sulfate. The residue obtained by removing the solvent was dissolved in methanol and treated with Dowex resin H * for 1 h. The crude product obtained by filtering the resin and removing the solvent was purified on silica gel column chromatography using 10% methanol in dichloromethane as the eluent to give the title compound (100 mg, 61%). [0327] TLC Rf = 0.6 (10% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 8.47 (s, 1), 8.02 (d, J = 8.1, 1), 7.80 (d, J = 8.1, 1), 5.27 (d, J = 5.4, 1), 4.31 (m, 2), 4.12 (dd, J = 5.7, 4.2, 1), 3.85 (d, J = 3.0, 1), 3.56 (dd, J = 3.3, 6.9, 1), 2.80 (m, 2), 2.24 (m, 1), 2.11 (s, 3), 1.67 (m, 2), 1.41 (m, 2), 1.00 (m, 9); MS(ESPOS): 413 [M+H]+.
[0328] PtO2 (50 mg, 0.22 mmol) was added to compound lib (m=2, R1=Me, R2=Me, R3=H, R9=butyl), (70 mg, 0.16 mmol) in methanol (2 mL), water (lOmL), and acetic acid (3 mL), and the mixture was hydrogenated at 50 psi overnight. The product obtained after filtering the catalyst and removing the solvent was purified on silica gel column chromatography using 30%) methanol in dichloromethane as the eluent (16 mg, 46%).
[0329] TLC Rf = 0.7 (30% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.16 (m, 3), 3.82 (d, J = 3.3, 1), 3.53 (m, 2), 2.93 (m, 2), 2.09 (s, 3), 1.93 (m, 1), 1.76 (m, 2), 1.50 (m, 1), 1.30 (m, 7), 0.92 (m, 9); MS(ESPOS): 419 [M+H]+.
Example 8 Preparation of 4-Pentyl-pyrrolidine-2-carboxyIic acid [2-methyI-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0330] Triethylamine (0.2 mL, 1.44 mmol, 3.6 equiv), followed by BSTFA (0.2 mL, 0.76 mmol, 1.9 equiv), were added to a stirred suspension of 2b (R1=Me, R2=Me) prepared by Method C (100.4 mg, 0.4 mmol, 1 equiv) in anhydrous DMF (2 mL) at 0°C and under nitrogen. The resulting mixture was stirred at 0°C for 10 min, and then at rt for 50 min. The resulting solution was cooled to 0°C and a solution of 6c (R9=pentyl) (Scheme 6) prepared as described in Birkenmeyer, R. D; et al; Journal of Medicinal Chemistry 1972, 15, 1255-1259. (144 mg, 0.51 mmol, 1.2 equiv) in anhydrous DMF (1.5 mL) was added, followed by solid HATU. The reaction mixture was allowed to warm to rt, and after 2 h, the reaction solution was evaporated to dryness under vacuum. The residual oil obtained was diluted with EtOAc (150 mL), washed sequentially with 10% citric acid (2 x 30 mL), 1:1 saturated aqueous NaHCO3, water (2 x 30 mL), and brine (30 mL), dried over Na2SO4, and evaporated to dryness. [0331] 1 ,2-Dichloroethane (8 mL), followed by dimethyl sulfide (180.3 μl), TFA (2.7 mL), and water (180.3 μl) were added to the crude product (267.5 mg) obtained above. The resulting mixture was stirred at rt for lh and evaporated to a minimal volume, diluted with DCE (3 x 30 mL), and evaporated to dryness. The residue obtained was purified by chromatography over silica gel, with a gradient eluent of 8-10% methanol ammonia in dichloromethane. The desired fractions were pooled together, evaporated to dryness, and lyophilized to furnish the title compound as a white fluffy powder (35.6 mg, 21.2%).
[0332] TLC, Rf = 0.15 (16% 0.25M methanolic ammonia in dichloromethane ). 1H NMR (300MHz, D2O) δ 5.4 (d, J = 5.8, 1), 3.91(s, 1), 3.69-3.66 (m, 3), 2.1 (s, 3), 1.32-1.15 (m, 3.37), 0.93-0.87 (m, 9.8); MS(ESPOS): 419.5 [M+H]+, (ESNEG): 417.45 [M-H]".
Example 9 Preparation of 4-(3-Methyl-butyi)-pyrrolidme-2-carboxyIic acid [2-methyI-l-(3,4,5- trihy droxy-6-methylsulf nyl-tetrahy dro-py ran-2-yl)-propy 1] -amide
[0333] Triethylamine (0.13 mL, 0.96 mmol, 3.2 equiv), followed by BSTFA (0.12 mL, 0.45 mmol, 1.5 equiv), were added to a solution of 2b (R1=Me, R2=Me) prepared by Method C (75 mg, 0.30 mmol, 1 equiv) in dry DMF (0.8 mL) at 0°C. The reaction mixture was stirred at 0°C for 10 min, and then at rt for 50 min. To the reaction mixture was added acid 7d (R9=2- methylbutyl), prepared by method J (160 mg, 0.56 mmol, 1.9 equiv), in a 25 mL round-bottom flask. Then HATU was added (227 mg, 0.60 mmol, 2 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (100 mL), washed with 10% citric acid (2 x 60 mL), water (60 mL), half sat. NaHCO3 (60 mL), and brine. The organic layer was dried over Na S0 and evaporated to give a yellow syrup. [0334] Trifluoroacetic acid (5 mL) and water (0.33 mL) were added to a solution of the above syrup in dichloromethane (15 mL) with methyl sulfide (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (75 mg, 60%) as a white solid.
[0335] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.4, 1), 4.18-3.99 (m, 4), 3.75 (d, J = 2.4, 1), 3.51 (dd, J = 3.3, 10.5, 1), 3.38-3.31 (m, 1), 2.68 (dd, J = 8.2, 10.6, 1), 2.23-2.05 (m, 3), 2.10 (s, 3), 1.97-1.87 (m, 1), 1.59-1.47 (m, 1), 1.46-1.34 (m, 2), 1.25-1.16 ( , 2), 0.92-0.88 (m, 12). MS(ESPOS): 419.5 [M+H]+, MS(ESNEG): 417.5 [M-H]".
Example 10 Preparation of 4-Pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2~yl)-propyl]-amide
[0336] Triethylamine (0.6 mL, 4.33 mmol, 3.6 equiv), followed by BSTFA (0.6 mL, 2.27 mmol, 1.9 equiv), were added to a stirred suspension of 2b (R1=Me, R2=Me) prepared by Method C (298.8 mg, 1.19 mmol, 1 equiv) in anhydrous DMF (5 mL) at 0°C and under nitrogen. The resulting mixture was stirred at 0°C for 10 min, and then at rt for 50 min. The resulting solution was cooled to 0°C and a solution of 7d (R =n-pentyl) was prepared by Method K (400.1 mg, 1.40 mmol, 1.2 equiv) in anhydrous DMF (5 mL) was added, followed by solid HATU (678.7 mg, 1.79 mmol, 1.5 equiv). The reaction mixture was allowed to warm to rt and after 2h the reaction solution was evaporated to dryness under vacuum. The residual oil obtained was diluted with EtOAc (400 mL), washed sequentially with 10% citric acid (2 x 100 mL), 1:1 saturated aqueous NaHC03, water (2 x 100 mL), and brine (100 mL), dried over Na2S0 , and evaporated to dryness.
[0337] 1,2-Dichloroethane (35 mL), followed by dimethylsulfide (768 μL), TFA (11.5 mL), and water (768 μL) were added to the crude product (1.14 g) obtained above. The resulting mixture was stirred at rt for lh, evaporated to a minimal volume, diluted with DCE (3 x 90 mL), and evaporated to dryness. One-third of the residue obtained was purified by chromatography over silica gel, with a gradient eluent of 8-12% methanol ammonia in dichloromethane. The desired fractions were pooled together, evaporated to dryness, treated with deuterium oxide/anhydrous acetonitrile, and lyophilized to furnish a white fluffy powder (68.2mg, 41.1%); TLC, Rf= 0.15 {16% 0.25M methanolic ammonia in dichloromethane}. 1H NMR (300 MHz, D20) δ 5.41 (d, J = 5.8, IH), 4.55 (m, 1), 4.24 (s, 2), 4.14 (m, 1), 3.91(d, J=3.3, 1), 3.70-3.66 (m, 2), 3.15 (m,l), 2.36-2.27 (m, 2), 2.19 (s, 5), 1.59-1.13 (m, 9), 0.93-0.88 (m, 9); 13C NMR (D20,): δ 170.4, 119.4, 88.4, 70.9, 69.3, 68.8, 68.2, 60.0, 53.4, 51.4, 37.3, 36.7, 31.3, 27.9, 27.2, 22.3, 20.1, 14.8, 13.7, 13.3; MS(ESPOS):419.6[M+H]+;(ESNEG):417.5[M-H]".
Example 11 Preparation of l-Methyl-4-propyl-pyrrolidine-2-carboxylic acid [2,2-difluoro-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0338] 30% Trifluoroacetic acid in dichloroethane (10 mL) and dimethylsulfide (0.5 mL) were added to lincosamine intermediate 5b (R1=Me), prepared by Method H (100 mg, 0.20 mmol). The mixture was stirred at rt for 1 h. The solvent was removed and the residue was kept under high vacuum for 1 h. N-Methyl-4-trans-propylproline (53 mg, 0.4 mmol) and HATU (114 mg, 0.30 mmol) were added to the dried compound in DMF (3 mL), and the mixture was stirred at rt overnight. DMF was removed and the residue obtained was then extracted with ethyl acetate (100 mL) and washed with saturated bicarbonate (50 mL). The organic portion was then dried using magnesium sulfate and the solvent was removed to obtain the crude product. The crude product was purified on silica gel column using ethyl acetate as the eluent (50 mg, 46%). The product (50 mg, 0.09 mmol) was then taken in methanol (2 mL) and water (1 mL), to which solid potassium carbonate (124 mg, 0.90 mmol) was added and the mixture was stirred at rt for 24 h. Solvents were then removed and the crude product was purified on silica gel column using 20%) methanol in dichloromethane as the eluent (20 mg, 52%>).
[0339] TLC: Rf = 0.57 (20% methanol in dichloromethane); MS(ESPOS): 427 [M+H]+. 1H NMR (300 MHz, CD3OD) δ 0.91 (m, 3), 1.34 (m, 4), 1.69 (t, J = 19.8, 3), 1.98 (s, 3), 2.20 (m, 2), 2.46 (s, 3), 3.18 (dd, J = 5.1, 10.20, 1), 3.93 (d, J = 3.0, 1), 4.08 (dd, J = 3.3, 10.20, 1), 4.40- 4.70 (m, 2), 5.28 (d, J = 5.4, 1).
Example 12 Preparation of 4-Pentyl-pyrrolidine-2-carboxylic acid [2,2-difluoro-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0340] Boc 4-trans-Pentylproline 7d (R9=n-pentyl) (179 mg, 0.631 mmol), HATU (299 mg, 0.789 mmol), and diethylisopropylamine (182 mg, 1.2 mmol) were added to lincosamine intermediate 5b (R1=Me) prepared by Method H (210 mg, 0.526 mmol) in DMF (3 mL) at 0°C. The mixture was stirred at rt overnight. After removing DMF, the residue was taken in ethyl acetate and washed with saturated bicarbonate (30 mL). The organic portion was then dried over sodium sulfate and the solvent was removed to obtain the crude product. The crude product was purified by column chromatography using 30% ethyl acetate in hexanes as the eluent (200 mg, 57%). Potassium carbonate (450 mg, 3.0 mmol) was added to the product (200 mg, 0.30 mmol) of the above reaction in methanol (3 mL) and water (1 mL), and the mixture was stirred at rt for 2 h. The solvent was removed and the residue obtained was taken in 30%) trifluoroacetic acid in dichloroethane (10 mL) and dimethyl sulfide (0.5 mL) and stirred for 1 h. After removing the solvent, the crude product obtained was purified by column using 10%) methanol in dichloromethane as the eluent (10 mg, 90%).
[0341] TLC: Rf - 0.56 (20% methanol in dichloromethane); JH NMR (300 MHz, CD3OD) δ 0.90 (m, 3), 1.31 (m, 7), 1.44 (m, 20), 1.73 (t, J = 19.5, 3), 2.02 (m, 1), 2.08 (s, 3), 2.24 (m, 2), 2.89 (t, J = 9.9, 1), 3.56 (m, 2), 3.86 (s, 1), 4.07 (dd, J = 6.0, 9.6, 1), 4.37 (m, 2), 4.63 (m, 1), 5.28 (d, J = 5.4, 1); MS(ESPOS): 441 [M+H]+.
Example 13 Preparation of 4-[3-(4-Fluoro-phenyl)-propyl]-pyrrolidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0342] Triethylamine (0.13 mL, 0.96 mmol, 3.2 equiv), followed by BSTFA (0.12 mL, 0.45 mmol, 1.5 equiv), were added to a solution of 2b (R1=Me, R =Me) prepared by Method C (75 mg, 0.30 mmol, 1 equiv) in dry DMF (0.8 mL) at 0 °C. The reaction mixture was stirred at 0 °C for 10 min, and then at rt for 50 min. The reaction mixture was added to acid 8c (R9=3-(4- fluorophenyl)propyl) prepared by Method L (120 mg, 0.34 mmol, 1.1 equiv) in a 25 mL round bottom flask, followed by HATU (160 mg, 0.42 mmol, 1.4 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (100 mL), and washed with 10% citric acid (2 x 60 mL), water (60 mL), half sat. NaHCO3 (60 mL), and brine. The organic layer was dried over Na2SO and evaporated to give a yellow syrup.
[0343] Trifluoroacetic acid (5 mL) and water (0.33 mL) were added to a solution of the above syrup in dichloromethane (15 mL) with methyl sulfide (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide 4-[3-(4-fluoro-phenyl)- propyl]-pyrrolidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl] -amide (90 mg, 62%) as a white solid.
[0344] 1H NMR (300 MHz, CDC13) δ 7.94 (brs, 1), 7.11 -7.06 (m, 2), 6.97-6.90 (m, 2), 5.31 (d, J = 5.4, 1), 4.10 (dd, J = 5.7, 9.9, 1), 3.96-3.82 (m, 3), 3.68-3.52 (m, 2), 3.10-3.20 (m, 1), 2.70-2.60 (m, 1), 2.56 (dd, J = 7.4, 7.4, 2), 2.36-2.24 (m, 1), 2.13 (s, 3), 2.10-1.93 (m, 2), 1.85- 1.73 (m, 1), 1.64-1.50 (m, 2), 1.40-1.30 (m, 2), 0.92-0.85 (m, 6). MS(ESPOS): 485.5 [M+H]+, MS(ESNEG): 483.5 [M-H]". Example 14 Preparation of 4-(3,3-Difluoro-propyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0345] Triethylamine (0.13 mL, 0.96 mmol, 3.2 equiv), followed by BSTFA (0.12 mL, 0.45 mmol, 1.5 equiv), were added to a solution of 2b (R =Me, R =Me) prepared by Method C (75 mg, 0.30 mmol, 1 equiv) in dry DMF (0.8 mL) at 0 °C. The reaction mixture was stirred at 0°C for 10 min, and then at rt for 50 min. The reaction mixture was added to acid 8c (R9=3,3- difluoropropyl) prepared by Method N (97 mg, 0.33 mmol, 1.1 equiv) in a 25 mL round bottom flask, followed by HATU (170 mg, 0.45 mmol, 1.5 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (100 mL), and washed with 10% citric acid (2 x 60 mL), water (60 mL), half sat. NaHCO3 (60 mL), and brine. The organic layer was dried over Na2SO4 and evaporated to give a yellow syrup. [0346] Trifluoroacetic acid (5 mL) and water (0.33 mL) were added to a solution of the above syrup in dichloromethane (15 mL) with methyl sulfide (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide 4-(3,3-difluoro-propyl)- pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propyl] -amide (81 mg, 64%) as a white solid.
[0347] 1H NMR (300 MHz, CDC13) δ 7.92 (d, J = 8.1, 1), 5.80 (dddd, J = 4.2, 4.2, 57, 57, 1), 5.31 (d, J = 5.7, 1), 4.11 (dd, J = 5.4, 9.9, 1), 3.96-3.82 (m, 3), 3.64-3.52 (m, 2), 3.23-3.10 (m, 1), 2.73-2.60 (m, 1), 2.36-2.23 (m, 1), 2.13 (s, 3), 2.18-1.95 (m, 2), 1.90-1.73 (m, 3), 1.56-1.43 (m, 2), 0.93-0.85 (m, 6). MS(ESPOS): 427.5 [M+H]+, MS(ESNEG): 425.5 [M-H]". Example 15 Preparation of 4-[3-(4-Chloro-phenyl)-propyl]-pyrrolidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0348] Triethylamine (88.3 μL, 0.64 mmol, 3.2 equiv), followed by BSTFA (79.2 mL, 0.30 mmol, 1.5 equiv), were added to a solution of 2b (R1=Me, R2=Me) prepared by Method C (50 mg, 0.20 mmol, 1 equiv) in dry DMF (0.5 mL) at 0°C. The reaction mixture was stirred at 0°C for 10 min, and then at rt for 50 min. The reaction mixture was added to 8c (R9=3-(4- chlorophenyl)propyl) prepared by Method M (97.3 mg, 0.26 mmol, 1.3 equiv) in a 25 mL round bottom flask, followed by HATU (123 mg, 0.32 mmol, 1.6 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), and washed with 10%) citric acid (2 x 40 mL), water (40 mL), half sat. NaHCO3 (40 mL), and brine. The organic layer was dried over Na2SO4 and evaporated to give a yellow syrup.
[0349] Trifluoroacetic acid (3 mL) and water (0.2 mL) were added to a solution of the above syrup in dichloromethane (9 mL) with methyl sulfide (0.2 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide 4-[3-(4-chloro-phenyl)-propyl]- pyrrolidine-2 -carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-metl ylsulfanyl-tetrahydro-pyran- 2-yl)-propyl]-amide (41.6 mg, 42%) as a white solid.
[0350] 1H NMR (300 MHz, CDC13) δ 7.26-7.21 (m, 2), 7.17-7.12 (m, 2), 5.23 (d, J = 5.7, 1), 4.10-4.00 (m, 3), 3.83-3.75 (m, 1), 3.74-3.70 (m, 1), 3.54-3.48 (m, 1), 3.25-3.18 (m, 1), 2.63- 2.50 (m, 3), 2.20-2.00 (m, 3), 2.09 (s, 3), 1.85-1.74 (m, 1), 1.68-1.55 (m, 2), 1.42-1.33 (m, 2), 0.95-0.85 (m, 6). MS(ESPOS): 501.5 [M+H]+ MS(ESNEG): 499.4 [M-H]". Example 16 Preparation 4-(2,2-Difluoro-pentyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0351] Triethylamine (88.3 mL, 0.64 mmol, 3.2 equiv), followed by BSTFA (79.2 mL, 0.30 mmol, 1.5 equiv), were added to a solution of 2b (R1=Me, R2=Me) prepared by Method C (50 mg, 0.20 mmol, 1 equiv) in dry DMF (0.5 mL) at 0°C. The reaction mixture was stirred at 0°C for 10 min, and then at rt for 50 min. The reaction mixture was added to the acid 9d (R9=2,2- difluoropentyl) prepared by Method O (67.7 mg, 0.21 mmol, 1.1 equiv) in a 25 mL round bottom flask, followed by HATU (101 mg, 0.27 mmol, 1.3 equiv). The reaction mixture was stirred at rt for 3h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), and washed with 10% citric acid (2 x 40 mL), water* (40 mL), half sat. NaHCO3 (40 mL), and brine. The organic layer was dried over Na SO4 and evaporated to give a yellow syrup. Trifluoroacetic acid (3 mL) and water (0.20 mL) were added to a solution of the above syrup in dichloromethane (9 mL) with methyl sulfide (0.20 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide 4-(2,2-Difluoro-pentyl)-pyrrolidine-2- carboxylic acid [2-met-hyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]- amide (56 mg, 62%) as a white solid.
[0352] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.17-4.04 (m, 3), 3.98 (dd, J = 3.3, 9.3, 1), 3.77 (d, J = 3, 1), 3.51 (dd, J = 3.4, 10.3, 1), 3.40 (dd, J = 6.9, 10.5, 1), 2.71 (dd, J = 10.2, 10.2, 1), 2.42-2.33 (m, 1), 2.23-2.11 (m, 2), 2.10 (s, 3), 2.08-1.73 (m, 5), 1.56-1.42 (m, 2), 0.99-0.89 (m, 9). MS(ESPOS): 455.5 [M+H]+; MS(ESNEG): 453.5 [M-H]".
Example 17 Preparation of 4-Propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsιιlfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0353] Triethylamine (0.18 mL, 1.26 mmol) and BSTFA (0.549 mL, 2.1 mmol) were added to the lincosamine intermediate 2b (R1=Me, R2=Me) prepared by Method D (102 mg, 0.42 mmol) in DMF (5 mL) at 0°C, and the mixture was stirred at rt for 3 h. Acid 10b (R9=propyl) prepared by Method P (200 mg, 0.84 mmol) and HATU (319 mg, 0.84 mmol) were added and the mixture was stirred for 4 h at rt. DMF was removed and the residue was extracted with ethyl acetate (100 mL) and washed with saturated bicarbonate (40 mL). The product obtained by removal of solvent was taken up in methanol and treated with Dowex H1" resin for 1 h. After filtering the resin, methanol was removed to obtain the crude product. The crude product was then purified on silica gel column using 10% methanol in dichloromethane as the eluent to provide pyridine lib (R1=Me, R2=Me, R3=H, R9=propyl) (117 mg, 58%). [0354] TLC: Rf = 0.81 (10% methanol in dichloromethane); 1H NMR (300 MHz, CDC13) δ 1.20 (t, J = 6.3, 6H), 2.19 (m, 2), 2.32 (s, 3), 2.43 (m, 1), 2.84-2.97 (m, 4), 3.74 (m, 1), 4.06 (m, 1), 4.31 (m, 1), 4.52 (m, 2), 5.42 (d, J = 5.7, 1), 7.33-7.61 (m, 5), 7.80 (m, 1), 8.15 (s, 1), 8.69 (d, J = 4.8, 1); MS (ESPOS): 475 [M+H]+.
[0355] PtO2 (100 mg, 0.44 mmol) was added to pyridine lib (R1=Me, R2=Me, R3=H, R9==propyl), (150 mg, 0.37 mmol) in methanol (2 mL), water (lOmL), and acetic acid (3 mL), and the mixture was hydrogenated at 50 psi overnight. The product obtained after filtering the catalyst and removing the solvent was purified by silica gel column chromatography using 30% methanol in dichloromethane as the eluent to provide the title compound (20 mg, 1 %). [0356] TLC: Rf = 0.7 (50% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 6.9, 1), 4.86 (m, 1), 4.13 (m, 2), 3.79 (d, J = 3.3, 1), 3.52 (dd, J = 3.3, 9.9, 1), 3.32 (m, 1), 3.17 (m, 1), 2.67 (m, 1), 2.17 (m, 1), 2.10 (s, 3), 1.97 (m, 1), 1.74 (m, 1), 1.54 (m, 1), 1.38 (m, 2), 1.31 (m, 2), 1.14 (m, 2), 1.02 (m, 9) MS(ESPOS): 405 [M+H]+. Example 18 Preparation of l-(2-Hydroxy-ethyl)-4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0357] DIEA (0.1 mL, 0.57 mmol) and liquid ethylene oxide (3 mL) were added to a stirred solution of crude 4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide, prepared as in Example 10 (237.4 mg), in anhydrous methanol (10 mL), at 0°C and under nitrogen. The resulting solution was stirred at - 4°C for 18 h and evaporated to dryness. The residue obtained was purified by chromatography over silica gel with an eluent of 5% methanol ammonia in dichloromethane. The desired fractions were evaporated and lyophilized (deuterium oxide/anhydrous acetonitrile, 1:1, v/v, lOmL) to furnish the title compound as a fluffy white powder (50.1mg, 30.2%); TLC, Rf= 0.68 (14% methanolic ammonia in dichloromethane); 1HNMR (300 MHz) δ 5.40 (d, J=5.8, 1), 4.55 (m, 1), 4.24 (s, 1), 4.17-4.11 (m, 1), 3.99-3.89 (m, 4), 3.69-3.65 (m, 1), 3.47 (d, J=4.4, 2), 3.01 (m, 1), 2.33 (br s,4), 2.18 (s,4), 1.57-1.32 (m,9), 0.94-0.87 (m, 9). MS(ESPOS):464[M+H]+;(ESNEG):497.5[M-H+HCTJ.
Example 19 Preparation of l-(2-Hydroxy-propyl)-4-pentyl-pyrrolidine-2-carbo-sylic acid [2-methyl-l- (3,4s5-trihydrosy-6-methyls«lfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0358] DIEA (O.lmL, 0.58 mmol, 1 equiv) and R(+)-propylene oxide (3 mL) were added to a stirred cool solution of crude 4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide (307.6 mg, 0.58 mmol, 1 equiv), prepared as in Example 10, in anhydrous methanol (lO L), at 0°C and under nitrogen. The resulting solution was stirred at -4°C for 18 h and evaporated to dryness. The residue obtained was purified by chromatography over silica gel, with an eluent of 6% methanol ammonia in dichloromethane. The desired fractions were evaporated, and lyophilized (deuterium oxide/anhydrous acetonitrile, 1:1, v/v, 20mL) to furnish the title compound as a fluffy white powder (91mg, 48%).
[0359] TLC, Rf = 0.7 (14% methanolic ammonia in dichloromethane); 1H N.MR(300 MHz, CD3OD) δ 5.44 (d, J = 5.5, 1), 4.31 (s, 2), 4.26-4.11 (m, 1), 3.97 (d, J = 3.3,1.1,1), 3.75 (dd, J = 3.3,3.3, 1), 3.39 (dd, J = 3.8,3.8, 1), 2.31 (s, 3), 1.5-0.95 (m, 12), 1.34 (d, J = 6.0, 4), 1.17-1.10 (m, 13); MS(ESPOS): 477.6 [M+H]+, (ESNEG): 475.6 [M-H]".
Example 20 Preparation of l-(2-Hydroxy-propyϊ)-4-pentyϊ-pyrrolidine-2-carboxylk acid [2-methyl-l- (3,4,5-trihydroxy-6-methylslfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0360] Dimethyl sulfide (62 μL), TFA (1 mL), and water (62 μL) were added to a stirred solution of the Boc-protected 4-pentyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide (92mg, 0.18 mmol), prepared as in Example 10, in anhydrous dichloroethane (3 mL). The resulting solution was stirred at rt for 1 h and evaporated to dryness. Anhydrous methanol (8 mL) and DIEA (31 μL, 0.18 mmol) were added to the residue obtained. The mixture was cooled to -4°C and S-(-)-ρropylene oxide (2 mL) was added. The resulting solution was stirred at -4°C for 18 h, evaporated to dryness, and purified by chromatography over silica gel, with an eluent of 6% methanol ammonia in dichloromethane. The desired fractions were evaporated and lyophilized (deuterium/anhydrous acetonitrile, 1:1, v/v, 8 mL) to furnish the title compound as a fluffy white powder (29.8 mg, 31.2%).
[0361] TLC, Rf = 0.7 (12% methanolic ammonia in dichloromethane); IH NMR (300 MHz, CD3OD) δ 5.44 (d, J = 5.5,1), 4.35-4.19 (m, 4), 4.02 (d, J = 3.3, 2), 2.75 (d, J = 6.3,2.2, 3), 2.3 (s, 3), 1.50 (m, 11), 1.4 (d, J = 6.0, 3.5, 3), 1.16-1.10 (m, 12). MS(ESPOS): 477.6 [M+H]+; (ESNEG) 475.4 [M-H]".
Example 21 Preparation of l-(3-Hydroxy-propyl)-4-pehtyl-pyrrolidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0362] Triethylamine (0.2 mL, 1.38 mmol, 3 equiv), followed by 3 -bromo- 1 -propanol (60 μL, 0.69 mmol, 1.5 equiv), were added to a stirred solution of crude 4-pentyl-pyrrolidine-2- carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]- amide (192.5 mg, 0.46 mmol, 1 equiv), prepared as in Example 10, in anhydrous acetonitrile (2 mL), under nitrogen. The resulting mixture was stirred at rt for 18 h and evaporated to dryness. The residue obtained was purified by chromatography over silica gel with an eluent of 5% methanol ammonia in dichloromethane. The desired fractions were pooled together, evaporated to dryness, and lyophilized to furnish the title compound as a white fluffy powder (13.5 mg, [0363] TLC, Rf = 0.75 (14% methanolic ammonia in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 5.44 (d, J = 5.8, 1), 4.33-4.26 (m, 4), 4.01 (d, J = 2.7, 1), 3.85-3.74 (m, 6), 2.29 (s, 3), 2.1 (m, 4), 1.54 (m, 8), 1.16-1.08 (m, 12); MS (ESPOS): 477.6 [M+H]+.
Example 22 Preparation of l-(2-Hydroxy-ethyl)-4-(3-methyl-butyl)-pyrrolidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Ethylene oxide (0.6 mL) was added to a solution of 4-(3-methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide (35.1 mg, 0.084 mmol), prepared as in Example 9, in methanol (3 mL), at 0°C. The reaction mixture was stirred at 4°C overnight. Additional ethylene oxide (0.6 mL) was added and stirred at 4°C overnight. The reaction mixture was concentrated and purified by chromatography to give a white solid, l-(2-hydroxy-ethyl)-4-(3-methyl-butyl)-pyrrolidine-2-carboxylic acid [2-methyl-l - (3,4,5-trihydroxy-6-methylsulf-----jιyl-tetrahydro-pyran-2-yl)-propyl]-amide (24.1 mg, 62%). [0364] 1H NMR (300 MHz, CDC13) δ 7.68 (d, J = 9.0, 1), 5.32 (d, J = 5.4, 1), 5.24 (d, J = 3.0, 1), 4.13-4.07 (m, 1), 4.01 (ddd, J = 2.8, 9.9, 9.9, 1), 3.86 (d, J = 10.8, 1), 3.78-3.68 (m, 2), 3.61-3.57 (m, 1), 3.56-3.48 (m, 1), 3.36-3.32 (m, 1), 3.27-3.21 (m, 1), 2.94-2.85 (m, 1), 2.76- 2.70 (m, 1), 2.55 (ddd, J = 3.6, 3.6, 12.6, 1), 2.41-2.37 (m, 1), 2.36-2.27 (m, 1), 2.15 (s, 3), 2.03- 1.95 (m, 2), 1.93-1.81 (m, 1), 1.54-1.42 (m, 1), 1.39-1.26 (m, 2), 1.22-1.10 (m, 2), 0.99-0.92 (m, 6), 0.90-0.84 (m, 6). MS(ESPOS): 463.5 [M+H]+ MS(ESNEG): 461.5 [M-H]". Example 23 Preparation of 4-(3,3-Difluoro-propyl)-l-(2-hydroxy-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyI]-amide
[0365] Ethylene oxide (0.4 mL) was added to a solution of 4-(3,3-difluoro-propyl)- pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propyl]-amide, prepared as in Example 14 (29.7 mg, 0.07 mmol), in methanol (2 mL), at 0°C. The reaction mixture was stirred at 4°C overnight. Additional ethylene oxide (0.4 mL) was added and stirred at 4°C overnight. The reaction mixture was concentrated and purified by chromatography to give a white solid, l-[2-(S)-4-(R)-(3-methylbut-l-yl)-N-(2-hydroxyeth-l- yl)pyrrolidin-2-yl]-N-{l-(R)- [2-(S)-3-(S), 4-(S), 5-(R)-trihydroxy-6-(R)- (methylthio)tetrahydropyran-2-yl]-2-methylprop-l-yl} acetamide (19.3 mg, 59%). [0366] 1H NMR (300 MHz, CDC13) δ 5.86 (dddd, J = 4.3, 4.3, 57, 57, 1), 5.23 (d, J = 5.7, 1), 4.13-4.04 (m, 3), 3.75 (d, J = 3.3, 1), 3.73-3.57 (m, 2), 3.53 (dd, J = 3.3, 10.2, 1), 3.42-3.36 (m, 1), 3.26-3.18 (m, 1), 2.88-2.78 (m, 1), 2.62-2.55 (m, 1), 2.17-2.00 (m, 4), 2.10 (s, 3), 1.94-l'.73 (m, 3), 1.55-1.45 (m, 2), 0.98-0.91 (m, 6). MS (ESPOS): 471.5 [M+H]+, MS (ESNEG): 469.4 [M-H]".
Example 24
Preparation of l-(2-Hydrosy-ethyl)-4-pentyl-pyrrolidine-2-carbo---ylic acid [2,2-diflHoro-l- (394,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0367] Ethylene oxide (1 mL, excess) was added to the title compound from Example 12 (60 mg, 0.136 mmol) in methanol (5 mL) at 0°C, and the mixture was stirred at 4°C overnight. The solvent was removed and the crude product was purified by silica gel column chromatography using 10% methanol in dichloromethane as the eluent (25 mg, 38%).
[0368] TLC: Rf = 0.76 (5% methanol in dichloromethane); 1H NMR (300 MHz, CD3OD) δ 0.89 (m, 3), 1.30 (m, 8), 1.68 (t, J = 19.4, 3), 1.78 (m, 1), 1.99 (m, 2), 2.07 (s, 3), 2.63 (m, 1), 2.73 (m, 1), 3.19 (m, 1), 3.58 (m, 3), 3.95 (m, 1), 4.08 (dd, J = 6.0, 9.90, 1), 4.44-4.60 (m, 2), 5.26 (d, J = 5.4, 1); MS (ESPOS): 485 [M+H]+.
Example 25 Preparation of 4-Pentyl-piperidine-2-carboxyIic acid [2-methyl-l-(3,4,5-trihyαroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0369] 4-Pentylpyridine-2-carboxylic acid (10b) (R9=pentyl) was made by employing Method P. To 4-pentylpyridine (3 g, 20 mmol) in acetic acid (30 mL), hydrogen peroxide (0.7 g, 30%, 20 mmol) was added and refluxed overnight. Removal of solvent resulted in residue which was dissolved in DCM (100 mL) dried over MgSO and filtered. Removal of DCM resulted in a brown liquid, 4-pentylpyridine-N-oxide, (3.3 g, 100%). To trimethylsilyl cyanide (2.37 g, 24 mmol), 4-pentylpyridine N-oxide (3.3 g, 20 mmol) in DCM (10 mL) was added followed by dropwise addition of dimethylcarbamoyl chloride (2.56 g, 24 mmol) in DCM (10 mL). After sti-rring at room temperature overnight, sodium bicarbonate (100 mL, 10 %) was added and the organic layer was separated. The aqueous layer was extracted twice with DCM (50 mL) and the combined organic layer was dried over magnesium sulfate. Removal of solvent resulted in compound 10a (R9=pentyl) (4.1 g, 100%).
[0370] 1H NMR (300 MHz, CD3OD) δ 8.52 (m, 1), 7.46 (s, 1), 7.27 (m, 1), 3.00 (m, 2), 2.60 (m, 2), 1.60 (m, 2), 1.27 (m, 2), 0.86 (m, 3). MS (ESPOS): 175 [M+H]+. [0371] The 4-Pentyl-2-cyanopyridine (10a) (R9=ρentyl) (3.4 g, 19.5 mmol) from the previous step was dissolved in HCl (6 N, 100 mL) and refluxed overnight. The residue obtained on removal of HCl was purified by column chromatography using 20% MeOH in DCM (3.7 g, 100%) to give product compound 10(b). 1H NMR (300 MHz, CD3OD) δ 8.74 (d, J=6.3, 1), 8.39 (s, 1), 8.06 (d, J=6.3, 1), 2.98 (t, J=7.2, 2), 1.77(m, 2), 1.39 (m, 4), 0.95 (t, J=7.2, 3H). MS (ESNEG): 192 [M-H]".
[0372] Then to 7-methyl α-thiolincosaminide 2b (R1=Me, R2=Me) (90 mg, 0.35 mmol) in DMF (2ml), TEA (72 mg, 0.7 mmol), BSTFA (276 mg, 1.05 mmol) were added at 0 °C and stirred at room temperature for 1.5 hr. Then the acid (10b) (R =pentyl) (138 mg, 0.7 mmol) and HATU (165 mg, 0.53 mmol) was added to the reaction mixture, and stirred at room temperature overnight. DMF was completely removed, the residue was taken up in EtOAc (50 mL), washed with sodium bicarbonate (10%, 50 mL), brine (50 mL). The product obtained after drying over magnesium sulfate and concentration was taken up in methanol (10 mL) and treated with NR-50 resin (150 mg) for 3 hr. The resin was filtered and the solvent was removed. Purification of the crude product was carried out silica gel column chromatography using 3% MeOH in DCM as eluent to obtain compound lib (R1=Me, R2=Me R3=H, R9=pentyl) (90 mg, 59%): [0373] 1HNMR (300 MHz, CD3OD) δ 8.50 (d, J=5.1, 1), 7.95 (s, 1), 7.11 (m, 1), 5.25 (d, J=5.7, 1), 4.20-4.87 (m, 3), 3.85 (d, J=3.3, 1), 3.55 (dd, J=3.3, 7.2, 1), 2.72 (m, 2), 2.16 (m, 4), 1.67 (m, 2), 1.35 (m, 4), 0.96 (m, 9). MS (ESPOS): 427 [M+H]+.
[0374] To pyridine lib (R*=Me, R2=Me R3=H, R9=pentyl) (90 mg, 0.7 mmol) in water (10 mL), AcOH (3 mL) and MeOH (2 mL), Pt02 (100 mg) was added, hydrogenated at 55 psi overnight. The solvent was removed to obtain the crude product. Purification of the crude product was carried out by silica gel column chromatography using 20%> MeOH in DCM to obtain the title compound (35 mg, 38%). [0375] 1H NMR (300 MHz, CD3OD) δ 5.23 (d, J=5.1, 1), 4.17 (m, 3), 3.79 (d, J=3.3, 1), 3.52 (m, 1), 3.38 (m, 1), 3.07 (m, 1), 2.68 (m, 1), 2.14 (m, 4), 1.88 (m, 1), 1.71 (m, 1), 1.52 (m, 1), 1.30 (m, 8), 1.07 (m, 3), 0.90 (m, 9); MS (ESPOS): 433 [M+H]+.
Example 26 Preparation of 4-Methoxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0376], , 4-Methoxypyridine-2-carboxylic acid, compound 10(b) (R9 = methoxy), was made employing Method P. To trimethylsilyl cyanide (0.95 g, 9.6 mmol), 4-methoxypyridine N-oxide (1 g, 8 mmol) in DCM (10 mL) was added, followed by dimethylcarbamoyl chloride (1.03 g, 9.6 mmol) in DCM (10 mL), dropwise. After stirring at room temperature overnight, sodium bicarbonate (100 mL, 10%) was added, and the organic layer was separated. The aqueous layer was extracted twice by DCM (50 mL each). The combined organic layer was dried over magnesium sulfate and the solvent was removed to obtain product, compound 10a (0.97 g,
[0377] 1H NMR (300 MHz, CD3OD) δ 8.52 (m, 1), 7.22 (m, 1), 7.01 (m, 1), 3.92 (s, 3H);
MS (ESPOS): 135 [M+H]+.
[0378] 4-Methoxy-2-cyanopyridine, compound 10a (R9 = methoxy), (0.97 g, 7.2 mmol) was dissolved in HCl (6N, 50 mL), and refluxed overnight. The HCl was evaporated and the resulting product was crystallized from acetonile, to give compound 10b (R9=methoxy) (0.6 g,
60%).
[0379] 1H NMR (300 MHz, CD3OD) δ 8.65 (m, 1), 7.99 (m, 1), 7.68 (m, 1), 4.21 (s, 3H).
MS (ESNEG): 152 [M-H]". [0380] To 7-methyl α-thiolincosaminide, compound 2b (R1=Me, R2=Me), (90 mg, 0.35 mmol) in DMF (2 mL), TEA (72 mg, 0.7 mmol), BSTFA (276 mg, 1.05 mmol) were added at 0°C and left stirred at room temperature for 1.5 hr. Then compound 10b (R9 = methoxy) (109 mg, 0.7 mmol) and HATU (165 mg, 0.53 mmol) were added to the reaction mixture, and stirred at room temperature overnight. The DMF was completely removed and the residue was taken up in EtOAc (50 mL), washed with sodium bicarbonate (10%, 30 mL), brine (30 mL), and dried over magnesium sulfate. The solvent was removed to obtain a brown oil-like liquid, which was dissolved in methanol (10 mL) and treated with NR-50 resin for 1 hr. The resin was filtered, and the solvent was removed to obtain the crude material. Purification was carried out on silica gel column chromatography using EtOAc as eluent to obtain compound lib (R1=Me, R2=Me R3=H, R9=methoxy) (100 mg, 72%).
[0381] 1H NMR (300 MHz, CD3OD) δ 8.42 (m, 1), 7.64 (m, 1), 7.07 (m, 1), 5.25 (d, J=5.4, 1), 4.07-4.87 (m, 3), 3.94 (m, 4), 3.56 (m, 1), 2.99 (m, 2), 2.80 (m, 1), 2.22 (m, 1), 2.11 (s, 3), 0.96 (m, 3). MS (ESPOS): 387 [M+H]+.
[0382] To compound lib (R^Me, R2=Me R3=H, R9=methoxy) (100 mg, 0.26 mmol) in water (10 mL), AcOH (3 mL) and MeOH (2 mL), Pt02 (100 mg) were added and hydrogenated ■ at 55 psi overnight. The solvent was removed to obtain the crude product. Purification of the crude product was carried out'by silica gel column chromatography using 20 % MeOH in DCM to obtain the title compound. (9 mg, 9%).
[0383] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.17 (dd, J=9.9; 3.3, 1), 4.07 (m, 2), 3.79 (m, 1), 3.52 (dd, J=10.5; 3.3, 1), 3.35 (s, 3), 3.18 (m, 2), 2.72 (m, 1), 2.16 (m, 1), 2.12 (s, 3) 1.99 (m, 2), 1.50 (m, 1), 1.24 (m, 2), 0.90 (d, J=6.9, 6); MS (ESPOS): 393 [M+H]+.
Example 27 Preparation of 4-(l-Ethyl-propyl)-piperidine-2-carfooxylic acid [2-methyl-l-(3,455- trihydroxy-6-methylSHlfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0384] 4-Isopentylpyridine-2-carboxylic acid, compound 10b (R9 = 1-ethyl-propyl), was made by employing Method P. To 4-(l-ethyl-propyl)-pyridine (8.5 g, 57 mmol) in acetic acid
(30 mL), hydrogen peroxide (17.8 g, 30 %, 57 mmol) was added. The resulting reaction mixture was refluxed overnight. The residue obtained on removal of solvent was dissolved in DCM (100 mL), dried over MgSO . After filtering, the solvent was removed to obtain a brown liquid, 4-(l- ethyl-propyl)-pyridine-N-oxide (9 g, 95%).
[0385] To a solution of trimethylsilyl cyanide (6.5 g, 65 mmol) and 4-(l-ethyl-propyl)- pyridine-N-oxide (9 g, 54 mmol) in DCM (25 mL) was added a solution of dimethylcarbamoyl chloride (7 g, 65 mmol) in DCM (10 mL), dropwise. After stirring at room temperature overnight, sodium bicarbonate (100 mL, 10%) was added, and the organic layer was separated.
The aqueous layer was extracted twice with DCM (50 mL). The combined organic layer was dried over magnesium sulfate and the solvent was removed to obtain the product, compound 10a
(R9 = 1-ethyl-propyl) (9.6 g, 100%).
[0386] JH NMR (300 MHz, CD3OD) δ 8.58 (m, 1), 7.46 (m, 1), 7.26 (m, 1), 2.42 (m, 1),
1.77 (m, 4)), 0.78 (t, J=7.5, 6). MS (ESPOS): 175 [M+H]+.
[0387] Compound 10a (R9 = 1-ethyl-propyl) (9.5 g, 54 mmol) was dissolved in HCl (6N, 50 mL) and refluxed overnight. HCl was evaporated and the resulting product, compound 10b (R9
= 1-ethyl-propyl), was crystallized from acetonitrile (10 g, 100%).
[0388] 1H NMR (300 MHz, CD3OD) δ 8.86 (m, 1), 8.45 (m, 1), 8.20 (m, 1), 2.92 (m, 1),
1.87 (m, 4), 0.84 (t, J=7.5, 6). MS (ESNEG): 192 [M-H]".
[0389] To the acid 10b (77 mg, 0.4 mmol) in DMF (2 mL), 7 -methyl α-thiolincosaminide, compound 2b (R1=Me, R2=Me), (100 mg, 0.4 mmol) was added, followed by HBTU (166 mg,
0.44 mmol) and DIEA (205 mg, 0.8 mmol). The mixture was stirred at room temperature for 2 hr. The product was obtained on removal of DMF and purified by silica gel column chromatography using ethyl acetate to provide compound lib (R1=Me, R2=Me R3=H, R9= 1- ethyl-propyl) (150 mg, 89%). [0390] lH NMR (300 MHz, CD3OD) δ 8.42 (d, J=5.1, 1), 7.37 (s, 1), 7.32 (m, 1), 5.27 (d,
J=4.8, 1), 4.21-4.88 (m, 3), 3.85 (d, J=3.6, 1), 3.56 (dd, J=3.3, 10.2, 1), 2.48 (m, 1), 2.11 (m, 4),
1.00 (m, 12). MS (ESPOS): 427 [M+H]+.
[0391] To compound lib (R =Me, R2=Me R3=H, R9= 1-ethyl-propyl) (130 mg, 0.3 mmol) in water (10 mL), AcOH (2 mL) and MeOH (2 mL), PtO2 (150 mg) was added and hydrogenated at 55 psi overnight. The solvent was removed to obtain the crude product.
Purification was carried by silica gel column chromatography using 20% MeOH in DCM to obtain the title compound (40 mg, 30%).
[0392] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.17 (dd, J=9.9; 3.3, 1), 4.10 (m,
2), 3.78 (m, 1), 3.51 (m, 2), 2.81 (m, 2), 2.16 (m, 1), 2.10 (s, 3) 1.90 (m, 2), 1.76 (m, 3), 1.40 (m,
8), 0.91 (m, 9); MS (ESPOS): 433 [M+H]+.
Example 28 Preparation of 4-Isopropyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0393] 4-Isopropylpyridine-2-carboxylic acid, compound (10b) (R9 = isopropyl), was made by employing Method P. To 4-isopropylpyridine (5 g, 0.041 mol) in acetic acid (60 mL), hydrogen peroxide (30%, 4.7 g, 0.13 mol) was added and refluxed over night. After removing the solvent, the residue was dissolved in DCM dried over magnesium sulfate and taken as such for the next step. To the resulting compound in dichloromethane (10 mL) trimethylsilyl cyanide (7.0 mL, 0.07 mol) and dimethylcarbamyl chloride (5.6 mL, 0.05 mol) were added and stirred at room temperature for 24 hours. Aqueous potassium carbonate (10%, 50 mL) was added and extracted with dichloromethane (100 mL). The crude product obtained on removal of solvent was taken up in hydrochloric acid (6N, 30 mL) and refluxed for 24 hours. Removal of acid followed by crystallization of the crude product from acetonitrile resulted in acid 10b (R9 = isopropyl) (5g, 75%).
[0394] 1H NMR (300 MHz, CD3OD): δ 8.78 (d, 3 = 6, 1), 8.42 (s, 1), 8.16 (d, J = 6.0, 1),
3.25 (m, 1), 1.33 (d, J = 9.0, 6) MS (ESNEG): 164 [M-H]".
[0395] To the amine, compound 2b (R^Me, R2=Me), (140 mg, 0.56 mmol) in DMF (3 mL),
BSTFA (0.59 mL, 2.24 mmol) and triethylamine (0.18 mL, 1.26 mmol) were added at 0°C and the reaction mixture was stirred at room temperature for 3 hours. Acid 10b (R = isopropyl)
(188 mg, 1.13 mmol) and HATU (319 mg, 0.84 mmol) were combined and left stirred for further 4 hours at room temperature. The DMF was removed and the residue was extracted with ethyl acetate (100 mL) and washed with saturated bicarbonate (40 mL). The product obtained on removal of solvent was taken up in methanol and treated with Dowex ϊt resin for 1 hour. After filtering the resin, methanol was removed to obtain the crude product. It was then purified on silica gel column using 10% methanol in dichloromethane as eluent to provide compound lib
(R1=Me, R2=Me R3=H, R9 = isopropyl) (120 mg, 53 %).
[0396] 1H NMR (300 MHz, CD3OD): δ 8.42 (d, J=5.1, 1), 7.37 (s, 1), 7.32 (m, 1), 5.27 (d,
J=4.8,l), 4.21-4.88 (m, 3), 3.85 (d, J=3.6, 1), 3.56 (dd, J=3.3, 10.2, 1), 2.48 (m, 1), 2.11 (m, 1),
2.10 (s, 3), 1.20 (m, 12). MS (ESPOS): 399 [M+H]+.
[0397] To lib (R1=Me, R2=Me R3=H, R9 = isopropyl) (100 mg, 0. 257 mmol) in methanol
(5 mL), water (10 mL) and acetic acid (5 mL), platinum dioxide (100 mg, 0.44 mmol) was added and hydrogenated at 60 psi for 16 h. After filtering the catalyst, the solvent was stripped off to obtain the crude product which was then purified on silica gel column chromatography using 10% methanol in dichloromethane as eluent. The lower Rf compound was the title compound (10 mg, 9%).
[0398] 1H MR (300 MHz, CD3OD): δ 5.24 (d, J=5.7, 1), 4.17 (dd, J=9.9; 3.3, 1), 4.10 (m,
2), 3.80 (m, 1), 3.51 (m, 1), 3.16 (m, 1), 2.61 (m, 1), 2.16 (m, 1), 2.10 (s, 3) 1.90 (m, 1), 1.76 (m,
1), 1.50- 1.09 (m, 5), 0.91 (m, 12); MS (ESPOS): 405 [M+H]+.
Example 29 Preparation of 4-Butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide
[0399] 4-Butylpyridine was prepared by adding potassium-t-butoxide (0.68 g, 6 mmol) to propylphosphonium bromide (Aldrich) (2.4 g, 6.0 mmol) in THF (10 mL), at 0 °C and stirring at room temperature for 1 hour. Pyridine-4-carbaldehyde (428 mg, 4 mmol) was added and the reaction mixture stirred for 2h. The reaction mixture was then poured into water and extracted with ethyl acetate. The product obtained after removing the solvent was taken as such in methanol (30 mL) to which palladium on carbon (10%, 300 mg) was added and hydrogenated at 1 atm pressure over night. Removal of solvent and purification on column chromatography using ethyl acetate resulted in pure product 4-butylpyridine (500 mg, 92%): [0400] 1H NMR (CDC13): δ 8.42 (d, J = 6.0, 2), 7.05 (d, J = 6.0, 1), 2.60 (t, J = 6.5, 2), 1.62 (m, 2), 1.37 (m, 2), 0.93 (t, J = 7.0, 3). MS (ESPOS): 136 [M+H]+. [0401] 4-Butylpyridine-2-carboxylic acid, compound (10b) (R9 = butyl), was made employing Method P. To 4-butylpyridine (2 g, 0.014 mol) in acetic acid (15 mL), hydrogen peroxide (30%>, 5 mL, 0.056 mol) was added and refluxed over night. After removing the solvent, the residue was dissolved in DCM dried over magnesium sulfate and taken as such for the next step. To the compound from the previous step in dicl loromethane (10 mL) trimethylsilyl cyanide (3.92 mL, 0.029 mol) and dimethylcarbamoyl chloride (2.67 mL, 0.028 mol) was added and stirred at room temperature for 24 hours. Aqueous potassium carbonate (10%), 50 mL) was added and extracted with dicliloromethane (100 mL). The crude product obtained on removal of solvent was taken up in hydrochloric acid (6N, 30 mL) and refluxed for 24 hours. Removal of acid followed by crystallization of the crude product from acetonitrile resulted in acid 10b (R9 = butyl) (1.5g, 60%).
[0402] 1H NMR (CDCI3): δ 8.92 (d, J = 6.0, 1), 8.65 (s, 1), 8.27 (m, 1), 3.23 (t, J = 6.5, 2), 1.98 (m, 2), 1.67 (m, 2), 1.20 (t, J = 7.0, 3). MS (ESNEG): 178 [M-H]".
[0403] To the amine, compound 2b (R1=Me, R2=Me), (140 mg, 0.56 mmol) in DMF (3 mL), BSTFA (0.59 mL, 2.24 mmol) and triethylamine (0.18 mL, 1.26 mmol) were added at 0 °C and then stirred at room temperature for 3 hours. Acid 10b (R9 = butyl) (203 mg, 1.13 mmol) and HATU (319 mg, 0.84 mmol) were added and the reaction mixture was stirred for 4 more hours at room temperature. The DMF was removed and the residue was extracted with ethyl acetate (100 mL) and washed with saturated bicarbonate (40 mL). The product obtained on removal of solvent was taken up in methanol and treated with Dowex ϊ resin for 1 hour. After filtering the resin, methanol was removed to obtain the crude product. The product was then purified on 1 "7 silica gel column using ethyl acetate as eluent to provide for compound lib (R =Me, R =Me
R3=H, R9 = butyl) (200 mg, 86 %).
[0404] 1H NMR (CDC13) δ 8.40 (d, J = 4.2, 1), 8.01 (s, 1), 7.29 (m, 1), 5.40 (d, J = 5.4, 1),
4.02-4.36 (m, 3), 4.80 (s, 1), 3.48-3.60 (m, 1), 3.72 (t, J = 6.0, 2), 2.49 (m, 1), 2.20 9s, 3), 1.67
(m, 4), 1.40 (m, 3), 0.98-1.18 (m, 9). Mass 413 [M+H]+.
[0405] To compound 1 lb (R =Me, R2=Me R3=H, R9 = butyl) (200 mg, 0. 49 mmol) in methanol (5 mL), water (10 mL) and acetic acid (5 mL), platinum dioxide (100 mg, 0.44 mmol) was added and hydrogenated at 60 psi for 16 h. After filtering the catalyst, the solvent was stripped off to obtain the crude product, which was then purified on silica gel column chromatography using 20% methanol in dichloromethane as eluent. The lower Rf fractions provided the title compound (60 mg, 29%).
[0406] ]H NMR (CDCI3): δ 5.20 (d, J = 3.6, 1), 4.20 (dd, J = 3.0, 4.8, 1), 4.04 (m, 2), 3.80
(d, J =3.0, 1), 3.61-3.66 (m, 1), 3.52 (dd, J = 3.3, 10.2), 2.88 (m, 1), 2.17 (m, 1), 2.14 (s, 3), 1.87
(m, 2), 1.62 (m, 2), 1.32 (m, 6), 0.89 (m, 9); MS (ESPOS): 419 [M+H]+.
Example 30 Preparation of 4-Cyclohexyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0407] 4-phenylpyridine-2-carboxylic acid, compound 10b (R9 =phenyl), was made by employing Method P. To 4-phenylpyridine-N-oxide (1 g, 5.84 mmol) in dichloromethane (10 mL) trimethylsilyl cyanide (1.5 mL, 11.6 mmol) and dimethylcarbamoyl chloride (1 mL, 11.6 mmol) were added and the reaction mixture was stirred at room temperature for 24 hours. An aqueous potassium carbonate solution (10%, 10 mL) was added and extracted with dichloromethane (100 mL). The crude product obtained on removal of solvent was taken up in hydrochloric acid (6N, 30 mL) and refluxed for 24 hours. Removal of acid followed by crystallization of the crude product from acetonitrile resulted in acid 10b (R9 = phenyl) (1 g, 86%).
[0408] MS (ESNEG): 198 [M-H]"; 1H NMR (300 MHz, CD3OD) δ 7.64-7.67 (m, 3), 8.02- 8.06 (m, 2), 8.53-8.56 (m, 1), 8.82 (s, 1), 8.82-8.90 (m, 1).
[0409] To the amine 2b (R^Me, R =Me) (102 mg, 0.42 mmol) in DMF (5 mL), BSTFA (0.549 mL, 2.1 mmol) and triethylamine (0.183 mL, 1.26 mmol) was added at 0 °C and then stirred at room temperature for 3 hours. Acid 10b (R9=phenyl) (158 mg, 0.80 mmol) and HATU (302 mg, 0.80 mmol) were added and the reaction was stirred for an additional 4 hours at room temperature. The DMF was removed and the residue was extracted with ethyl acetate (100 mL) and washed with saturated, bicarbonate (40 mL). The product obtained on removal of solvent was taken up in methanol and treated with Dowex H1" resin for 1 hour. After filtering the resin, methanol was removed to obtain the crude product. The resulting residue was then purified by silica gel chromatography using 10% methanol in dichloromethane as eluent to provide compound llb(R1=Me, R2=Me R3=H, R9' = phenyl) (50 mg, 58 %). [0410] TLC: Rf = 0.70 (10% MeOH/DCM); MS (ESPOS): 435 [M+H ; 1H NMR (300 MHz, CDC13) δ 1.01 (t, J = 6.6, 6), 2.12 (s, 3), 2.28 (m, 1), 3.56 (dd, J = 3.3, 10.5, 1), 3.90 (d, J = 3.3, 1), 4.12 (dd, J = 5.4, 10.5, 1), 4,27-4,36 (m, 2), 4.52 (m, 2), 5.26 (d, J = 5.7, 1), 7.48-7.55 (m, 3), 7.77-7.80 (m, 2), 7.83-7.85 (m, 1), 8.37 (s, 1), 8.69 (d, J = 5.4, 1). [0411] To compound 1 lb (R*=Me, R2=Me R3=H, R9' = phenyl) (40 mg, 0.09 mmol) in methanol (5 mL), water (10 mL) and acetic acid (5 mL), platinum dioxide (100 mg, 0.44 mmol) was added and the reaction mixture shaken at 60 psi hydrogen for 16 h. The catalyst was removed by filtration, and the solvent was evaporated to obtain the crude product, which was then purified on silica gel column chromatography using 10%) methanol in dichloromethane to provide the title compound (10 mg, 25%).
[0412] TLC: Rf = 0.22 (20% MeOH/DCM); MS (ESPOS): 447 [M+H]+; 1H NMR (300 MHz, CD3OD) δ 0.90 (d, J = 6.8, 6), 0.93-1.05 (m, 5), 1.20 (m, 6), 1.33-1.47 (m, 4), 1.75 (m, 6), 2.10 (s, 3), 2.18-2.22 (m, 1), 2.97 (t, J = 12.3, 1), 3.39-3.52 (m, 2), 3.70-3.78 (m, 2), 4.05-4.21 (m, 3), 5.23 (d, J = 5.7, 1).
Example 31 Preparation of 4-Ethyl-l-(2-hydroxy-ethyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0413] To the product of Example 1 (28 mg, 0.07 mmol) in methanol (2 mL), ethylene oxide (0.5 mL) was added and stirred at 4 °C overnight. The solvent was removed and the resulting product was purified by column, chromatography using 20%> MeOH in DCM as eluent to obtain the title compound (16 mg, 51%) as a white powder.
[0414] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 6, 1), 4.27 (m, 1), 4.10 (m, 2), 3.95 (m, 1), 3.79-3.50 (m, 4), 3.85 (m, 1) 3.74 (m, 1), 3.26 (m, 1), 2.91 (m, 2), 2.33 (m, 1), 2.13 (m, 4), 1.92 (m, 1), 1.71 (m, 1), 1.17 (m, 7), 0.94 (m, 9); MS (ESPOS): 435 [M+H]+.
Example 32 Preparation of l-(2-Hydroxy-ethyl)-4-pentyl-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydro--^-6-methylsu- any!-tetrahydro-pyran-2-yl)-propyI]-amide
[0415] The title compound was prepared using the procedures of Example 32 with the product from Example 25 as the starting material.
[0416] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=6.0, 1), 4.19 (m, 3), 3.79 (d, J=3.3, 1), 3.74 (m, 1), 3.65 (m, 1), 3.54 (dd, J=3.0, 10.2, 1), 3.25 (m, 2), 2.82 (m, 2), 2.14 (m, 4), 1.89 (m, 1), 1.72 (m, 1), 1.28 (m, 12), 0.94 (m, 9); MS (ESPOS): 477 [M+H]+.
Example 33 Preparation of l-(2-Hydroxy-ethyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0417] The title compound was made using the procedures of Example 32 with the product of Example 17 as the starting material.
[0418] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=6.0, 1), 4.19 (dd, J=9.6; 3.3, 1), 4.11 (m, 2), 3.79 (d, J=3.3, 1), 3.75 (m, 1), 3.65 (m, 1), 3.54 (m, 1), 3.28 (m, 1), 2.82 (m, 2), 2.27 (m, 5) 1.90 (m, 1), 1.71 (m, 1), 1.36 (m, 8), 0.94 (m, 9); MS (ESPOS): 449 [M+H]+.
Example 34 Preparation of 2-[2-Methyl-l-(3,4,5-trihydrosy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidin&-l-carbosylic acid 91-f-fluoren-9-ylmet-hyl ester
[0419] To the product of Example 17 (50 mg, 0.123 mmol) in water (3 mL) and dioxane (3 mL), Fmoc-Cl (38mg, 0.197 mmol) and sodium carbonate (25 mg, 0.246 mmol) were added and the reaction mixture was stirred over night at room temperature. The solvents were removed and the crude material was loaded into a silica gel column and eluted with ethyl acetate to obtain the title compound as a white solid (30 mg, 51%).
[0420] TLC: Rf = 0.5 (EtOAc). MS (ESPOS): 627 [M+H]+, 649 [M+Naf; 1H NMR (CD3OD, 200 MHz): 7.79 (d, J = 4.6 Hz, 2), 7.59-7.62 (m, 2), 7.28-7.41 (m, 4), 5.19 (d, J = 3.8 Hz, 1), 4.45 (m, 2), 4.24 (t, J = 4.2, 1), 3.99-4.15 (m, 4), 3.93 (m, 1), 3.47-3.50 (m, 2), 2.05 (s, 3), 1.87 (m, 1), 1.67 (s, 2), 1.50 (m, 1), 1.30 (m, 4), 0.86-0.91 (m, 9).
Example 35 Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methyl-sulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine-l-carboxyIic acid ethyl ester
[0421] To the product of Example 17 (50 mg, 0.123 mmol) in water (3 mL) and dioxane (3 mL), ethyl chloroformate (20 mg, 0.147 mmol) and sodium carbonate (25 mg, 0.246 mmol) were added and stirred over night at room temperature. The solvents were removed and the crude material was loaded into a silica gel column and eluted with ethyl acetate to obtain the title compound as a white solid (40 mg, 52%).
[0422] TLC: Rf = 0.28 (EtOAc). MS (ESPOS): 477 [M+H]+; 499 [M+Naf; 1H NMR (CD3OD, 200 MHz): 5.22 (d, J = 3.6 Hz, 1), 4.27 (m, 1), 4.03-4.14 (m, 5), 3.96 (bs, 1), 3.62 (m, 1), 3.54 (d, J = 2.2 Hz, 1), 3.52 (d, J = 2.2 Hz, 1), 2.08 (s, 3), 1.93-2.03 (m, 2), 1.75-1.85 (m, 3), 1.61 (m, 2), 1.33 (m, 4), 1.22-1.28 (m, 3), 0.90-0.94 (m, 9).
Example 36 Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid phenyl ester
[0423] To the product of Example 17 (50 mg, 0.123 mmol) in water (3 mL) and dioxane (3 mL), phenyl chloroformate (40 mg, 0.246 mmol) and sodium carbonate (25 mg, 0.246 mmol) were added and the reaction mixture was stirred over night at room temperature. The solvents were removed and the crude material was loaded into a silica gel column and eluted with ethyl acetate to obtain the title compound as a white solid (30 mg, 47%>). [0424] TLC: Rf = 0.4 (EtOAc). MS (ESPOS): 526 [M+H]+, 548 [M+Naf; 1H NMR (CD3OD, 200 MHz): 7.36 (t, J = 3.8 Hz, 2), 7.17-7.23 (m, 10), 7.10 (d, J = 3.6 Hz, 2), 5.20 (d, J = 3.6 Hz, 1), 4.09 (m, 3), 3.93 (d, J = 2.2 Hz, 1), 3.82 (m, 2), 3.46 (m, 2), 2.01 (s, 3), 2.00 (m, 1), 1.71 (m, 1), 1.46-1.36 (m, 4), 0.96-0.90 (m, 9).
Example 37 Preparation of 4-(4,4-Difluoro-pentyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydrosy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0425] To a solution of aldehyde 8a, prepared by first step in Method L, (510 mg, 1.47 mmol, 1 equiv) in benzene (8 mL) was added l-friphenylphosphoranylidene-2-propanone (Aldrich) (702 mg, 2.2 mmol, 1.5 equiv). The reaction mixture was refluxed overnight and the solvent was removed under vacuum. The residue was purified by chromatography to give 4-(4- oxo-pent-2-enyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester as an oil (237 mg, 42 %).
[0426] MS (ESPOS): 410.2 [M + Naf , 288.3 [M - Boc + H]+; MS (ESNEG): 386.2 [M-H]". [0427] To a solution of 4-(4-oxo-pent-2-enyl)-pyrrolidine- 1 ,2-dicarboxylic acid 1 -tert-butyl ester 2-methyl ester (193 mg, 0.5 mmol, 1 equiv) in benzene (0.9 mL) was added a solution of friphenylphosphine-copper (I) hydride hexamer in benzene (3.6 mL). The mixture was stirred at rt overnight and hexane (13 mL) was added. The mixture was filtered and the filtrate was evaporated. The residue was purified by chromatography to give 4-(4-oxo-pentyl)-pyrrolidine- 1,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (127 mg, 65 %). [0428] 1H NMR (300 MHz, CDC13) δ 7.35-7.30 (m, 5), 5.25-5.04 (m, 2), 4.42-4.25 (m, 1), 3.77-3.62 (m, 1), 3.00-2.85 (m, 1), 2.39 (t, J = 7, 2), 2.34-1.47 (m, 7), 2.10 (s, 3), 1.43 (s, 3H), 1.31 (s, 6H); MS (ESPOS): 412.3 [M + Naf, 290.3 [M - Boc + H]+; MS (ESNEG): 388.4 [M-
H]-.
[0429] To a solution of 4-(4-oxo-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (155 mg, 0.40 mmol, 1 equiv) in dichloromethane (1.5 mL) at -78 °C was added DAST (0.21 mL, 1.60 mmol, 4 equiv). The reaction mixture was warmed to rt and stirred at rt for 3 h, followed by additional DAST (0.32 mL, 2.4 mmol, 6 equiv) at -78 °C. The mixture was warmed to rt and stirred overnight. Then the mixture was diluted with dichloromethane, washed with sat. aqueous NaHC03 (lx), dried, and evaporated. The residue was purified by chromatography to give 4-(4,4-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester as a yellow oil (88 mg, 54 %).
[0430] MS (ESPOS): 434.2 [M + Naf, 312.3 [M - Boc + H]+.
[0431] To a solution of 4-(4,4-difluoro-pentyl)-pyrrolidine- 1 ,2-dicarboxylic acid 1 -tert-butyl ester 2-methyl ester (88 mg, 0.21 mmol, 1 equiv) in THF (1.2 mL) and water (0.4 mL) was added lithium hydroxide monohydrate (45 mg, 1.07 mmol, 5 equiv). The reaction mixture was stirred at rt overnight. The THF was removed under vacuum. The residue was diluted with water and washed with ether. The aqueous layer was taken up in ethyl acetate, partitioned with
10% citric acid. The organic layer was washed with water (1 x), brine (1 x), dried and concentrated to give 4-(4,4-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester
(66 mg, 96 %).
[0432] 1H NMR (300 MHz, CDC13) δ 4.39-4.34 (m, 1), 3.57-3.48 (m, 1), 2.92-2.83 (m, 1),
2.57-2.50 (m, 1), 2.30-2.18 (m,l), 1.91-1.73 (m, 3), 1.64-1.36 (m, 7), 1.48 (s, 9); MS (ESPOS):
344.3 [M + Naf, 222.3 [M - Boc + H]+; MS (ESNEG): 320.2 [M-H]".
[0433] To a solution of compound 2b (R1=Me, R2=Me) (50 mg, 0.20 mmol, 1 equiv) in dry
DMF (0.5 mL) at 0 °C was added triethylamine (88.3 up, 0.64 mmol, 3.2 equiv), followed by the addition of BSTFA (79.2 μL, 0.30 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. The reaction mixture was added to the acid (66 mg, 0.21 mmol, 1 equiv) in a 25 mL round bottom flask, followed by the addition of
HATU (96.8 mg, 0.25 mmol, 1.25 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid, water, sat. NaHC03 and brine. The organic layer was dried over Na S0 and evaporated and used in the next step without additional purification.
[0434] To a solution of the above Boc protected lincosamide in DCM (9 mL) with methyl sulfide (0.20 mL) were added trifluoroacetic acid (3 mL) and water (0.20 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound
(68 mg, 75 %) as a white solid.
[0435] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.4, 1), 4.16 (dd, J = 3.3, 9.9, 1), 4.11-
4.00 (m, 3), 3.75 (d, J = 3.3, 1), 3.51 (dd, J = 3.3, 10.2, 1), 3.40-3.32 (m, 1), 2.71 (dd, J = 8.2,
10.6, 1), 2.23-2.05 (m, 3), 2.10 (s, 3), 1.98-1.76 (m, 3), 1.63-1.39 (m, 7), 0.94-0.87 (m, 6). MS
(ESPOS): 455.3 [M+Hf . Example 38 Preparation of 4-(3,3-Difluoro-butyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0436] Ethyl triphenylphosphonium bromide (Aldrich) (2.92 g, 7.86 mmol, 3.9 equiv) and potassium-t-butoxide (0.61 g, 5.44 mmol, 2.7 equiv) were suspended in toluene (26 mL) under nitrogen with vigorous stirring. After 4 h, a solution of aldehyde 8a prepared by the first step in Method L (700 mg, 2.01 mmol, 1 equiv) in toluene (17 mL) was added dropwise. The reaction mixture was stirred at rt for 2 h and diluted with ethyl acetate (150 mL). The organic layer was washed with water (2x), brine,. dried and concentrated. The residue was purified by ■ chromatography to give a clear oil 4-but-2-enyl-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (360 mg, 50 %). [0437] MS (ESPOS): 260.3 [M+H - Bocf .
[0438] To a solution of 4-but-2-enyl-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2- methyl ester (149 mg, 0.42 mmol, 1 equiv) in DMF (1.4 mL) and water (0.2 mL) were added palladium(II)chloride (7.4 mg, 0.042 mmol, 0.1 equiv) and copper(I) chloride (41.1 mg, 0.42 mmol, 1 equiv). The mixture was stirred at 50 °C overnight with oxygen bubbling into the mixture. The mixture was filtered and the filtrate was concentrated under high vacuum. The residue was diluted with ethyl acetate, washed with water (lx), brine (lx), dried and concentrated. The residue was purified by preparative TLC to give 4-(3-oxo-butyl)-pyrrolidine- 1,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (110 mg, 71 %>). [0439] 1H NMR (300 MHz, CDC13) δ 7.35-7.28 (m, 5), 5.24-5.03 (m, 2), 4.43-4.25 (m, 1), 3.75-3.61 (m, 1), 3.01-2.87 ( , 1), 2.44-2.35 (m,2), 2.28-2.15 (m, 1), 2.11 (s, 3), 2.09-1.98 (m, 1), 1.91-1.51 (m, 3), 1.43 (s, 3.4H), 1.31 (s, 5.6H). MS (ESPOS): 398.3 [M + Naf, 276.3 [M - Boc + H] . [0440] To a solution of the 4-(3 -oxo-butyl)-pyrrolidine- 1 ,2-dicarboxylic acid 1 -tert-butyl ester 2-methyl ester (110 mg, 0.29 mmol, 1 equiv) in dichloromethane (1.1 mL) at -78 °C was added DAST (0.16 mL, 1.17 mmol, 4 equiv). The reaction mixture was warmed to rt and stirred at rt for 3 h, followed by additional DAST (0.23 mL, 1.76 mmol, 6 equiv) at -78 °C. The mixture was warmed to rt and stirred overnight. Then the mixture was diluted with dichloromethane, washed with sat. aqueous NaHC03 (lx), dried, evaporated. The residue was purified by chromatography to give 4-(3,3-Difluoro-butyl)-pyrrolidine-l,2-dicarboxylic acid 1- tert-butyl ester 2-methyl ester (92.7 mg, 80 %>). [0441] MS (ESPOS): 420.3 [M + Naf.
[0442] To a mixture of 4-(3 ,3 -difluoro-butyl)-pyrrolidine- 1 ,2-dicarboxylic acid 1 -tert-butyl ester 2-methyl ester (92.7 mg, 0.23 mmol, 1 equiv) in THF (1.2 mL) and water (0.4 mL) was added lithium hydroxide monohydrate (49 mg, 1.17 mmol, 5 equiv). The reaction mixture was stirred at rt overnight, the THF was removed under vacuum. The residue was diluted with water, washed with ether. The aqueous layer was taken up in ethyl acetate, partitioned with 10%> citric acid. The organic layer was washed with water (1 x), brine (1 x), dried and concentrated to give a white solid, 4-(3,3-difluoro-butyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester (59.7 mg," 83 %). -
[0443] ■ 1H NMR (300 MHz, CDC13) δ 4.40-4.36 (m, 1), 3.59-3.52 (m, 1), 2.94-2.86 (m, 1); 2.55-2.48 (m, 1), 2.33-2.15 (m, 1), 1.92-1.73 (m, 3), 1.66-1.40 (m, 5), 1.47 (s, 9); MS (ESPOS): 330.2 [M + Naf, 208.2 [M - Boc + H]+ ; MS (ESNEG): 306.1 [M-H]". [0444] To a solution of compound 2b (R1=Me, R2=Me) (50 mg, 0.20 mmol, 1 equiv) in dry DMF (0.5 mL) at 0 °C was added triethylamine (88.3 up, 0.64 mmol, 3.2 equiv), followed by the addition of BSTFA (79.2 μL, 0.30 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. The reaction mixture was added to the acid 4-(3,3-difluoro-butyl)-pyrrolidine-l,2-diearboxylic acid 1-tert-butyl ester (59.7 mg, 0.20 mmol, 1 equiv) in a 25 mL round bottom flask, followed by the addition of HATU (93.3 mg, 0.25 mmol, 1.25 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid, water, sat. NaHC03 and brine. The organic layer was dried over Na S04 and evaporated to give a syrup. [0445] To a solution of the above syrup in DCM (9 mL) with methyl sulfide (0.20 mL) were added trifluoroacetic acid (3 mL) and water (0.20 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (63 mg, 72 %) as a white solid.
[0446] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.22-4.13 (m, 2), 4.10-4.04 (m,
2), 3.76 (d, J = 2.4, 1), 3.54-3.42 (m, 2), 2.84-2.76 (m, 1), 2.29-1.83 (m, 5), 2.10 (s, 3), 1.67-1.51
(m, 6), 0.95-0.87 (m, 6). MS (ESPOS): 441.3 [M+H]+
Example 39 Preparation of 4-(3,3-Difluoro-pentyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0447] To a solution of compound 7c (R9 =2-penteneyl) prepared using methods of Method K (323.7 mg, 0.87 mmol, 1 equiv) in DMF (2.8 mL) and water (0.4 mL) at 0 °C were added palladium (II) chloride (15.4 mg, 0.087 mmol, 0.1 equiv) and copper(I) chloride (85.9 mg, 0.87 mmol, 1 equiv). The mixture was stirred at 50 °C overnight with oxygen bubbling into the mixture. The mixture was filtered and the filtrate was concentrated under high vacuum. The residue was diluted with ethyl acetate, washed with water (lx), brine (lx), dried and concentrated. The residue was purified by preparative TLC to provide 4-(3-oxo-pentyl)- pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (242 mg, 72 %). [0448] MS (ESPOS): 412.3 [M + Naf, 290.3 [M - Boc + H]+.
[0449] To 4-(3-oxo-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (242 mg, 0.62 mmol, 1 equiv) in dichloromethane (2.3 mL) at -78 °C was added DAST (0.33 mL, 2.49 mmol, 4 equiv). The reaction mixture was warmed to rt and stirred at rt for 3 h, followed by an addition of more DAST (0.49 mL, 3.73 mmol, 6 equiv) at -78 °C. The mixture was warmed to rt and stirred overnight. Then the mixture was diluted with dichloromethane, washed with sat. aqueous NaHCθ3 (lx), dried, evaporated. The residue was purified by chromatography to 4-(3,3-Difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2- methyl ester (117 mg, 46%).
[0450] !H NMR (300 MHz, CDC13 ) δ 7.35-7.26 (m, 5), 5.25-5.04 (m, 2), 4.44-4.27 (m, 1), 3.79-3.64 (m, 1), 3.02-2.89 (m, 1), 2.32-2.17 (m,l), 2.13-2.02 (m, 1), 1.91-1.68 (m, 5), 1.57-1.47 (m, 2), 1.44 (s, 3.5H), 1.31 (s, 5.5H), 0.97 (t, J = 7.5, 3). MS (ESPOS): 434.3 [M + Naf, 312.3 [M - Boc + H]+.
[0451] To a solution of 4-(3,3-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester 2-methyl ester (106 mg, 0.26 mmol, 1 equiv) in THF (2.4 mL) and water (0.8 mL) was added lithium hydroxide monohydrate (54 mg, 1.29 mmol, 5 equiv). The reaction mixture was stirred at rt overnight. THF was removed under vacuum. The residue was diluted with water (10 mL), washed with ether (20 mL). The aqueous layer was taken up in ethyl acetate (50 mL), partitioned with 10% citric acid (25 mL). The organic layer was washed with water (1 x), brine (1 x), dried and concentrated to give 4-(3,3-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1- tert-butyl ester as a clear oil (82.1 mg, 99%).
[0452] 1HNMR (300 MHz, CDCl3) δ 4.40-4.36 (m, 1), 3.58-3.51 (m, 1), 2.94-2.86 (m, 1), 2.57-2.51 (m, 1), 2.30-2.15 (m,l), 1.92-1.72 (m, 5), 1.62-1.53 (m, 2), 1.48 (s, 9), 0.99 (t, J = 7.5, 3); MS (ESPOS): 344.3 [M +Naf , 222.3 [M - Boc + H]+.
[0453] To a solution of compound 2b (R1=Me, R2=Me) (50 mg, 0.20 mmol, 1 equiv) in dry DMF (0.5 mL) at 0 °C was added triethylamine (88.3 μL, 0.64 mmol, 3.2 equiv), followed by the addition of BSTFA (79.2 μL, 0.30 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. The reaction mixture was added to the acid 4-(3,3-difluoro-pentyl)-pyrrolidine-l,2-dicarboxylic acid 1-tert-butyl ester (76.6 mg, 0.24 mmol, 1.2 equiv) in a 25 mL round bottom flask, followed by the addition of HATU (111.9 mg, 0.29 mmol, 1.5 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate (60 mL), washed with 10% citric acid (30 mL), water (30 mL), sat. NaHC03 (30 mL) and brine. The organic layer was dried over Na2S04 and evaporated to give a yellow oil.
[0454] To a solution of the above oil in DCM (9 mL) with methyl sulfide (0.20 mL) were added trifluoroacetic acid (3 mL) and water (0.20 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (72 mg, 80%) as a white solid. [0455] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.20-4.04 (m, 4), 3.76 (d, J = 2.7, 1), 3.51 (dd, J = 3.4, 10.3, 1), 3.43 (dd, J = 6.9, 10.8, 1), 2.77 (dd, J = 8.4, 10.8, 1), 2.30-2.05 (m, 3), 2.10 (s, 3), 2.03-1.76 (m, 5), 1.64-1.54 (m, 2), 1.03-0.89 (m, 9); MS (ESPOS): 455.4
Example 40 Preparation of 4-(3,3-Difluoro-pentyl)-l-(2-hydroxy-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0456] To a solution of the product of Example 42 (17.9 mg, 0.039 mmol) in MeOH (2 mL) at 0 °C was added ethylene oxide (0.4 mL). The reaction mixture was stirred at 4 °C overnight. The reaction mixture was concentrated and purified by chromatography to give the title compound as a white solid (8.2 mg, 42%).
[0457] 1H NMR (300 MHz, CD3OD) δ 5.23 (d, J = 5.7, 1), 4.13-4.05 (m, 3), 3.75 (d, J = 3.6, 1), 3.72-3.57 (m, 2), 3.53 (dd, J = 3.3, 10.2, 1), 3.41-3.36 (m, 1), 3.22 (dd, J = 3.3, 10.8, 1), 2.88- 2.78 (m, 1), 2.63-2.54 (m, 1), 2.18-1.99 (m, 4), 2.10 (s, 3), 1.93-1.75 (m, 5), 1.57-1.46 (m, 2), 1.01-0.90 (m, 9); MS (ESPOS): 499.6 [M+H] +; MS (ESNEG): 497.5 [M-H]".
Example 41 Preparation of 4-(3,3-Difluoro-propyl)-piperidine-2-carboxylic add [2-methyl-l-(3,4,5- trilιydro-sy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0458] Compound 14c (R9 = 2,2-difluoroethyl) is prepared using the methods described in Method R.
[0459] To a dry flask were added compound 14a (1.4 g, 5.32 mmol, 1 equiv), , triphenylphosphine (111.6 mg, 0.43 mmol, 0.08 equiv), copper (I) iodide (81 mg, 0.43 mmol, 0.08 equiv), palladium acetate (47.7 mg, 0.21 mmol, 0.04 equiv) and triethylamine (20 mL). The mixture was deaerated with nitrogen, followed by addition of propiolaldehyde diethyl acetyl (1.36 g, 10.65 mmol, 2 equiv). The mixture was stirred at rt for 3 hrs. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give a yellow oil, compound 14b (R9 = 3,3-Diethoxy-prop-l-ynyl) (1.4 g, 100%>). [0460] 1HNMR (300 MHz, CDC13) δ 8.69 (dd, J = 0.8, 5.0, 1), 8.15 (dd, J = 0.8, 1.4, 1), ■ 7.49 (dd, J = 1.7, 5.0, 1), 5.48 (s, 1), 3.99 (s, 3), 3.82-3.73 (m, 2), 3.71-3.62(m, 2), 1.26 (t, J = 7.2, 6). MS (ESPOS): 264.5 [M+Hf .
[0461] To a solution of 14b (R9 = 3,3-Diethoxy-prop-l-ynyl) (1.4 g, 5.32 mmol) in methanol (100 mL) was added 10 % palladium on carbon (0.3 g). The mixture was purged and charged with hydrogen (1 atm) and shaken at rt overnight. The palladium was removed by filtration and the filtrate was concentrated to give 14c (R9=3,3-diethoxy propyl) as an oil (1.39 g, 98%>). [0462] 1H NMR (300 MHz, CDC13) δ 8.60 (d, J = 5.1, 1), 7.98 (d, J = 0.9, 1), 7.31-7.28 (m, 1), 4,45 (t, J = 5.4, 1), 3.98 (s, 3), 3.72-3.58 (m, 2), 3.52-3.39(m, 2), 2.79-2.72 (m, 2), 1.99-1.90 (m, 2), 1.22-1.15 (m, 6).
[0463] To a mixture of 14c (R9=3,3-diethoxy propyl) (0.68 g, 2.55 mmol) in acetic acid (8 mL) and water (2 mL) was added cone, hydrochloric acid (2 drops). The mixture was stirred at rt overnight and the solvent was removed under high vacuum. The residue was diluted with ethyl acetate, washed with sat. sodium bicarbonate (lx), brine (lx). The organic layer was dried and concentrated to give 4-(3-Oxo-propyl)-pyridine-2-carboxylic acid methyl ester as a yellow oil (0.27 g, 55 %). [0464] To a solution of aldehyde 4-(3-oxo-propyl)-pyridine-2-carboxylic acid methyl ester (0.27 g, 1.4 mmol, 1 equiv) in DCM (5 mL) at -78 °C was added DAST (0.91 g, 5.6 mmol, 4 equiv). The mixture was warmed to rt and stirred overnight. The mixture was diluted with dichloromethane (60 mL), washed with sat. aqueous NaHC03 (lx), dried, and evaporated. The residue was purified by prep.
[0465] TLC (5% MeOH in DCM) to 4-(3,3-difluoro-ρropyl)-pyridine-2-carboxylic acid methyl ester (137 mg, 45%): 1H NMR (300 MHz, CDC13) δ 8.64 (d, J = 5.1, 1), 8.00-7.98 (m, 1), 7.33-7.29 (m, 1), 5.85 (dddd, J = 4.1, 4.1, 56.4, 56.4, 1), 3.99 (s, 3), 2.90-2.83 (m, 2), 2.28- 2.09 (m, 2); MS (ESPOS): 216.4 [M+Hf .
[0466] To a solution of 4-(3,3-difluoro-propyl)-pyridine-2-carboxylic acid methyl ester (130 mg, 0.6 mmol) (or compound 14c (R9 = 2,2-difluoroethyl) prepared in the previous steps) in MeOH (3 mL) and water (3 mL) were added cone. HCl (0.25 mL, 3.0 mmol, 5 equiv) and platinum oxide (65 mg). The mixture was purged and charged with hydrogen (1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to give a clear syrup. To the above residue were added 2N NaOH (1.21 mL) and t-butyl alcohol (0.7 mL). The mixture was stirred at rt for 2 hrs. Then di-t-butyl dicarbonate (0.16 g, 0.73 mmol) was added. The mixture was stirred at rt overnight. The solvent was removed under vacuum. The residue was diluted with water (10 mL), was washed with ether (20 mL). The aqueous layer was acidified with 2N HCl to pH = 2.0, and extracted with ethyl acetate (2x). The combined organic layers were dried and concentrated to give 4-(3,3-Difluoro-propyl)-piperidine-l,2- dicarboxylic acid 1-tert-butyl ester as a clear syrup (163 mg, 88 %>)
[0467] 1H NMR (300 MHz, CDC13) δ 5.77 (dddd, J = 4.2, 4.2, 56.6, 56.6, 1), 4.34 (t, J = 6.4, 1), 3.62-3.50 (m, 1), 3.41-3.30 (m, 1), 2.05-1.96 (m, 1), 1.92-1.73 (m, 4), 1.70-1.60 (m, 1), 1.52- 1.32 (m, 3), 1.43 (s, 9); MS (ESPOS): 330.5 [M + Na] +; MS (ESNEG): 306.5 [M-H]". [0468] To a mixture of the HCl salt of compound 2b (R =Me, R2=Me) (140 mg, 0.49 mmol, 1 equiv) in dry DMF (1.2 mL) at 0 °C was added triethylamine (0.34 mL, 2.43 mmol, 5 equiv), followed by the addition of BSTFA (0.20 mL, 0.74 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. To the reaction mixture were added the 4-(3,3-Difluoro-propyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester (153 mg, 0.50 mmol, 1.0 equiv) and HATU (235 mg, 0.62 mmol, 1.26 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10%) citric acid (lx), water (lx), sat. aHCθ3 (lx) and brine. The organic layer was dried over Na S0 and evaporated to give a pink syrup which was used without purification.
[0469] To a solution of the above syrup in DCM (15 mL) with methyl sulfϊde (0.33 mL) were added trifluoroacetic acid (5 mL) and water (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (lower isomer, 93 mg,
43%) as a white solid.
[0470] 1H NMR (300 MHz, CD3OD) δ 5.90 (dddd, J = 4.2, 4.2, 56.7, 56.7, 1), 5.24 (d, J =
6, 1), 4.21 (dd, J = 3.5, 9.8, 1), 4.11-4.04 (m, 2), 3.84-3.77 (m, 2), 3.51 (dd, J = 3.2, 10.3, 1),
3.45-3.37 (m, 1), 3.07-2.98 (m, 1), 2.23-2.12 (m, 2), 2.11 (s, 3), 1.98-1.66 (m, 4), 1.52-1.26 (m,
4), 0.94-0.88 (m, 6). MS (ESPOS): 441.7 [M+H]+ .
Example 42 Preparation of 4-(4,4-Difluoro-butyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0471] To a solution of methyl sulfoxide (0.58 mL, 8.16 mmol, 2.4 equiv) in dichloromethane (1.8 mL) at -72 °C was added a 2 M solution of oxalyl chloride in dichloromethane (2.04 mL, 4.08 mmol, 1.2 equiv) over a period of 1 minute. The mixture was stirred at -72 °C for 25 minutes, followed by the dropwise addition of a solution of the alcohol 14c (R9=4-hydroxybutyl), prepared using the procedures in Method R, (0.71 g, 3.4 mmol, 1 equiv) in dichloromethane (4.8 mL) over a period of 2 minutes. The reaction mixture was stirred at -72 °C for 25 minutes, then warmed to -50 °C and stirred for an additional 2 h. Triethylamine (1.89 mL, 13.6 mmol, 4.0 equiv) was added and stirred at -50 °C for 25 minutes. The mixture was diluted with ethyl acetate, washed with water (lx), sat. aqueous NaHC03 (lx), brine (l ), dried, evaporated and co-evaporated with anhydrous toluene to give aldehyde 4-(4- oxo-butyl)-pyridine-2-carboxylic acid methyl ester as an oil (0.66 mg, 94%).
[0472] 1H NMR (300 MHz, CDC13) δ 9.78 (s, 1), 8.62 (d, J = 5.1, 1), 7.97 (s, 1), 7.29 (d, J =
5.1, 1), 3.99 (s, 3), 2.72 (t, J = 7.8, 2), 2.50 (t, J = 7.2, 2), 2.04-1.93 (m, 2); MS (ESPOS): 230.4
[0473] To a solution of 4-(4-oxo-butyl)-pyridine-2-carboxylic acid methyl ester (0.66 g, 3.19 mmol, 1 equiv) in DCM (12 mL) at -78 °C was added DAST (1.69 mL, 12.75 mmol, 4 equiv).
The mixture was warmed to rt and stirred overnight. The mixture was diluted with dichloromethane, washed with sat. aqueous NaHC03 (lx), brine (lx), dried, evaporated. The residue was purified by chromatography to provide 4-(4,4-difluorobutyl)-pyridine-2-carboxylic acid methyl ester (0.54 g, 74%>).
[0474] 1H NMR (300 MHz, CDC13) δ 8.62 (d, J = 5.1, 1), 7.97-7.95 (m, 1), 7.29-7.26 (m, 1),
5.81 (dddd, J = 3.9, 3.9, 56.6, 56.6, 1), 3.98 (s, 3), 2.74 (t, J = 7.2, 2), 1.93-1.77 (m, 4). MS
(ESPOS): 230.4 [M+Hf , 252.4 [M + Naf.
[0475] To a mixture 4-(4,4-difluorobutyl)-pyridine-2-carboxylic acid methyl ester (0.54 g,
2.36 mmol, 1 equiv) in MeOH (8 mL) and water (8 mL) were added cone. HCl (0.59 mL, 7.07 mmol, 3 equiv) and platinum oxide (0.2 g). The mixture was purged and charged with hydrogen
(1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to give a residue:
[0476] MS (ESPOS): 236.6 [M+H]+ .
[0477] To the residue prepared above were added 2N NaOH (4.72 mL) and t-butyl alcohol
(2.5 mL). The mixture was stirred at rt for 2 hrs. Then di-t-butyl dicarbonate (0.77 g, 3.54 mmol) was added. The mixture was stirred at rt overnight. The solvent was removed under vacuum. The residue was diluted with water (10 mL), was washed with ether (20 mL). The aqueous layer was acidified with 2N HCl to pH = 2.0, extracted with ethyl acetate (2x). The combined organic layers were dried and concentrated to give 4-(4,4-difluoro-butyl)-piperidine-l,
2-dicarboxylic acid 1-tert-butyl ester (0.67 g, 89 %).
[0478] 1H NMR (300 MHz, CDC13) δ 5.77 (dddd, J = 4.3, 4.3, 56.8, 56.8, 1), 4.30 (t, J = 6.8,
1), 3.58-3.47 (m, 1), 3.41-3.31 (m, 1), 2.05-1.96 (m, 1), 1.87-1.68 (m, 4), 1.65-1.56 (m, 1), 1.51-
1.30 (m, 5), 1.43 (s, 9); MS (ESPOS): 344.5 [M + Naf.
[0479] To a mixture of the HCl salt of compound 2b (R1=Me, R2=Me) (153 mg, 0.53 mmol,
1 equiv) in dry DMF (1.3 mL) at 0°C was added triethylamine (0.37 mL, 2.66 mmol, 5 equiv), followed by the addition of BSTFA (0.21 mL, 0.80 mmol, 1.5 equiv). The reaction mixture was stirred at 0°C for 10 minutes, and then was stirred at rt for 50 minutes. To the reaction mixture were added the 4-(4,4-difluoro-butyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester (196 mg, 0.61 mmol, 1.15 equiv) and HATU (293 mg, 0.77 mmol, 1.45 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid (lx), water (lx), sat. NaHCO3 (lx) and brine. The organic layer was dried over Na SO and evaporated to give a syrup. The residue was dissolved in methanol (20 mL), then dried and washed Dowex resin (100 mg) was added. The mixture was stirred at rt for 30 minutes, and filtered. The filtrate was concentrated to give a clear syrup, which was purified by chromatography to give a clear syrup (0.25g, 85 %). [0480] MS (ESPOS): 555.8 [M+Hf .
[0481] To a solution of the above syrup in DCM (15 mL) with methyl sulfide (0.33 mL) were added trifluoroacetic acid (5 mL) and water (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (lower isomer, 70 mg, 34%>) as a white solid.
[0482] 1H NMR (300 MHz, CD3OD) δ 5.88 (dddd, J = 4.4, 4.4, 57, 57, 1), 5.24 (d, J = 5.4, 1), 4.20 (dd, J = 3.2, 10.1, 1), 4.12-4.03 (m, 2), 3.90-3.80 (m, 2), 3.52 (dd, J = 3.5, 10.3, 1), 3.46- 3.39 (m, 1), 3.09-2.98 (m, 1), 2.25-2.12 (m, 2), 2.11 (s, 3), 1.98-1.67 (m, 4), 1.56-1.30 (m, 6), 0.95-0.87 (m, 6); MS (ESPOS): 455.7 [M+H]+.
Example 43 Preparation of 4-(5,5-Difluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- tri-hydro-sy-6-methy-I-s-ϊlfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0483] Method R is used to prepare compound 14c (R9' = 5-hydroxypentyl) (To a dry flask was added compound 14a (2 g, 7.60 mmol, 1 equiv), triphenylphosphine (159.4 mg, 0.61 mmol, 0.08 equiv), copper (I) iodide (115.8 mg, 0.61 mmol, 0.08 equiv), palladium acetate (68.2 mg, 0.30 mmol, 0.04 equiv) and triethylamine (28 mL). The mixture was deaerated with nitrogen, followed by addition of 4-pentyn-l-ol (1.28 g, 15.21 mmol, 2 equiv). The mixture was stirred at rt overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 14b (R9' = 5-hydroxypent-l-yn-yl). [0484] To a solution of 14b (R9' = 5-hydroxypent-l-yn-yl) in methanol (60 mL) was added 10% palladium on carbon (0.62 g). The mixture was purged and charged with hydrogen (1 atm) and stirred at rt overnight. The palladium was removed by filtration and the filtrate was concentrated to give a yellow oil 14c (R9 = 5-hydroxypentyl) (1.34 g, 79 %). [0485] 1H NMR (300 MHz, CDC13) δ 8.62 (d, J = 4.8, 1), 7.97 (s, 1), 7.31 (dd, J = 1.6, 5, 1), 3.99 (s, 3), 3.63 (t, J = 6.5, 2), 2.70 (t, J = 7.7, 2), 1.74-1.53 (m, 4), 1.46-1.34 (m, 2). [0486] To a solution of methyl sulfoxide (0.46 mL, 6.42 mmol, 2.6 equiv) in dichloromethane (1.4 mL) at -72 °C was added a 2 M solution of oxalyl chloride in dichloromethane (1.61 mL, 3.21 mmol, 1.3 equiv) over a period of 1 minute. The mixture was • stirred at -72 °C for 25 minutes, followed by the dropwise addition of a solution of pyridine 14c (R9 = 5-hydroxypentyl) (0.55 g, 2.47 mmol, 1 equiv) in dichloromethane (3.8 mL) over a period of 2 minutes. The reaction mixture was stirred at -72 °C for 25 minutes, then warmed to -50 °C and stirred for an additional 2 h. Triethylamine (1.48 mL, 10.7 mmol, 4.33 equiv) was added and stirred at -50 °C for 25 minutes. The mixture was diluted with ethyl acetate, washed with water (2x), sat. aqueous NaHCO3 (lx), brine (lx), dried, evaporated and co-evaporated with anhydrous toluene to 4-(5-oxo-pentyl)-pyridine-2-carboxylic acid methyl ester (0.48 mg, 88 %). [0487J 1H NMR (300 MHz, CDC13) δ 9.75 (t, J = 1.4, 1), 8.61 (d, J = 5.1, 1), 7.97-7.95 (m, 1), 7.28 (dd, J = 1.7, 5, 1), 3.99 (s, 3), 2.73-2.67 (m, 2), 2.50-2.44 (m, 2), 1.71-1.63 (m, 4). [0488] To a solution of the to 4-(5-oxo-pentyl)-pyridine-2-carboxylic acid methyl ester oil (0.48 g, 2.19 mmol, 1 equiv) in DCM (8 mL) at -78°C was added DAST (1.41 g, 8.74 mmol, 4 equiv). The mixture was warmed to rt and stirred overnight. The mixture was diluted with dichloromethane, washed with sat. aqueous NaHCO3 (lx), dried, and evaporated. The residue was purified by chromatography to 4-(5,5-Difluoro-pentyl)-pyridine-2-carboxylic acid methyl ester (278 mg, 52 %) [0489] 1H NMR (300 MHz, CDC13) δ 8.61 (dd, J = 0.6, 4.8, 1), 7.97-7.95 (m, 1), 7.28 (dd, J = 1.5, 4.8, 1), 5.78 (dddd, J = 4.3, 4.3, 57, 57, 1), 3.99 (s, 3), 2.70 (t, J = 7.7, 2), 1.94-1.66 (m, 4), 1.55-1.43 (m, 2). [0490] MS (ESPOS): 244.2 [M+Hf
[0491] To a mixture of 4-(5,5-difluoro-pentyl)-pyridine-2-carboxylic acid methyl ester (278 mg, 1.14 mmol) in MeOH (5 mL) and water (5 mL) were added cone. HCl (0.286 mL, 3.43 mmol, 3 equiv) and platinum oxide (140 mg). The mixture was purged and charged with hydrogen (1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to 4-(5,5-difluoro-pentyl)-piperidine-2-carboxylic acid 2-methyl ester. [0492] MS (ESPOS): 250.2 [M+H]+.
[0493] To the above residue 4-(5,5-difluoro-pentyl)-piperidine-2-carboxylic acid-2-methyl ester were added 2N NaOH (2.3 mL) and t-butyl alcohol (1.2 mL). The mixture was stirred at rt for 2 hrs. Then di-t-butyl dicarbonate (0.37 g, 1.72 mmol) was added. The mixture was stirred at rt overnight. The solvent was removed under vacuum. The residue was diluted with water, was washed with ether. The aqueous layer was acidified with 2N HCl to pH = 2.0, extracted with ethyl acetate (2x). The combined organic layers were dried and concentrated to 4-(5,5- Difluoro-pentyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester (310 mg, 81 %). [0494] 1H NMR (300 MHz, CD3OD) δ 5.84 (dddd, J = 4.5, 4.5, 57, 57, 1), 4.31 (t, J = 6.3, 1), 3.65-3.56 (m, 1), 3.35-3.25 (m, 1), 2.03-1.63 (m, 5), 1.48-1.30 (m, 8), 1.43 (s, 9). [0495] To a mixture of the HCl salt of compound 2b (R!=Me, R2=Me) (223.7 mg, 0.78 mmol, 1 equiv) in dry DMF (1.9 mL) at 0°C was added triethylamine (0.54 mL, 3.89 mmol, 5 equiv), followed by the addition of BSTFA (0.31 mL, 1.17 mmol, 1.5 equiv). The reaction mixture was stirred at 0°C for 10 minutes, and then was stirred at rt for 50 minutes. To the reaction mixture were added 4-(5,5-difluoro-pentyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester (272 mg, 0.81 mmol, 1.05 equiv) and HATU (391 mg, 1.03 mmol, 1.32 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid (lx), water (lx), sat. NaHCO3 (lx) and brine. The organic layer was dried over Na SO and evaporated to give a residue. The residue was dissolved in methanol (30 mL), then dry and washed Dowex resin (150 mg) was added. The mixture was stirred at rt for 1 h and filtered. The filtrate was concentrated to give a clear syrup, which was purified by chromatography to give a clear syrup (0.26 g, 72 %). [0496] To a solution of the above syrup in DCM (15 mL) with methyl sulfide (0.33 mL) were added trifluoroacetic acid (5 mL) and water (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (lower isomer, 40 mg, 15 %) as a white solid.
[0497] 1H NMR (300 MHz, CD3OD) δ 5.86 (dddd, J = 4.5, 4.5, 57, 57, 1), 5.24 (d, J = 5.7, 1), 4.21 (dd, J = 3.3, 9.9, 1), 4.11-4.04 (m, 2), 3.86-3.78 (m, 2), 3.51 (dd, J = 3.5, 10.4, 1), 3.47- 3.38 (m, 1), 3.07-2.97 (m, 1), 2.23-2.12 (m, 2), 2.11 (s, 3), 1.98-1.64 (m, 4), 1.50-1.27 (m, 8), 0.94-0.87 (m, 6); MS (ESPOS): 469.4 [M+H]+.
Example 44 Preparation of 4-(5-Fluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0498] To a solution of compound 14c (R9= 5-hydroxypentyl) (0.66 g, 2.96 mmol, 1 equiv), prepared as described in Method R and in the synthesis of Example 47, in DCM (11 mL) at - 78°C was added DAST (1.91 g, 11.85 mmol, 4 equiv). The mixture was warmed to rt and stirred overnight. The mixture was diluted with dichloromethane, washed with sat. aqueous NaHC03 (lx), dried, and evaporated. The residue was purified by chromatography to give 4-(5- fluoro-pentyl)-pyridine-2-carboxylic acid methyl ester (254 mg, 38 %) [0499] 1H NMR (300 MHz, CDC13) δ 8.62 (d, J = 4.8, 1), 7.97 (d, J = 1.2, 1), 7.30 (dd, J = 1.7, 5, 1), 4.50 (t, J = 5.9, 1), 4.34 (t, J = 6, 1), 3.99 (s, 3), 2.70 (t, J = 7.7, 2), 1.80-1.62 (m, 4), 1.50-1.41 (m, 2).
[0500] To a mixture of 4-(5-fluoro-ρentyl)-pyridine-2-carboxylic acid methyl ester (254 mg, 1.13 mmol) in MeOH (5 mL) and water (5 mL) were added cone. HCl (0.28 mL, 3.39 mmol, 3 equiv) and platinum oxide (130 mg). The mixture was purged and charged with hydrogen (1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to give 4-(5-fluoro-pentyl)-piperidine-2-carboxylic acid 2-methyl ester. [0501] MS (ESPOS): 232.4 [M+Hf .
[0502] To 4-(5-fluoro-pentyl)-piperidine-2-carboxylic acid 2-methyl ester was added 2N NaOH (2.43 mL) and t-butyl alcohol (1.3 mL). The mixture was stirred at rt for 2 hrs. Then di- t-butyl dicarbonate (0.40 g, 1.82 mmol) was added. The mixture was stirred at rt overnight. The solvent was removed under vacuum. The residue was diluted with water, was washed with ether. The aqueous layer was acidified with 2N HCl to pH = 2.0, extracted with ethyl acetate (2x). The combined organic layers were dried and concentrated to give 4-(5-fluoro-pentyl)- piρeridine-l,2-dicarboxylic acid 1-tert-butyl ester as a syrup (254 mg, 71 %). [0503] 1HNMR (300 MHz, CDC13) δ 4.52-4.06 (m, 3), 3.55-3.30 (m, 2), 2.03-1.94 (m, 1), 1.81-1.54 (m, 4), 1.45-1.20 (m, 8), 1.43 (s, 9). MS (ESPOS): 218.3 [M + Na - Boc] + [0504] To a mixture of the HCl salt of compound 2b (R*=Me, R2=Me) (213.8 mg, 0.74 mmol, 1 equiv) in dry DMF (1.8 mL) at 0 °C was added triethylamine (0.52 mL, 3.72 mmol, 5 equiv), followed by the addition of BSTFA (0.30 mL, 1.12 mmol, 1.5 equiv). The reaction mixture was stirred at 0 °C for 10 minutes, and then was stirred at rt for 50 minutes. To the ■ * reaction mixture were added the 4-(5-fluoro-pentyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester as a syrup (244 mg, 0.77 mmol, 1.04 equiv) and HATU (370 mg, 0.97 mmol, 1.31 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated to dryness, taken up in ethyl acetate, washed with 10% citric acid (lx), water (lx), sat. NaHCθ3 (lx) and brine. The organic layer was dried over Na S0 and evaporated to give a residue. The residue was dissolved in methanol (30 mL), then dry and washed Dowex resin (140 mg) was added. The mixture was stirred at rt for 1 h and filtered. The filtrate was concentrated to give a clear syrup, which was purified by chromatography to give a clear syrup (212 mg, 52 %). [0505] To a solution of the above syrup in DCM (15 mL) with methyl sulfide (0.33 mL) were added trifluoroacetic acid (5 mL) and water (0.33 mL). The reaction mixture was stirred at rt for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the title compound (lower isomer, 40 mg, 17 %) as a white solid.
[0506] 1H MR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.49 (t, J = 5.9, 1), 4.33 (t, J = 6, 1), 4.20 (dd, J = 3.5, 10.1, 1), 4.11-4.04 (m, 2), 3.83-3.77 (m, 2), 3.51 (dd, J = 3.3, 10.2, 1), 3.44- 3.36 (m, 1), 3.06-2.94 (m, 1), 2.23-2.13 (m, 2), 2.11 (s, 3), 1.98-1.88 (m, 1), 1.77-1.59 (m, 3), 1.45-1.27 (m, 8), 0.94-0.87 (m, 6). [0507] MS (ESPOS): 451.4 [M+Hf Example 45 Preparation of 4-(4-Fluoro-butyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0508] To a solution of compound 14c (R9 = 4-hydroxybutyl) (0.76 g, 3.62 mmol, 1 equiv), prepared as described-in Method R and in the synthesis of Example 47, in DCM (14 mL) at -78 °C was added DAST (1.9 mL, 14.47 mmol, 4 equiv). The mixture was warmed to rt and stirred overnight. The mixture was diluted with dichloromethane, washed with sat. aqueous NaHCθ3 (lx), brine (lx), dried, evaporated. The residue was purified by chromatography to provide 4- (4-Fluoro-butyl)-pyridine-2-carboxylic acid methyl ester as a yellow oil (0.24 g, 31 %). [0509] 1H NMR (300 MHz, CDC13) δ 8.57 (d, J = 4.8, 1), 7.92 (d, J = 1.2, 1), 7.27-7.23 (m, 1), 4.49 (t, J = 5.6, 1), 4.33 (t, J = 5.6, 1), 3.94 (s, 3), 2.69 (t, J = 7.5, 2), 1.79-1.59 (m, 4). [0510] To a mixture of 4-(4-fluoro-butyl)-pyridine-2-carboxylic acid methyl ester (0.24 g) in THF (3 mL) and water (1 mL) was added lithium hydroxide monohydrate (71.3 mg, 1.7 mmol, 1.5 equiv). The mixture was stirred at rt overnight and diluted with methanol (20 mL). Then H1" resin was added and the mixture was shaken for 10 minutes. The resin was washed with methanol (lx), 1:1 acetonitrile/water (lx), and acetonitrile (lx). The product was eluted with 5%> TEA in methanol (4x) and acetonitrile (lx). The combined organic solvents were evaporated and co-evaporated with toluene to provide 4-(4-Fluoro-butyl)-pyridine-2-carboxylic acid (0.22 g, 65 %).
[0511] 1H NMR (300 MHz, CD3OD) δ 8.46 (d, J = 4.8, 1), 7.95 (s, 1), 7.39-7.35 (m, 1), 4.52 (t, J = 5.6, 1), 4.36 (t, J = 5.9, 1), 3.22 (q, J = 7.3, 2.5H, TEA), 2.77 (t, J = 7.5, 2), 1.84-1.62 (m, 4), 1.28 (t, J = 7.2, 3.8H, TEA). [0512] To a solution of 4-(4-fluoro-butyl)-pyridine-2-carboxylic acid (0.22 g, 0.73 mmol, 1 equiv) in dry acetonitrile (4 mL) at 0°C was added triethylamine (74 mg, 0.73 mmol, 1 equiv), followed by the addition of isobutyl chloroformate (100 mg, 0.73 mmol, 1 equiv). The reaction mixture was stirred at 0°C for 15 minutes, and then was stirred at 4°C for 2 h. To the reaction mixture was added a solution of the HCl salt of compound 2b (R1=Me, R2=Me) (263 mg, 0.91 mmol, 1.25 equiv) and triethylamine (93 mg, 0.91 mmol, 1.25 equiv) in a 1:1 acetone/water (4 mL). The reaction mixture was stirred at 4 °C overnight. The reaction mixture was evaporated to dryness, taken up in DCM, washed with sat. NaHCO3 (lx). The organic layer was dried over Na2SO and evaporated. The residue was purified by chromatography to give a clear solid (110 mg, 35 %).
[0513] To a solution of the above solid (110 mg, 0.25 mmol, 1 equiv) in MeOH (6 mL) and water (4 mL) were added cone. HCl (20.2 μL, 0.24 mmol, 0.95 equiv) and platinum oxide (220 mg). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to give a residue, which was purified by chromatography to provide the title compound (lower isomer, 33 mg, 30%>) as a white solid.
[0514] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J = 5.4, 1), 4.51 (t, J = 6, 1), 4.35 (t, J = 5.9, 1), 4.21 (dd, J = 3.3, 10.2, 1), 4.10-4.04 (m,'2), 3.93-3.80 (m, 2), 3.52 (dd, J = 3.3, 10.2, 1), 3.46- 3.38 (m, 1), 3.11-2.98 (m, 1), 2.26-2.13 (m, 2), 2.11 (s, 3), 2.00-1.92 (m, 1), 1.80-1.60 (m, 3), 1.54-1.27 (m, 6), 0.95-0.87 (m, 6). [0515] MS (ESPOS): 437.4 [M+Hf
Example 46 Preparation of 4-(3-Ethyl-3-hydro2y-peiityl)-piperidme-2-carbo-sylk acid [2-methyl-l- (3,4,5-t-rihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0516] To a dry flask was added compound 13b (R1=Me, R2=Me, and R3=H) made using
Method Q (130 mg, 0.27 mmol, 1 equiv), friphenylphosphine (45.3 mg, 0.17 mmol, 0.64 equiv), copper (I) iodide (32.9 mg, 0.17 mmol, 0.64 equiv), palladium acetate (19.4 mg, 0.086 mmol,
0.32 equiv) and triethylamine (1.5 mL). The mixture was deaerated with nitrogen, followed by addition of 3-ethyl-l-pentyn-3-ol (174 μL, 1.35 mmol, 5 equiv). The mixture was stirred at 50
°C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 13c (R1=Me, R2=Me, R3=H, R9=3-Ethyl-3-hydroxy-pent-l- ynyl).
[0517] MS (ESPOS): 467.7 [M+H]+ ; MS (ESNEG): 465.5 [M-H]".
[0518] To a mixture of the above syrup in MeOH (12 mL) and water (8 mL) were added platinum oxide (300 mg) and cone. HCl (26 μL). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filfrate was evaporated. The residue was purified by chromatography to give the title compound as a white solid (19 mg, 15 %).
[0519] 1HNMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.17 (dd, J = 3.1, 10.0, 1), 4.10-
4.02 (m, 2), 3.80 (d, J = 3, 1), 3.53-3.48 (m, 1), 3.42-3.35 (m, 1), 3.23-3.15 (m, 1), 2.75-2.64 (m,
1), 2.22-2.11 (m, 1), 2.10 (s, 3), 2.04-1.97 (m, 1), 1.80-1.72 (m, 1), 1.50-1.40 (m, 6), 1.31-1.06
(m, 5), 0.94-0.80 (m, 12); MS (ESPOS): 477.8 [M+Hf ; MS (ESNEG): 475.6 [M-H]".
Example 47 Preparation of 4-Butoxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0520] To trimethylsilyl cyanide (5.2 g, 52 mmol), 4-benzyloxy-pyridine 1 -oxide (8.8 g, 44 mmol) in DCM (20 mL) was added, followed by dimethylcarbamoyl chloride (5.6 g, 52 mmol) in DCM (10 mL), dropwise, stirred at room temperature overnight. Sodium bicarbonate (100 mL, 10%) was added, stirred for 10 minutes and extracted twice with DCM (50 mL). The combined organic layer was dried over magnesium sulfate, solvent was removed to obtain the product, compound 10a (R9 = Benzyloxy) (10.5 g, 100%).
[0521] 1H NMR (300 MHz, CDC13) δ 8.34 (d, J=5.7, 1), 7.24 (m, 5), 7.11 (t, J=2.4, 1), 6.90
(dd, J=5.7, 2.4, 1), MS (ESPOS): 211[M+H]+.
[0522] Compound 10a (R9 = Benzyloxy) (5 g, 23 mmol) was dissolved in HCl (6N, 70 mL) and refluxed overnight. The crude product 4-hydroxypyridine-2-carboxylic acid, compound 10b
(R9 = hydroxy) obtained on removal of HCl was crystallized from acetonitrile (2.6 g, 80%).
[0523] 1H NMR (300 MHz, CD3OD) δ 8.55 (d, J=6.6, 1), 7.78 (d, J=3.0, 1), 6.90 (dd, J=2.7,
6.9, 1), MS (ESNEG): 138 [M-H]".
[0524] The synthesis of title compound was completed using the synthetic sequence found in Method S starting from 4-hydroxypyridine-2-carboxylic acid, as prepared above.
[0525] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.4, 1), 4.22 (dd, J=10.2; 3.3, 1), 4.08 (m,
2), 3.81 (d, J=3.0, 1), 3.70 (m, 1), 3.54 (m, 4), 3.43 (m, 2), 2.90 (m, 1), 2.41 (m, 1), 2.19 (m, 1),
2.10 (s, 3) 1.45 (m, 6), 0.92 (m, 9); MS (ESPOS): 435 [M+H]+.
Example 48 Preparation of 4-Pentyloxy-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0526] The title compound was made using the synthetic sequence found in Method S starting from 4-hydroxypyridine-2-carboxylic acid 10b (R9 = hydroxy) substituting n-pentyl bromide as the alkylating agent.
[0527] Compound 15a (R9 = pentoxy): 1H NMR (300 MHz, CD3OD) δ 8.38 (d, J=5.1, 1), 7.64 (s, 1), 7.10 (d, J=3.3, 1), 4.18 (t, J=6.6, 2), 1.85 (m, 2), 1.49 (m, 4), 0.96 (t, J=7.2, 3). MS (ESNEG): 208 [M-H]". •
[0528] Compound 15b (R*=Me, R2=Me, R9= pentoxy): 1H NMR (300 MHz, CD3OD) δ 8.41 (d, J=5.7, 1), 7.61 (d, J=2.4, 1), 7.07 (dd, J=2.4, 5.4, 1), 5.27 (d, J=5.4, 1), 4.05-4.31 (m, 5), 3.85 (d, J=3.0, 1), 3.57 (dd, J=3.3, 7.2, 1), 2.11 (m, 4), 1.81 (m, 2), 1.49 (m, 4), 1.00 (m, 9). MS (ESPOS): 443 [M+Hf .
[0529] Title compound (20 mg, 10%): 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.22 (dd, J=9.9; 3.3, 1), 4.10 (m, 2), 3.76 (m, 3), 3.51 (m, 3), 3.39 (m, 1), 3.02 (m, 2), 2.43 (m, 1), 2.15 (m, 1), 2.10 (s, 3) 1.95 (m, 2), 1.69 (m, 2), 1.53 (m, 2), 1.34 (m, 2), 0.93 (m, 9); MS (ESPOS): 449 [M+Hf.
Example 49 Preparation of 4-(4-Fluoro-butoxy)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide
[0530] The title compound was made using the synthetic sequence found in Method S starting from 4-hydroxypyridine-2-carboxylic acid 10b (R9 = hydroxy) substituting 4- fluorobutyl bromide as the alkylating agent.
[0531] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.53 (t, J=5.7, 1), 4.37 (t, J=5.7, 1), 4.21 (dd, J=3.3, 6.6, 1), 4.07 (m, 2), 3.80 (d, J=3.3, 2), 3.60 (m, 5), 2.88 (m, 1), 2.38 (m, 1), 2.18 (m, 1), 2.10 (s, 3) 1.33-1.83 (m, 8), 0.92 (m, 6); MS (ESPOS): 453 [M+Hf..
Example 50 Preparation of 4-Butyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-allyl]-amide
[0532] To a solution of Boc 7-Methylene MTL (P=Boc, R*=Me, R2=CH2) prepared from compound 2a (P=Boc, R1=Me) by Method D (391mg, 1.1 mmol) in a solution of dichloroethane (10 mL) and dimethylsulfide (0.4 mL, 2.5 mmol) was added, TFA (5 mL) containing water (0.4 mL) and the reaction mixture stirred at rt for 45min. The solvent was removed and the residue evaporated twice from DCE to obtain the crude product. The product was obtained as an HCl salt by precipitation from ethyl acetate (4 mL) at 0 °C by addition of 2M HCl in ether, and dried under vacuum (351mg g, 86%). The white solid product was used in the next reaction without additional purification. [0533] MS (ESPOS): 350 [M+H]+.
[0534] Triethylamine (0.68 mL, 4.9 mmol, 4.0 equiv), followed by BSTFA (0.58 mL, 2.20 mmol, 1.8 equiv), were added to a stirred suspension of compound prepared above (351 mg, 1.22 mmol, 1 equiv) in anhydrous DMF (5 mL) at 0 °C and under nitrogen. The resulting mixture was stirred at 0 °C for 10 min, and then at rt for 50 min. The resulting solution was cooled to 0 °C and a solution of compound 7d (R9=n-butyl) prepared by Method K (400 mg, 1.47^ mmol, 1.2 equiv) in anhydrous DMF (5 mL) was added, followed by solid HATU (741 mg, 1.95 mmol, 1.6 equiv). The reaction mixture was allowed to warm to rt and after 2h the reaction solution was evaporated to dryness under vacuum. The residual oil obtained was diluted with EtOAc (200 mL), washed sequentially with 10%> citric acid , 1:1 saturated aqueous NaHC03, water, and brine dried over Na S04, and evaporated to dryness.
[0535] To a solution of 50mg crude coupling product in 1,2-dichloroethane (6 mL), was added dimethylsulfide (200 μL), followed by TFA (11.5 mL), and water (768 μL). The resulting mixture was stirred at rt for lh, evaporated to a minimal volume, diluted with DCE (3 x 10 mL), and evaporated to dryness. The residue was purified by column chromatography 8%) to 12%> 0.25M methanolic ammonia in dichloromethane to provide the title compound (10.0 mg, 25 %). [0536] 1H NMR (300 MHz, CD3OD) δ 5.22 (d, J = 5.8, 1), 5.00 (s, 1), 4.95 (s, 1), 4.58 (d, J = 8.8, 1) 4.19 (d, J = 8.8, 1), 4.09 (dd, J = 5.8, 10.1, 1) 3.85-3.77 (m, 2), 3.57-3.52 (m, 1), 3.26- 3.29 (m, 1), 2.59-2.53 (m, 1), 2.10-1.98 (m, 4), 1.80 (s, 3), 1.36-1.51-1.11 (m, 7), 0.91(t, , J=6.9, 3); MS (ESPOS): 403.6 [M+Hf.
Example 51 Preparation of l,4-Diethyl-piperidine-2-carboxylk acid [2-methyl-l-(3,495-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0537] To the product of Example 1 (30 mg, 0.07 mmol) in DMF (1 mL), DIEA (43 mg, 0.35 mmol) was added at room temperature and stirred overnight. Then removed the solvent and the resulting product was purified by column chromatography using 20%) MeOH in DCM to obtain the title compound (20 mg, 66%>) as a white powder.
[0538] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 6.0, 1), 4.26 (dd, J=3.6, 9.6, 1), 3.79 (d, J = 3.0, 1), 3.55 (dd, J=3.3, 10.2, 1) 2.85 (m, 2), 2.13 (m, 4), 1.37 (m, 12), 0.94 (m, 9); MS (ESPOS): 420 [M+Hf.
Example 52 Preparation of 4-(3-Fluoro-propoxy)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0539] The title compound was made using the synthetic sequence found in Method S starting from 4-hydroxypyridine-2-carboxylic acid substituting 3-fluoropropyl bromide as the alkylating agent.
[0540] Compound 15a (R9 = 3-fluoropropoxy): 1H NMR (300 MHz, CD3OD) δ 8.41 (d,
J=5.1, 1), 7.65 (d, J=2.1, 1), 7.14 (dd, J=2.1, 5.7, 1), 4.59 (m, 2), 4.24 (t, J=6.0, 2), 1.91 (m, 2).
MS (ESNEG): 212 [M-H]".
[0541] Compound 15b (R*=Me, R2=Me, R9 = 3-fluoropropoxy): 1H NMR (300 MHz,
CD3OD) δ 8.44 (d, J=5.7, 1), 7.65 (d, J=2.4, 1), 7.12 (dd, J=2.4, 5.7, 1), 5.48 (d, J=5.7, 1), 4.87
(m, 2), 4.30 (m, 2), 4.12 (dd, J=3.0, 10.2, 1), 3.85 (d, J=3.3, 1), 3.56 (dd, J=9.9, 3.3, 1), 2.26 (m,
1), 2.11 (s, 3), 1.37 (m, 4), 1.00 (t, J=5.1, 6). MS (ESPOS): 443 [M+Hf.
[0542] Title compound (60 mg, 31%): 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1),
4.50 (m, 2), 4.21 (dd, J=3.3, 9.9, 1), 4.06 (m, 2), 3.80 (d, J=2.7, 1), 3.66 (m, 3), 3.59 (m, 1), 3.33
(m, 1), 2.87 (m, 1), 2.41 (m, 1), 2.18 (m, 1), 2.10 (s, 3) 1.91 (m, 4), 1.51 (m, 2), 0.92 (m, 6); MS
(ESPOS): 439 [M+Hf.
Example 53
Preparation of 4-(3,3,3-Trifluoro-propoxy)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0543] The title compound was made using the synthetic sequence found in general Method S starting from 4-hydroxypyridine-2-carboxylic acid substituting 2-trifluoroethyl bromide as the alkylating agent. [0544] Compound 15a (R9 = 3,3,3-trifluoropropoxy): 1H NMR (300 MHz, CD3OD) δ 8.67
(m, 1), 7.92 (s, 1), 7.43 (m, 1), 4,65 (m, 2), 3.01 (m, 2). MS (ESNEG): 234[M-H]". [0545] Compound 15b (R2=Me, R2=Me, R9 = 3,3,3-trifluoropropoxy): 1H NMR (300 MHz, CD3OD) δ 8.46 (d, J=6.0, 1), 7.65 (d, J=2.7, 1), 7.13 (dd, J=2.7, 6.0, 1), 5.27 (d, J=5.7, 1), 4.39 (t, J=6.0, 2), 4.30 (m, 2), 4.11 (m, 1), 3.85 (d, J=3.0, 1), 3.57 (dd, J=3.0, 10.2, 1), 2.88 (m, 2), 2.25 (m, 1), 2.11 (s, 3), 1.00 (t, J=6.9, 6). MS (METHOD ESPOS): 469 [M+H]+. [0546] Title compound (10 mg, 10%): 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.18 (dd, J=3.0, 9.9, 1), 4.15 (m, 2), 3.80 (d, J=3.6, 1), 3.74 (m, 2), 3.52 (dd, J=3.3, 10.2, 2), 3.38 (m, 2), 3.18 (m, 1), 2.66 (m, 1), 2.66 (m, 1), 2.44 (m, 2), 2.22 (m, 1), 2.10 (s, 3) 1.34 (m, 2), 0.91 (d, J=7.2, 6); MS (ESPOS): 475 [M+Hf.
Example 54 Preparation of 4-Isobutyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0547] 4-Isobutyl-2-cyanopyridine is prepared as follows. To isobutylfriphenylphosphomum iodide (Aldrich) (8 g, 18.5 mmol) in THF (20 mL) at 0 °C, potassium-tert-butoxide (1.8 g, 16 mmol) was added, stirred at room temperature for 1 hr. pyridine-4-carboxlaldehyde (1 g, 9.3 mmol) was added, stirred at room temperature for overnight. Reaction mixture was then poured to water (100 mL) and extracted with EtOAc (100 mL). The product obtained on removal of solvent was purified by column chromatography using 40% EtOAc in hexane as eluent (1.05 g, 84%). To the resulting product (4.2 g, 31.5 mmol), 10%> Pd/C (0.4 g) was added and hydrogenated at 1 atm pressure overnight. Removal of solvent and purification on column chromatography using 30% EtOAc in hexanes resulted in 4-isobutylpyridine (3.8 g, 90%>). [0548] The intermediate, 4-isobutylpyridine-2-carboxylic acid, compound 10b, (R9 =isobutyl) was made by employing Method P. To 4-isobutylpyridine (2 g, 14.8 mmol) in acetic acid (20 mL), hydrogen peroxide (3.35 g, 30 %, 30 mmol) was added and refluxed overnight. After removing the solvent, the residue was dissolved in DCM dried over magnesium sulfate and taken as such for the next step. To the compound in DCM (10 mL) trimethylsilyl cyanide (1.73 g, 17.4 mmol) and dimethylcarbonyl chloride (1.89 g, 17.4 mmol) was added and stirred at room temperature for 24 hours. Aqueous potassium bicarbonate (100 mL, 10%>) was added and extracted twice with DCM (50 mL each). The crude product obtained on removal of solvent was taken in HCl (6N, 100 mL) and refluxed for 24 hours. Removal of acid and purification of crude product by column chromatography using 30% MeOH in DCM resulted in acid 10b (R9 =isobutyl) (1.5 g, 100%).
[0549] 1H NMR (300 MHz, CD3OD) δ 8.78 (d, J=5.7, 1), 8.44 (s, 1), 8.13 (d, J=5.7, 1), 2.92 (d, J=7.5, 1), 2.15 (m, 1), 0.98 (d, J=6.6, 6). MS (ESNEG): 178[M-H]\ [0550] To the amine 2b (R*=Me, R =Me) (200 mg, 0.79 mmol) in DMF (2ml), TEA (161 mg, 1.6 mmol), BSTFA (614 mg, 2.4 mmol) was added at 0 °C and stirred at room temperature for 1.5 hr. Acid 10b (R9 = isobutyl) (214 mg, 1.2 mmol) and HATU (368 mg, 1.2 mmol) was added and let stirred at room temperature for 4 hours. DMF was removed and the residue was extracted with EtOAc (50 mL), washed with sodium bicarbonate (10%, 50 mL), brine (50 mL) and dried over magnesium sulfate. The product obtained on removal of solvent was dissolved in methanol (10 mL) and treated with NR-50 resin (300 mg) for 3 hr. After filtering the resin, methanol was removed to obtain the crude product. It was then purified on silica gel column chromatography using 3% MeOH in DCM to obtain compound lib (R1=Me, R2=Me, R3=H, R9 =isobutyl) (200 mg, 60%).
[0551] 1H NMR (300 MHz, CD3OD) δ 8.41 (d, J=4.8, 1), 8.28 (d, J=9.6, 1), 7.95 (s, 1), 5.35 (d, J=5.4, 1), 4.25 (m, 2), 3.99 (d, J=10.8, 1), 3.78 (d, J=3.6, 1), 3.55 (dd, J=3.6, 10.8, 1), 2.52 (m, 3), 2.15 (s, 3), 1.93 (m, 1), 1.02 (m, 12). MS (ESPOS): 413 [M+Hf. [0552] To compound lib (R1=Me, R2=Me, R3=H, R9 =isobutyl) (200 mg, 0.48 mmol) in water (10 mL), AcOH (2 mL) and MeOH (2 mL), Pt02 (200 mg), was added, hydrogenated at 55 psi for 16 hours. After filtering the catalyst, the solvent was removed to obtain the crude material which on purification over silica gel column using 20 %> MeOH in DCM as eluent. The lower Rf fraction provided the title compound (70 mg, 34%). [0553] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.20 (dd, J=9.9; 3.3, 1), 4.07 (m, 2), 3.80 (d, J=3.0, 1), 3.60 (m, 2), 3.34 (m, 2), 2.84 (m, 1), 2.17 (m, 1), 2.10 (s, 3) 2.01 (m, 1), 1.77 (m, 3), 1.40 (m, 4), 0.91 (m, 12); MS (ESPOS): 419 [M+Hf.
Example 55 Preparation of 4-Propyl-piperidine-2-carboxylic acid [2,2-difluoro-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0554] To acid 10b (R9 = propyl) prepared by Method P (53 mg, 0.32 mmol) in DMF (3 mL), lincosamide intermediate, compound 5b (R1 = Me), prepared by Method I (63 mg, 0.16 mmol) and HATU (121 mg, 0.32 mmol), triethylamine (70 mg, 0.48 mmol) was added at 0°C and stirred at room temperature 16 hours. DMF was removed and the residue was taken in ethyl acetate and washed with saturated bicarbonate (30 mL). The product obtained on removal of solvent was purified on silica gel column using 30% ethyl acetate in hexanes (40 mg, 45 %>). To the pure product in methanol (5 mL), water (5 mL), acetic acid (5 mL) and platinum dioxide (50 mg, mmol) was added and hydrogenated at 60 psi for 16 hours. After filtering tlie catalyst, tl e solvent was removed to obtain the crude product which was taken in methanol (3 mL). Potassium carbonate (125 mg, 0.83 mmol) in water (1 mL) was added to it and stirred 16 hours. Solvents were then removed and the crude product purified on column chromatography using 20 %> methanol in dichloromethane. The lower Rf fraction resulted in the title compound (10 mg, 33%). [0555] 1H NMR (300 MHz, CD3OD) δ 5.28 (d, J = 5.4, 1), 4.73 (s, 1), 4.57-4.65 (m, 1), 4.33-4.42 (m, 1), 4.05 (m, 1), 3.89 (s, 1), 3.53-3.57 (m, 2), 2.83 (t, J = 12.3, 1), 2.09 (s, 3), 1.63- 1.84 (m, 5), 1.16-1.37 (m, 6), 0.93 (m, 3). MS(ESPOS): 427 [M+Hf.
Example 56 Preparation of 4-Fluoro-4-propyl-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0556] To a stirred solution of (2S, 4R)-4-hydroxyproline (Aldrich) (25 g, 108 mmol) in methanol (50 mL) at 0 °C was added frimethylsilyldiazomethene (24.6 g, 216 mmol). The mixture was stirred at 0 °C for 1 hour. The residue obtained on removal of solvent and purification by column chromatography using 50% ethyl acetate in hexanes (27 g, 100%) was used in the next step. To oxalyl chloride (15 g, 118 mmol) in DCM (15 mL) at -78 °C, DMSO (18.6 mL, 236 mmol) was added slowly over 15 minutes. After the completion of addition, the above product (2S, 4R)-N-Boc-4-hydroxyproline methylester (26.5 g, 108 mmol) in DCM (100 mL) was added at -78 °C for 20 minutes. Triethylamine (54.6 g, 540 mmol) was added followed by stirring at room temperature for 2 hours. The reaction mixture was then washed with 10% aq HCl (200 mL) and the organic layer was separated and dried over sodium sulfate. The crude product obtained on removal of solvent was purified on silica gel column chromatography using 50%) EtoAc in hexanes to obtain (2S, 4R)-N-Boc-4-Ketoproline methylester (20 g, 78%). [0557] 1H NMR (300 MHz, CDC13) δ 4.80 (m, 1), 3.88 (d, J=8.7, 2), 3.77 (s, 3), 2.98 (m, 1), 2.58 (m, 1), 1.45 (s, 9); MS (ESPOS): 244 [M+Hf. [0558] To a stirred solution of (2S, 4R)-N-Boc-4-Ketoproline methylester (1 g, 4.11 mmol) in THF (10 mL), tetraallyltin (1.08 mL, 4.52 mmol) in dry THF was added, then cooled to 0 °C before borontrifluoride etherate (0.520 mL, 4.11 mmol) was added drop wise. The mixture was stirred at 0 °C for lh and then at room temperature for an additional 2 hours. Potassium fluoride (360 mg in 5mL water) and celite (1 g) was added and the reaction mixture was stirred for an hour. The reaction mixture was filtered and concentrated to dryness and the residue was dissolved in DCM (200 mL), washed with water (lOOmL) and brine 100 mL), dried over MgS0 and evaporated to dryness. T he residue obtained on removal of solvent was purified by silica gel column chromatography using 50% EtOAc in hexanes to obtain 4-Hydroxy-4-allylproline methylester (0.94 g, 80%).
[0559] 1H NMR (300 MHz, CDC13) δ 5.87 (m, 1), 5.19 (m, 2), 4.34 (m, 1), 3.75 (d, J=4.8, 3), 3.50 (m, 3), 2.37 (m, 1), 2.21 (m, 1), 1.39 (d, J=12.9, 9); MS (ESPOS): 308 [M+Naf. [0560] To a stirred solution of DAST (1.06 g, 6.58 mmol) in DCM (10 mL) at -78 °C, 4-hydroxy-4-allylproline methylester (940 mg, 3.3 mmol) in dry DCM (10 mL) was added slowly. The mixture was then stirred at -78 °C for lh, then at -10 °C for an additional lh. DCM (50 mL) was added, quenched withN-H-tCl (10%>, 150 mL) and the organic layer was separated, dried over sodium sulfate and evaporated to dryness. The residue obtained on removal of solvent was purified by silica gel column chromatography using 5%» EtOAc in hexanes as eluent to obtain 4-fluoro~4-allylproline methylester (330 mg, 34%).
[0561] 1H NMR (300 MHz, CDC113) δ 5.82 (m, 1), 5.12 (m, 2), 4.43 (m, 1), 3.66 (s, 3), 3.47 (m, 1), 2.37 (m, 1), 2.43 (m, 4), 1.37 (dd, J=4.5, 13.8, 9); MS (ESPOS): 310 [M+Naf. [0562] To a solution of 4-fluoro-4-allylproline methylester (0.33 g, 1.15 mmol) in MeOH (15 mL) was added 10% Pd/C (40 mg) and hydrogenated at 1 atmosphere. The catalyst was filtered through celite and washed with methanol. To the product obtained on removal of solvent (330 mg, 1.15 mmol) in THF (12 mL) was added aq lithium hydroxide monohydrate (60 mg, 1.38 mmol). The reaction mixture was stirred at room temperature overnight. THF was removed and the residue was taken up in ethyl acetate (50 mL), washed with 10% citric acid (100 mL) and brine (20 mL). Removal of solvent resulted in 4-fluoro-4-propylproline (310 mg, 100%>). [0563] 1H NMR (300 MHz, CD3OD) δ 4.43 (m, 1), 3.71 (m, 6), 2.51 (m, 2), 1.98 (m, 3), 1.45 (m, 9), 0.96 (m, 3); MS (ESNEG): 274 [M-H]".
[0564] To a solution of 4-fluoro-4-propylproline (310 mg, 1.15 mmol) in DMF (3 mL) at 0 °C, 7-Methyl MTL 2b (R1 =Me, R2=Me) (272 mg, 1.15 mmol), HBTU (469 mg, 1.3 mmol) and DIEA (290 mg, 2.3 mmol) was added, left stirred at room temperature for 16 hours. DMF was removed and the residue obtained was purified by 3%> MeOH in DCM(40 mg, 93%). The product from the column purification was taken in DCE (6 mL), to which triethylsilane (0.16 mL), TFA (2 mL) and water (0.16 mL) was added and stirred at room temperature for 1.5 hours. Removal of solvent followed by purification on silica gel column chromatography using 10% MeOH in DCM resulted in the title compound as isomeric mixtures with lower RF fraction (160 mg, 50%).
[0565] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.46 (m, 1), 4.24 (dd, J=5.7, 10.2, 1), 4.08 (m, 2), 3.81 (d, J=2.4, 1), 3.52 (m, 3), 2.73 (m, 1), 2.10 (m, 4), 1.88 (m, 2), 1.50 (m, 2), 0.99 (t, J=7.5, 3), 0.91 (dd, J=3.0, 6.9, 6); MS (ESPOS): 409 [M+Hf; and higher Rf fraction (40 mg, 12%). 1H NMR (300 MHz, CD3OD) δ 5.38 (d, J=5.4, 1), 4.46 (m, 1), 4.24 (dd, J=2.7, 7.2, 1), 4.08 (m, 2), 3.81 (d, J=2.4, 1), 3.64 (m, 3), 2.73 (m, 1), 2.11 (m, 4), 1.84 (m, 2), 1.47 (m, 2), 0.98 (t, J=7.5, 3), 0.91 (dd, J=3.0, 6.9, 6); MS (METHOD ESPOS): 409 [M+Hf.
Example 57 Preparation of 4-Butyl-4-fluoro-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0566] To ethyl acetylene (140 mg, 2.6 mmol) in THF (5 mL) at -78 °C, n-butyllithium (1.1 mL, 2.6 mmol) was added with stirring at -78 °C for 1 hour. Then n-(tert-Butoxycarbonyl)-L- proline-4-ketone (described in the example 56) (570 mg, 2.3 mmol) in THF (5 mL) was added at -78 °C with stirring for 2 hours and then let it warm to -40 °C over 1 hour. The reaction mixture was extracted with EtOAc (20 mL), washed with saturated NH CI (5 mL) and dried over sodium sulfate. Purification of the crude product was carried out by silica gel chromatography using 50% EtOAc in hexane to obtain the N-boc-4-butyl-4-hydroxy-prolinemethyl ester (520 mg, 73%). To the DAST (556 mg, 3.4 mmol) in DCM (5 mL) at -78 °C, was added a solution of the above ester (520 mg, 1.7 mmol) in DCM (5 mL) at -78 °C and stirred for 1 hour. The reaction mixture was extracted with DCM (50 mL) and washed with NaHC03 (30 mL, 10%). The product obtained after removal of solvent was purified by silica gel chromatography using 5% EtOAc in hexanes to obtain N-(tert-butoxycarbonyl)-L-proline-4-fluoro-4-butane (276 mg, 52%).
[0567] 1H NMR (300 MHz, CD3OD) δ 4.41 (m, 1), 3.83 (m, 1), 3.71 (s, 3), 3.45 (m, 1), 2.55-1.54 (m, 8), 1.39 ( , 9), 0.89 (m, 3); MS (ESPOS): 326 [M+Naf. [0568] To a solution of N-(tert-butoxycarbonyl)-L-proline-4-fluoro-4-butane (270 mg, 0.89 mmol) in THF (12 mL) and water (4 mL), was added lithium hydroxide monohydrate (45 mg, 1.07 mmol). The reaction mixture was stirred at room temperature for 16 hours. THF was removed under vacuum and the residue was taken up in ethyl acetate (150 mL), washed with 10% citric acid (100 mL) and brine (20mL). Removal of solvent resulted in N-(tert-butoxy)-L- proline-4-fluoro-4-butyl-2-carboxylic acid (260 mg, 100%).
[0569] 1H NMR (300 MHz, CD3OD) δ 4.32 (m, 1), 3.72 (m, 2), 2.58 (m, 2), 2.10-1.63 (m, 6), 1.42 (m, 9), 0.93 (t, J=6.6, 3); MS (ESNEG): 288 [M-H]". '
[0570] To a solution of N-(tert-butoxy)-L-proline-4-fluoro-4-butyl-2-carboxylic acid (135 ' mg, 0.46 mmol) in DMF (3 mL) at 0 °C, 7-Methy MTL 2b (R1 =Me, R2=Me) (135 mg, 0.46 * mmol), HBTU (194 mg, 0.51 mmol), DIEA (120 mg, 0.93 mmol) was added, left it at room temperature for 16 hours. The product obtained after removing DMF and purification by column chromatography using 5% MeOH in DCM (189 mg, 77%>) was taken in DCE (6 mL). Triethylsilane (0.16 mL), TFA (2 mL) and water (0.16 mL) was added, stirred at room temperature for 1.5 hours. The residue obtained on removal of solvent was purified by column chromatography using 10% MeOH in DCM to obtain the title compound (156 mg, 96%). [0571] 1H NMR (300 MHz, CD3OD) δ 5.26 (d, J=5.7, 1), 4.55 (m, 1), 4.27 (dd, J=3.3, 10.2, 1), 4.08 (m, 2), 3.82 (d, J=3.0, 1), 3.58 (m, 3), 2.79 (m, 1), 2.22 (m, 1), 2.10 (s, 3), 1.89 (m, 3), 1.40 (m, 4), 0.91 (m, 9); MS (ESPOS): 423 [M+Hf.
Example 58 Preparation of 4-Fluoro-4-propyl-pyrrolidine-2-carboxylic acid [2-hydroxy-l-(3,4,5- trihydroxy-6-methylsuIfanyI-tetrahydro-pyran-2-yl)-propyI]-amide
[0572] N-(tert-butoxycarbonyl)-4-fluoro-4-propyl-L-proline was prepared as described in the previous example (except using n-propyl lithium in the place of n-butyl lithium) (164 mg, 0.57 mmol) was suspended in dry acetonitrile (4 mL). Triethylamine (332 μL, 3.02 mmol) was added and the reaction mixture was cooled to 0°C. Isobutyl chloroformate (78 μL, 0.57 mmol) was added and after 10 min the reaction was allowed to warm to 4°C. After 1.5 h a solution of MTL (151 mg, 0.57 mmol) in 1 :1 acetone: water (4 mL) was added and the reaction mixture was stirred for lOh at rt. The reaction mixture was evaporated to dryness and chromatographed on silica 95:5 dichloromethane/MeOH to 95:8 dichloromethane/MeOH to provide the product as a colorless oil (137 mg, 45%): TLC Rf 0.32 (9:1 dichloromethane/MeOH). [0573] To a solution of the above boc protected lincosamide (125 mg,) in DCM (2.0 mL) was added a solution of DCE (10.0 mL), trifluoroacetic acid (5 mL) methyl sulfide (0.3 mL), and water (0.3 mL). The reaction mixture was stirred at rt for 40 min then diluted with DCE (25.0 mL). The solvent was removed under vacuum and co-evaporated with DCE twice. The residue was purified by chromatography on fluorosil 20% MeOH (0.25M NH3) in DCM to provide the product as a colorless solid (30.0 mg, 30%>).
Example 59
Preparation of 4-(2-methoxyethoxy)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methyIsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0574] The title compound was made using the synthetic sequence found in Method S starting from 4-hydroxypyridine-2-carboxylic acid, substituting 2-methoxyethyl bromide as the alkylating agent.
[0575] Compound 15a (R9 = 2-methoxyethoxy): 1H NMR (300 MHz, CD3OD) δ 8.40 (d,
J=6.0, 1), 7.69 (d, J=2.4, 1), 7.20 (dd, J=2.7, 6.3, 1), 4.35 (m, 2), 3.80 (m, 2), 3.40 (s, 3). MS
(ESNEG): 196[M-H]\
[0576] Compound 15b (RJ=Me, R2=Me, R9= 2-methoxyethoxy): 1H NMR (300 MHz,
CD3OD) δ 8.43 (d, J=5.7, 1), 7.65 (d, J=2.4, 1), 7.12 (dd, J=2.4, 5.7, 1), 5.27 (d, J=5.4, 1), 4.10-
4.87 (m, 4), 3.85 (d, J=3.3, 1), 3.77 (m, 2), 3.55 (m, 1), 3.41 (s, 3), 2.26 (m, 1), 2.11 (s, 1), 0.998
(m, 6). MS (ESPOS): 431 [M+Hf.
[0577] Title compound (10 mg, 10%). 1H NMR (300 MHz, D2O) δ 5.18 (d, J=6.0, 1), 4.00
(m, 3), 3.70 (m, 1), 3.56 (m, 1), 3.45 (m, 3), 3.26 (m, 1), 3.16 (m, 3), 3.10 (m, 1), 2.80 (m, 1),
2.48 (m, 1), 2.22 (m, 1), 1.96 (m, 4), 1.17-1.72 (m, 4), 0.70 (m, 6); MS (ESPOS): 437 [M+Hf.
Examples 60-62
Example 60
Preparation of 4-ButyI-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Example 61
Preparation of 4-(4,4-Difluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Example 62 ion of 4-(3-Fluorθ' piperidine-2-earboxylfc acid [2-methyl-l-(3,4,5- trihydroxy-6-methyl§ulfanyl •tetrahydro-pyran-2-yl)-propyl]-amide
[0578] Examples 60 to 62 were prepared using methods and techniques describe herein utilizing commercially available starting materials where appropriate.
Example 63
Preparation of 4-Fluoro-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0579] Lincosamine 2b (R1=Me, R2=Me) was coupled to carbamate 21d (P2=Boc, m = 2, R9 = «-propyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R =R =Me, R3=H, R9=propyl/fluoro, P1=H, P2=carboxylic acid-t-butyl ester, m=2) which was deprotected under acidic conditions to provide the title compound.
[0580] HPLC: C18 3.5 μm, 4.6 x 30 mm Column; gradient eluent 2%-98% MeCN over 10 min; 1.5 mL/min): Rt = 3.696 min); 1H NMR (300 MHz, CD3OD) δ 5.36 (d, J= '5.8, 1), 4.24 (dd, J= 3.3, 12.9, 1), 4.20 (s, 2), 4.12 (dd, J= 6.0, 10.4, 1), 3.88 (d, /= 3.3, 1), 3.65 (dd, J= 3.0, 10.4, 1), 3.48 (dd, J= 4.1, 13.1, 1), 3.33 (ddd, J= 3.6, 3.6, 13.5, 1), 2.43-2.45 (m, 1), 2.22-1.58 (m, 6), 2.16 (s, 3), 1.48 (m, 2), 0.94 (t, J= 7.1, 3), 0.88 (d, J= 6.9, 6); 19F NMR (CD3OD) δ- 158.8-δ-159.0 (sextuplet), MS (ESPOS): 423.2.
Example 4
Preparation of 4-(2-Fluoroethosy)-piperidine-2-carbosylic acid [2-methyl-l-(3,4,5- trihydro ]-amide
[0581] The title compound was made using the synthetic sequence found in general method S starting from 4-hydroxypyridine-2-carboxylic acid 10b (R9 = hydroxy) substituting 2- fluoroethyl bromide as the alkylating agent.
[0582] 15a (R9'=2-fluoroethoxy): 1H NMR (300 MHz, CD3OD) δ 8.40 (d, J=6.0, 1), 7.73 (d, J=2.7, 1), 7.24 (dd, J=2.4, 6.0, 1), 4.87 (m, 2), 4.50 (m, 2). MS (ESNEG): 184 [M-H]". [0583] 15b (R*=R2 =Me, R3=H, R9 =2-fluoroethoxy): 1H NMR (300 MHz, CD3OD) δ 8.45 (d, J=5.7, 1), 7.67 (d, J=2.4, 1), 7.14 (dd, J=2.7, 5.7, 1), 5.27 (d, J=5.7, 1), 4.68 (m, 1), 4.43 (m, 1), 4.12-4.33 (m, 3), 4.10 (dd, J=6.0, 10.2, 1), 3.18-3.84 (m, 3), 2.26 (m, 1), 2.11 (s, 3), 1.00 (t, J=5.1, 6). MS (ESPOS): 419 [M + H]+ .
[0584] Title compound (20 mg, 14%): 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.59 (t, J=4.2, 1), 4.43 (t, J=4.2, 1), 4.18 (dd, J=3.3, 6.6, 1), 4.07 (m, 2), 3.80 (m, 2), 3.73 (m, 1), 3.55 (m, 2), 3.48 (m, 1), 3.21 (m, 1), 2.74 (m, 1), 2.32 (m, 1), 2.18 (m, 1), 2.10 (s, 3) 1.38 (m, 3), 0.93 (m, 6); MS (ESPOS): 425 [M + H]+.
Example 65
[0585] There is no Example 65.
Example 66
Preparation of 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0586] Compound 14c (R =2-cyclopropylethyl) was prepared using the methods described in general method R.
[0587] To a stirred suspension of 14a (0.5 g, 1.9 mmol, 1 equiv), triphenylphosphine (39.9 mg, 0.15 mmol, 0.08 equiv), copper (I) iodide (28.9 mg, 0.15 mmol, 0.08 equiv), palladium acetate (17 mg, 0.076 mmol, 0.04 equiv) in triethylamine (7 mL) under dry nitrogen, was added cyclopropyl acetylene (Aldrich) (0.25 g, 3.8 mmol, 2 equiv). The mixture was stirred at rt overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by column chromatography to give 14b (R9'=2-cyclopropyl-eth-l-ynyl) (0.39 g, 100%) as a yellow oil. [0588] 1H NMR (300 MHz, CDC13) δ 8.65-8.56 (m, 1), 8.06-7.99 (m, 1), 7.40-7.32 (m, 1),
3.98 (s, 3), 1.50-1.40 (m, 1), 0.96-0.81 (m, 4). MS (ESPOS): 202.0 [M+Hf.
[0589] To a solution of 14b (R9 =2-cycloproρyl-eth-l-ynyl) (0.39 g, 1.9 mmol) in methanol (15 mL) was added 10% palladium on carbon (0.2 g). The mixture was purged and charged with hydrogen (1 atm) and stirred at rt overnight. The palladium was removed by filtration and the filtrate was concentrated to give 14c (R9=2-cyclopropylethyl) (0.38 g, 97%) as a yellow oil. [0590] 1H NMR (300 MHz, CDC13) δ 8.60 (d, J=4.5, 1), 8.00-7.96 (m, 1), 7.34-7.29 (m, 1),
3.99 (s, 3), 2.78 (t, J=7.6, 2), 1.58-1.49 (m, 2), 0.71-0.59 (m, 1), 0.47-0.38 (m, 2), 0.06-0.02 (m, 2); MS (ESPOS): 228.2 [M + Naf.
[0591] To a mixture of 14c (R9=2-cycloρropylethyl) (0.38 g) in MeOH (8 mL) and water (8 mL) were added cone. HCl (158 μL) and platinum oxide (0.2 g). The mixture was purged and charged with hydrogen (1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate was evaporated to give a light yellow solid.
[0592] To the above residue was added 2N NaOH (3.8 mL) and t-butylalcohol (2 mL). The reaction mixture was stirred at rt for 2 h, di-t-butyl dicarbonate (0.62 g, 2.85 mmol) was then, added and the mixture stirred overnight. The solvent was removed under vacuum and the resulting residue was diluted with water, then washed with ether. The aqueous layer was acidified with 2N HCl to pH=2.0, and extracted with ethyl acetate (2 x). The combined organic layers were dried and concentrated to give 4-(2-cyclopropylethyl)-piperidine-l,2-dicarboxylic acid 1-tert-butyl ester (0.42 g, 77 %) as a clear syrup. [0593] MS (ESPOS): 320.3 [M + Naf. MS (ESNEG): 296.2 [M-H]". [0594] To a solution of 4-(2-cyclopropylethyl)-piperidine- 1 ,2-dicarboxylic acid- 1 -tert-butyl ester (115 mg, 0.387 mmol, 1 equiv) and 2b (R =Me, R2=Me) HCl salt (111.4 mg, 0.387 mmol, 1 equiv) in DMF (2.4 mL) at r.t. was added DIEA (100 μL, 0.774 mmol, 2 equiv), followed by HBTU (162 mg, 0.426 mmol, 1.1 equiv). The reaction mixture was stirred at rt for 3 hrs then evaporated under high vacuum to dryness. The residue was diluted with ethyl acetate, washed with 1:1 10% citric acid/brine (1 x), sat. aqueous sodium bicarbonate (1 x), brine (1 x), dried and concentrated. The residue was purified by chromatography to give the desired l'-Boc protected lincosamide product Ila (R1=R2 =Me, R3=H, R9=2-cyclopropylethyl, P1=H, P2=carboxylic acid- t-butyl ester) (126 mg, 61%) as a clear syrup. [0595] MS (ESPOS): 531.3 [M+Hf.
[0596] To a solution of the above Ila (R!=R2 =Me, R3=H, R9=2-cyclopropylethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=2) in DCE (12 mL) at 0 °C was added a solution of trifluoroacetic acid (3 mL) and water (0.375 mL). The reaction mixture was stirred at 0 °C for 5 min and at rt for 35 min. The reaction solvent was removed under vacuum and co-evaporated with toluene twice, and the resulting residue was purified by semi-preparative HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60%> acetonitrile in H 0 w/ 0.1% AcOH over 30 min, 20 mL/min flow rate) to provide the title compound of example 66 (25 mg, 27 %) as a white solid.
[0597] 1HNMR (300 MHz, CD3OD) δ 5.24 (d, J=6, 1), 4.19 (dd, .7=3.4, 10, 1), 4.10-4.04 (m, 2), 3.81-3.72 (m, 2), 3.51 (dd, .7=3.3, 10.2, 1), 3.42-3.33 (m, 1), 3.02-2.90 (m, 1), 2.20-2.12 (m, 1), 2.11 (s, 3), 1.94-1.86 (m, 1), 1.76-1.64 (m, 1), 1.48-1.20 (m, 7), 0.94-0.85 (m, 6), 0.73- 0.62 (m, 1), 0.47-0.38 (m, 2), 0.05-0.02 (m, 2). MS (ESPOS): 431.3 [M+Hf.
Example 67
Preparation of 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0598] 4-Cyclopropylmethylpyridine-2-carboxylic acid, compound 10b (R9=cyclopropylmethyl), was made by employing Method P using the starting material 4- Cyclopropylmethylpyridine prepared as described by Osuch et al, Journal of the American Chemical Society, 1955, 78, 1723.
[0599] To a -78 °C solution of 4-picoline (1.1 g, 11.8 mmol) in THF (5 mL) was added a solution of LDA 2M in THF/heptane/ethylbenzene (Aldrich) (5.9 mL, 11.8 mmol). The resulting reaction mixture was stirred at -78 °C for 3 h then -40 °C for 1 h. Cyclopropyl bromide (1.43 g, 11.8 mmol) was added at -78 °C, the reaction mixture was alloed to warm up to room temperature and was stirred at room temperature for 1 h. To the reaction mixture was added saturated aqueous NKUC1 (10 mL), the aqueous phase was extracted with EtOAc (10 x 2 mL), and the combined organic extracts dried over Na2S04. 4-Cyclopropylmethylpyridine-2- carboxylic acid (0.5 g, 31%>) was obtained after the solvent was removed and used without further purification.
[0600] To a 0°C solution of 4-Cyclopropylmethylpyridine-2-carboxylic acid (10b) (R9=cyclopropylmethyl) (147 mg, 0.83 mmol), 2b (R!=Me, R2=Me) HCl salt (238 mg, 0.83 mmol) and TEA (231 μL, 1.66 mmol) in DMF (2 mL) was added solid HBTU (346 mg, 0.91 mmol), and the resulting reaction mixture was stirred overnight. Reaction solvents were removed, and the residue purified by silica gel column chromatography at 50-100%
EtOAc/Hexane to provide the desired lincosamide product lib (R =R =Me, R =H, R9=cycloproρylmethyl, P1=H) (260 mg, 76%).
[0601] To a solution of lib (R!=R2 =Me, R3=H, R9=cyclopropylmethyl, P1=H) (250 mg, 0.6 mmol) in water (10 mL), AcOH (2 mL), MeOH (3 mL) was added Pt02 (200 mg), and the reaction mixture was hydrogenated at 50 psi for 5 hr. The solvent was removed to obtain the crude lincosamide product. Purification was carried by silica gel column chromatography (20% MeOH/DCM), then HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 . mm ID x 100 mm, 5-60% acetonitrile in H20 w/ 0.1% AcOH over 30 min, 20 mL/min flow rate) to provide the title compound (13 mg, 5%) as a colorless solid.
[0602] 1H NMR (300 MHz, CD3OD) δ 5.23 (d, J=6, 1), 4.26-4.10 (m, 3), 3.83-3.78 (m, 2), 3.55-3.33 (m, 2), 3.07-2.95 (m, 1), 2.34-2.17 (m, 2), 2.12 (s, 3), 2.00-1.80 (m, 4), 1.42-1.15 (m, 4), 1.00-0.90 (m, 6), 0.79-0.66 (m, 1), 0.53-0.45 (m, 2), 0.12-0.04 (m, 2); MS (ESPOS): 417.3
Example 68
Preparation of 4-(2-Cyclobutyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyI-tetrahydro-pyran-2-yl)-propyl]-amide
[0603] 4-(2-Cyclobutyl-ethyl)-pyridine-2-carboxylic acid, compound 10b (R9=2-cyclobutyl- ethyl), was made by employing Method P using the starting material 4-(2-Cyclobutyl-ethyl)- pyridine prepared as described by Osuch et al, Journal of the American Chemical Society, 1955, 78, 1723.
[0604] Lincosamine 2b (R1=Me, R2=Me) was coupled to 4-(2-cyclobutyl-ethyl)-pyridine-2- carboxylic acid 10b (R9=cyclobutyl-ethyl) as depicted in general coupling scheme 11 to provide intermediate lib (R1=R2 =Me, R3=H, R9= 2-cyclobutyl-ethyl, P1=H) which was reduced by catalytic hydrogenation to the title compound. [0605] MS (ESPOS): 445.2 [M+Hf.
Example 69 ion of 4-Cyclobutylm' .-piperidine-2-earboxyl acid [2-methyl-l-(3,495- trihydrosy-6-methylsulfanyi etrahydro-pyran-2-yl)-propyl]-amide
[0606] 4-Cyclobutylmethylpyridine-2-carboxylic acid, compound (10b) (R9=4- cyclobutylmethyl) was made by employing Method P using the starting material 4- cyclobutylmethylpyridine prepared as described by Osuch et al, Journal of the American Chemical Society, 1955, 78, 1723.
[0607] Lincosamine 2b (R1=Me, R2=Me) was coupled to 4-cyclobutylmethylpyridine-2- carboxylic acid, compound 10b (R9=4-cyclobutylmethyl), as depicted in general coupling scheme 11 to provide intermediate lib (R1=R2 =Me, R3=H, R9= cyclobutyl-methyl, P1=H), which was reduced by catalytic hydrogenation to the title compound. [0608] MS (ESPOS): 431.3 [M+Hf.
Example 70
Preparation of 3-Butyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0609] To a solution of azetedine acid 16f (R -butyl) (52 mg, 0.20 mmol, 1 equiv), aminosugar (58 mg, 0.20 mmol, 1 equiv) and HBTU (84 mg, 0.22 mmol, 1.1 equiv) in DMF (2.0 mL) at 23 °C was added DIPEA (88 μL, 0.51 mmol, 2.5 equiv). After stirring for 12 h at 23 °C, DMF was removed in vacuo, then the residue was partitioned between EtOAc (100 mL) and 1:1 brine: 10% aqueous citric acid (100 mL). The organic layer was separated and washed with 1:1 brine/saturated aqueous NaHC03 (100 mL), brine (50 mL), dried (MgSQ ), filtered and concentrated to furnish 82 mg (0.17 mmol, 84%) Ila (R1=R2 =Me, R3=H, R9=butyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) as a glassy solid. [0610] MS (ESPOS): [M+Hf.
[0611] To a solution of carbamate Ila (R =R2 =Me, R3=H, R9=butyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) (82 mg, 0.17 mmol, 1 equiv) in 1,2-dichloroethane (10 mL) at 23 °C was added H20 (0.40 mL) followed by TFA (4.0 mL). After stirring for 20 min at 23 °C, toluene (50 mL) was added and the resulting solution was concentrated to dryness. The residue was purified by semi-preparative HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60% acetonitrile in H20 w/ 0.1% HCl over 30 min, 20 mL/min flow rate) to give 41 mg of title compound as a white solid.
[0612] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.60 (d, J=7.2, 1), 4.25 (dd, J=3.3, 9.9, 1), 4.16-4.05 (m, 2), 3.99 (t, J=9.0, 1), 3.81 (d, J=3.0, 1), 3.74 (dd, J=8.4, 9.9, 1), 3.50 (dd, j=3.3, 10.2, 1), 2.90-2.74 (m, 1), 2.23-2.10 (m, 1), 2.10 (s, 3), 1.90-1.67 (m, 2), 1.44-1.24 (m, 4), 1.00-0.86 (m, 9 H). MS (ESPOS): 391.4 [M+Hf.
Example 71
Preparation of 3-CyclopropylmethyI-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0613] Lincosamine 2b (R =Me, R =Me) was coupled to azetedine acid 16f (R9=cyclopropylmethyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R!=R2 =Me, R3=H, R9=cyclopropylmethyl, P*=H, P2=carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound.
[0614] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.65 (d, J=7.5, 1), 4.25 (dd, J=3.9, 9.6, 1), 4.13-3.99 (m, 3), 3.86-3.79 (m, 2), 3.51 (dd, J=3.3, 10.2, 1), 3.00-2.87 (m, 1), 2.21- 2.09 (m, 1), 2.11 (s, 3), 1.83-1.72 (m, 1), 1.62-1.51 (m, 1), 0.94 (d, J=6.9, 3), 0.89 (d, .7=6.9, 3), 0.82-0.70 (m, 1), 0.58-0.47 (m, 2), 0.18-0.08 (m, 2 H). MS (ESPOS): 389.2 [M+Hf.
Example 72
Preparation of 3-Propyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0615] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=propyl) as depicted in general coupling scheme 11 to provide intermediate Ila R1=R2 =Me, R3=H, R =propyl, P =H, P =carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound.
[0616] 1H NMR (300 MHz, CD3OD) δ 5.26 (d, J=5.4, 1), 4.60 (d, J=7.5, 1), 4.26 (dd, J=3.3, 9.6, 1), 4.13-3.99 (m, 3), 3.81 (d, J=3.3, 1), 3.76 (dd, .7=5.1, 10.2, 1), 3.51 (dd, .7=3.3, 9.9, 1), 2.93-2.76 (m, 1), 2.24-2.09 (m, 1), 2.11 (s, 3), 1.84-1.67 (m, 2), 1.48-1.30 (m, 2), 1.01-0.87 (m, 9 H). MS (ESPOS): 377.0 [M+Hf.
Example 73
Preparation of 3-Butyl-l-(2-hydroxy-ethyl)-azetidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0617] A sample of 3-Butyl-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide prepared in example 70 was alkylated with ethyleneoxide as depicted in scheme 12 (R6=2-hydroxyethyl) to provide the title compound. [0618] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.4, 1), 4.13-4.05 (m, 3), 3.83-3.77 (m, 2), 3.64 (t, J=9.9, 1), 3.58-3.48 (m, 2), 3.37-3.30 (m, 1), 2.78-2.55 (m, 2), 2.44-2.34 (m, 1), 2.25-2.10 (m, 2), 2.11 (s, 3), 1.80-1.40 (m, 2), 1.39-1.20 (m, 4), 0.95-0.84 (m, 9 H). MS (ESPOS): 435.1 [M+Hf.
Example 74
Preparation of 3-PentyI-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0619] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=pentyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R1=R2 =Me, R3=H, R =pentyl, P =H, P =carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound.
[0620] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=5.7, 1), 4.55 (d, J=6.9, 1), 4.25 (dd, J=3.3, 9.9, 1), 4.13-4.04 (m, 2), 4.00 (t, J=9.3, 1), 3.80 (d, J=3.0, 1), 3.73 (dd, J=7.8, 9.9, 1), 3.51 (dd, J=3.3, 10.5, 1), 2.88-2.75 (m, 1), 2.23-2.10 (m, 1), 2.11 (s, 3), 1.84-1.60 (m, 2), 1.44-1.26 (m, 6), 0.97-0.86 (m, 9 H). MS (ESPOS): 405.4 [M+Hf.
Example 75
Preparation of 3-(3-Methyl-butyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0621] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=3-methyl- butyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R1=R2 =Me, R3=H, R9=3 -methyl-butyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound.
[0622] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J=6.0, 1), 4.55 (d, J=7.5, 1), 4.26 (dd, J=3.6, 10.2, 1), 4.13-4.04 (m, 2), 4.00 (t, J=9.3, 1), 3.80 (d, J=3.3, 1), 3.73 (dd, J=8.1, 10.2, 1), 3.51 (dd, J=3.3, 10.2, 1), 2.87-2.72 (m,T), 2.24-2.10 (m, 1), 2.11 (s, 3), 1.84-1.70 (m, 2), 1.66-1.50 (m, 1), 1.26-1.10 (m, 2), 0.97-0.86 (m, 12 H). MS (ESPOS): 405.0 [M+Hf.
Example 76
Preparation of 3-(3-Cyclobutyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methyIsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0623] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=3- cyclobutyl-propyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R =R =Me, R =H, R =(3 -cyclobutyl-propyl), P =H, P =carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound. [0624] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, .7=5.7, 1), 4.24-4.14 (m, 2), 4.11-4.04 (m, 2), 3.78 (d, .7=3.3, 1), 3.71 (t, J=9.0, 1), 3.54-3.43 (m, 1), 2.76-2.60 (m, 1), 2.34-2.10 (m, 2), 2.10 (s, 3), 2.09-1.97 (m, 2), 1.90-1.51 (m, 7), 1.44-1.11 (m, 3), 0.90 (d, J=2.1, 3), 0.88 (d, J=2.1, 3 H). MS (ESPOS): 431.3 [M+Hf.
Example 77
Preparation of 3-(2-Cyclobutyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0625] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=2- cyclobutyl-ethyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R =R =Me, R3=H, R9=2-cyclobutyl-ethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound. [0626] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, .7=6.0, 1), 4.51 (d, .7=10.2, 1), 4.25 (dd, J=3.3, 9.9, 1), 4.12-4.03 (m, 2), 3.96 (t, --7=9.3, 1), 3.79 (d, .7=3.0, 1), 3.69 (dd, .7=8.1, 10.2, 1), 3.50 (dd, J=3.3, 10.2, 1), 2.85-2.69 (m, 1), 2.36-2.10 (m, 2), 2.10 (s, 3), 2.09-2.00 (m, 2), 1.95- 1.33 (m, 10), 0.92 (d, .7=6.9, 3), 0.89 (d, J=6.9, 3 H). MS (ESPOS): 417.3 [M+Hf.
Example 78
Preparation of 3-(2-Cyclopropyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0627] Lincosamine 2b (R1=Me, R2=Me) was coupled to azetedine acid 16f (R9=2- cyclopropyl-ethyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R!=R2 =Me, R3=H, R9=2-cycloρropyl-ethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound. [0628] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=5.7, 1), 4.53 (d, J=7.2, 1), 4.24 (dd, J=3.3, 9.9, 1), 4.11^1.03 (m, 2), 3.98 (t, J=9.6, 1), 3.79 (d, J=3.0, 1), 3.73 (dd, J=8.1, 9.9, 1), 3.50 (dd,. J=3.3, 10.2, 1), 2.92-2.77 (m, 1), 2.22-2.09 (m, 1), 2.10 (s, 3), 1.95-1.77 (m, 2), 1.33-1.18 (m, . 2), 0.91 (d, .7=6.9, 3), 0.88 (d, .7=6.9, 3), 0.73-0.60 (m, 1), 0.48-0.40 (m, 2), 0.08-0.01 (m, 2 H). MS (ESPOS): 403.3 [M+H]+ .
Example 79
Preparation of 3-(3-Cyclopropyl-propyl)-azetidine-2-carboxylic add [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0629] Lincosamine 2b (R =Me, R =Me) was coupled to azetedine acid 16f (R =3- cyclopropyl-propyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R^R2 =Me, R3=H, R9=3 -cyclopropyl-propyl, P1=H, P2=carboxylic acid-t-butyl ester, m=0) which was deprotected under acidic conditions to provide the title compound. [0630] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, .7=5.7, 1), 4.54 (d, J=7.5, 1), 4.24 (dd, .7=3.3, 9.9, 1), 4.11^1.03 (m, 2), 3.97 (t, J=9.6, 1), 3.79 (d, .7=3.0, 1), 3.71 (dd, J=7.8, 9.9, 1), 3.50 (dd, .7=3.0, 10.2, 1), 2.88-2.74 (m, 1), 2.23-2.09 (m, 1), 2.10 (s, 3), 1.90-1.68 (m, 2), 1.57-1.33 (m, 2), 1.23 (q, J=6.9, 2), 0.91 (d, .7=6.9, 3), 0.89 (d, J=6.9, 3), 0.75-0.60 (m, 1), 0.45-0.38 (m, 2), 0.04-D.02 (m, 2 H). MS (ESPOS): 417.3 [M+Hf.
Example 80
Preparation of 3-Butyl-l-methyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0631] Azetedine acid 16f (R =butyl) was deprotected and N-methylated by treatment with formic acid and formaldehyde under conditions known to persons skilled in the art to provide 3- butyl- l-methyl-azetidine-2-carboxylic acid. Lincosamine 2b (R =Me, R =Me) was coupled to 3-butyl-l-methyl-azetidine-2-carboxylic acid as depicted in general coupling scheme 11 to provide the title compound.
[0632] 1H NMR (300 MHz, D2O) δ 5.36 (d, .7=6.0, 1), 4.67 (d, .7=8.7, 1), 4.31 (t, J=9.3, 1), 4.22 (s, 2), 4.10 (dd, .7=6.0, 10.5, 1), 3.86 (d, .7=3.3, 1), 3.79 (t, --7=9.6, 1), 3.64 (dd, .7=3.3, 10.2, 1), 3.04-2.84 (m, 1), 2.93 (s, 3), 2.25-2.05 (m, 1), 2.14 (s, 3), 1.87-1.68 (m, 2), 1.40-1.20 (m, 4), 0.87 (t, --7=7.5, 9 H). MS (ESPOS): 405.4 [M+H]+. Example 81
Preparation of Phosphoric acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester
[0633] Synthesis of the title compound, 5 (wherein R1=R2 =Me, R3=H, R9=4-ct,s-n-propyl, Rn=PO(OH)2) is detailed in Method N.
[0634] 1H ΝMR (300 MHz, CD3OD) δ 5.41 (d, .7=5.7, 1), 4.53^1.43 (m, 1), 4.10-3.88 (m, 4), 3.73 (dd, .7=3.0, 9.6, 1), 3.44-3.30 (m, 2), 2.25-2.10 (m, 2), 2.11 (s, 3), 2.00-1.88 (m, 1), 1.86-1.70 (m, 1), 1.44-1.25 (m, 6), 0.98-0.88 (m, 9). MS (ESPOS): 485.1 [M+Hf. MS (ESΝEG): 483.0 [M - H]\
Example 82
Preparation of Succinic acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester
[0635] To a suspension R9=4-ct-s-n-propyl, m=2) (721 mg, 1.16 mmol, 1 equiv), succinic anhydride (174 mg, 1.74 mmol, 1.5 equiv) and DMAP (71 mg, 0.58 mmol, 0.5 equiv) in 1,2-dichloroethane (3.0 mL) at 23 °C was added Et3Ν (0.34 mL, 2.43 mmol, 2.1 equiv). After stirring for 2 h at 23 °C, the reaction mixture was partitioned between EtOAc (100 mL) and 10% aqueous citric acid (100 mL). The organic layer was separated, washed with brine (50 mL), dried (MgS04) filtered and concentrated to furnish 840 mg of the desired product 18c (wherein R1=R2 =Me, R3=H, R^-cz'-s-n-propyl, Rn=succinic acid mono ester) (1.16 mmol, 100%) as a white foam. The product was used without further purification.
[0636] MS (ESPOS): 745.2 [M + Naf. MS (ESNEG): 721.2 [M - H]\ [0637] To a solution of protected succinate 18c (wherein R1=R2 =Me, R3=H, R9=4-ct-s-n- propyl, Rπ=succinic acid mono ester) (251 mg, 0.35 mmol, 1 equiv) in 1,2-dichloroethane (20 mL) at 23 °C was added H20 (1.0 mL) followed by TFA (8.0 mL). After stirring for 25 min at 23 °C, toluene (30 mL) was added and the resulting solution was concentrated to 20 mL volume. This solution was further diluted with toluene (30 mL) then concentrated to dryness. The product was purified by semi-preparative HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60% acetonitrile in H20 w/ 0.1% AcOH over 30 min, 20 mL/min flow rate) to give 94 mg of the title compound as a white solid. [0638] 1H NMR (300 MHz, D2O) δ 5.55 (d, J=6.0, 1), 5.15 (dd, J=6.0, 10.5, 1), 4.26-4.16 (m, 2), 3.98-3.87 (m, 3), 3.51 (br d, J=13.5, 1), 3.07 (br t, J=9.9, 1), 2.69 (t, J=7.2, 2), 2.56 (t, J=7.2, 2), 2.28-2.21 (m, 1), 2.16-2.06 (m, 1), 2.13 (s, 3), 2.04-1.96 (m, 1), 1.84-1.70 (m, 1), 1.54-1.28 (m, 6), 0.94-0.84 (m, 9 H). MS (ESPOS): 505.1 [M+Hf. MS (ESNEG): 503.2 [M - H]-.
Example 83
Preparation of i¥-(2-Morpholin-4-yl-etlιyl)--5 ιccinanιic acid 4,5-dihydro-ay-6-{2- methyl-l-[(4-propyl-piperidin-- 2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro- pyran-3-yl ester
[0639] To a solution of protected succinate 18c (wherein R1=R2 =Me, R3=H, R9=4-cw-n- propyl, R11=succinic acid mono ester) (222 mg, 0.31 mmol, 1 equiv) in DMF (3.0 mL) at 0 °C was added 4-(2-aminoethyl)morpholine (60 μL, 0.46 mmol, 1.5 equiv) followed by EDCΗC1 (89 mg, 0.46 mmol, 1.5 equiv) followed by HOBt-H2O (56 mg, 0.37 mmol, 1.2 equiv). After stirring for 20 min at 0 °C the reaction was warmed to 23 °C. After stirring for a further 16 h, volatiles were removed in vacuo and the residue was partitioned between EtOAc (100 mL) and 1:1 brine/10% aqueous citric acid (100 mL). The organic layer was separated, washed with brine (80 mL), dried (MgS04) filtered and concentrated to furnish 242 mg (0.29 mmol, 94%) of 18c (wherein R1=R2 =Me, R3=H, R9=4-ct-s-n-propyl, R11=N-(2-Morpholin-4-yl-ethyl)-succinic acid mono ester) as a clear oil. The product was used without further purification. [0640] MS (ESPOS): 835.5 [M+Hf.
[0641] To a solution of protected succinate 18c (wherein R =R =Me, R =H, R =4-cis-n- propyl, R11=N-(2-Mo holin-4-yl-ethyl)-succinic acid mono ester) (242 mg, 0.29 mmol, 1 equiv) in 1,2-dichloroethane (20 mL) at 23 °C was added H20 (1.0 mL) followed by TFA (8.0 mL). After stirring for 25 min at 23 °C, toluene (70 mL) was added and the resulting -solution wa& concentrated to dryness. The product was purified by semi-preparative HPLC (Waters Νova-Pak® HR Cι8, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60% acetonitrile in H 0 w/ 0.1% HCl over 30 min, 20 mL/min flow rate) to give 47 mg of the title compound as a white solid.
[0642] 1H ΝMR (300 MHz, D20) δ 5.52 (d, J=6.0, 1), 5.12 (dd, J=6.0, 10.5, 1), 4.24-4.16 (m, 2), 3.95-3.85 (m, 3), 3.63 (t, J=6.0, 2), 3.50 (br d, J=13.5, 1), 3.34 (t, J=6.0, 2), 3.05 (dt, J=3.0, 13.5, 1), 2.76 (t, J=6.0, 2), 2.61 (t, J=6.0, 2), 2.28-2.21 (m, 1), 2.16-2.06 (m, 1), 2.11 (s, 3), 2.04-1.96 (m, 1), 1.84-1.70 (m, 1), 1.54-1.28 (m, 6), 0.94-0.84 (m, 9 H); morpholine protons are visible but very broad, causing baseline humps between δ 4.15—3.14 (8 E). MS (ESPOS): 617.5 [M+Hf. MS (ESΝEG): 651.2 [M + CI]". Example 84
Preparation of Dimethylamino-acetic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester
[0643] To a solution of alcohol 18b (wherein m=2) (393 mg, 0.63 mmol, 1 equiv) in CH2C12 (5.0 mL) at 23 °C was added NN-dimethylglycine (72 mg, 0.70 inmol, 1.1 equiv) followed by DCC (390,mg, 1.90 mmol, 3 equiv) followed by DMAP (54 mg, 0.44 mmol, 0.7 equiv). After stirring for ,12 h at 23 °C, the reaction mixture was filtered through a PTFE membrane (0.45 μm) with the aid of CH2C1 (20 mL). The filtrate was partitioned between EtOAc (100 mL) and saturated aqueous ΝaHC03 (80 mL). The organic layer was separated, washed with 1:1 brine: 10% aqueous citric acid (60 mL), brine (50 mL), dried (MgS04) filtered and concentrated to furnish 600 mg of brown solid which was determined to contain the desired product. The crude product was used without further purification.
[0644] MS (ESPOS): 708.5 [M+H]+.
[0645] To a solution of protected glycinate (600 mg, crude) in 1,2-dichloroethane (20 mL) at 23 °C was added H20 (1.0 mL) followed by TFA (8.0 mL). After stirring for 20 min at 23 °C, toluene (100 mL) was added and the resulting solution was concentrated to dryness. The residue was dissolved/suspended in H20 (10 mL) and filtered through a PTFE membrane (0.45 μm) with the aid of H20 (10 mL). The resulting solution was purified by semi-preparative HPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60% acetonitrile in H20 w/ 0.1% HCl over 30 min, 20 mL/min flow rate) to give 189 mg the title compound as a white solid. [0646] 1H NMR (300 MHz, D20) δ 5.62 (d, .7=6.3, 1), 5.30-5.23 (m, 1), 4.27-4.16 (m, 4), 3.95-3.89 (m, 3), 3.50 (br d, .7=12.3, 1), 3.04 (br t, J=13.2, 1), 3.00 (s, 6), 2.28-2.19 (m, 1), 2.16-2.06 (m, 1), 2.13 (s, 3), 2.04-1.96 (m, 1), 1.84-1.70 (m, 1), 1.54-1.28 (m, 6), 0.93-0.83 (m, 9 H). MS (ESPOS): 490.4 [M+Hf. MS (ESNEG): 524.2 [M + CI]".
Example 85
Preparation of 4-Cyclopropylmethyl-pyrrolidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0647] Amino acid intermediate (2S, R)-4-cyclopropylmethyl-pyrrolidine- 1,2-dicarboxylic acid- 1-tert-butyl ester was prepared by the synthetic sequence described by Goodman et al.
Journal of Organic Chemistry, 2003, 68, 3923.
[0648] Lincosamine 2b (R1=Me, R2=Me) was coupled to (2S, 4R)-4-cyclopropylmethyl- pyrrolidine- 1,2-dicarboxylic acid- 1-tert-butyl ester as depicted in general coupling scheme 11 to provide intermediate Ila (R1=R2 =Me, R3=H, R9=cyclopropylmethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=l) which was deprotected under acidic conditions to provide the title compound.
[0649] MS (ESPOS): 403.3 [M+Hf.
Example 86
Preparation of 4-(2-Cyclobutyl-ethyl)-pyrrolidine-2-carbosylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0650] Amino acid intermediate (2S, ¥R)-4-(2-cyclobutyl-ethyl)-pyrrolidine- 1,2- dicarboxylic acid- 1-tert-butyl ester was prepared by general method K by alkylation of pyroglutamic acid ester 7a with (2-bromo-ethylidene)-cyclobutane. The allylic halide (2-bromo- ethylidene)-cyclobutane starting material was prepared from cyclobutanone in two steps as disclosed in U.S. patent 3,711,555.
[0651] Lincosamine 2b (R1=Me, R2=Me) was coupled to protected amino acid 7d (R9=2- cyclobutyl-ethyl) to provide intermediate carbamate Ila (R1=R2 =Me, R3=H, R9=2-cyclobutyl- ethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=l) which was deprotected under acidic conditions to provide the title compound. [0652] MS (ESPOS): 432.3 [M+H]+.
Example 87
Preparation of 4-(2-Cyclopropyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0653] Lincosamine 2b (R1=Me, R2=Me) was coupled to protected amino acid 8c (R9=2- cyclopropyl-ethyl) prepared by method M to provide intermediate carbamate Ila (R =R =Me, R3=H, R9= 2-cyclopropyl-ethyl, P1=H, P2=carboxylic acid-t-butyl ester, m=l) which was deprotected under acidic conditions to provide the title compound. 54] MS (ESPOS): 417.3 [M+Hf.
Example 88
Preparation of 5-Propyl-azepane-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0655] To a solution of protected cyclic amino acid 19f (R12=H, RI3=propyl), (81.4 mg, 0.29 mmol) prepared by general method W, DIEA (100 μL, 0.57 mmol) and 2b (R =Me, R2=Me) HCl salt (83 mg, 0.29 mmol) in anhydrous DMF (2 mL) and HBTU (120 mg, 0.32 mmol) were added. The resulting reaction mixture was stirred for 6 hours at room temperature under N then concentrated to a residue. The residue was taken up in ethyl acetate and washed with 10%) citric acid, saturated sodium bicarbonate, and brine. The organic layer was dried and evaporated to dryness to give the desired l'-N protected lincosamide Ila (R1=Me, R2=Me, R3=H, R9=propyl, P1=H, P2=carboxylic acid-t-butyl ester) (148 mg, 100 %>).
[0656] To a solution of l'-N protected lincosamide Ila (R*=Me, R2=Me, R3=H, R9=propyl, P1=H, P2=carboxylic acidt-butyl ester) (147 mg, 0.285 mmol) in anhydrous DCE (2.0 mL) was added a solution of TES (10 μL, 0.13 mmol) in TFA (7.5% by weight water) 0.44 mL,). The resulting reaction mixture was stirred for one hour at room temperature under N2. The reaction mixture was concentrated and co-evaporated with toluene to furnish the desired product 5- Propyl-2,3,6,7-tefrahydro-lH-azepine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide (151 mg, 100 %) as a white solid which was used in the subsequent reaction without further purification. [0657] MS(ESPOS): 517 [M+Η]+.
[0658] To a solution of 5-Propyl-2,3,6,7-tetrahydro-lH-azepine-2-carboxylic acid [2- methyl-l-(3,4,5-frihydroxy-6-methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide (112.4 mg, 0.212 mmol) in methanol (8.0 mL) was added 10% Pd/C degusa wet form (100 mg). The resulting suspension was hydrogenated at 50 psi for two days. The reaction mixture was filtered through celite, then a 0.45 μm PTFE membrane and evaporation to dryness to furnish the crude product (96 mg, 88 %). The crude product was purified by semi-preparative ΗPLC (Waters Nova-Pak® HR C18, 6 μm particle size, 60 A pore size, 20 mm ID x 100 mm, 5-60% acetonitrile in H20 w/ 0.1% AcOH over 30 min, 20 mL/min flow rate) to provide the title compound (23.5 mg, 21.5 %) as a white solid.
[0659] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, 1). 4.20-4.27 (m, 1), 4.04-4.15 (m, 2), 3.84 (d, 1), 3.41-3.55 (m, 2), 3.10-3.18 (t, 1), 1.88-2.20 (m, 6), 1.50-1.61 (m, 2), 1.25-1.39 (m, 3), 0.88-0.89 (m, 6); MS (ESPOS): 419.4 [M+Hf.
Example 89
Preparation of 4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0660] Lincosamine 2b (R1=Me, R2=Me) was coupled to amino acid intermediate 20o,
(wherein R =H, R =4-tra«-s-n-butyl, n=l) as prepared by method X, as depicted in general 1 9 Q coupling scheme 11 to provide intermediate carbamate Ila (R =R =Me, R =H, R =4-trans-n- 1 9 butyl, P =H, P =carboxylic acid-t-butyl ester, m=2) which was deprotected under acidic conditions to provide the title compound. [0661] MS (ESPOS): 420 [M+H]+.
Example 90
Preparation of 4-(3-Cyclopentyl-propyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsuIfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0662] To a mixture of 13c (R!=R2 =Me, R3=H, R9'=3-Cyclopentyl-prop-l-ynyl) prepared by method Q (100 mg, 0.22 mmol) in MeOH (6 mL) and water (4 mL) were added platinum oxide (200 mg) and cone. HCl (25 μL). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filfration and the filfrate was evaporated. The residue was purified by column chromatography to give the title compound (26.8 mg, 26%) as a white solid. [0663] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J= 5.4, 1), 4.17 (dd, J= 3.3, 10.2, 1), 4.10-
4.03 (m, 2), 3.81 (d, J= 3.3, 1), 3.57-348 (m, 2), 3.27-3.20 (m, 1), 2.84-2.74 (m, 1), 2.19-2.13 (m, 1), 2.10 (s, 3), 2.07-2.00 (m, 1), 1.82-1.70 (m, 4), 1.64-1.50 (m, 5), 1.38-1.25 (m, 6), 1.23-
1.04 (m, 4), 0.94-0.88 (m, 6). MS (ESPOS): 473.7 [M+H]+ .
Example 91 of 4-(3-Methoxy-propyl)-piperidine-2-carboxylic acid [2-ι 1- (334,5-trihydroj-y-6-methyls«lfa]-iyl-tetra-hydro-pyra-n-2-yl)-propyl]-am:
[0664] To a dry flask was added 13b (R =R2 =Me, R3=H) (130 mg, 0.27 mmol, 1 equiv), triphenylphosphine (45.3 mg, 0.17 mmol, 0.64 equiv), copper (I) iodide (32.9 mg, 0.17 mmol, 0.64 equiv), palladium acetate (19.4 mg, 0.086 mmol, 0.32 equiv) and triethylamine (1.5 mL). The mixture was deaerated with nitrogen, followed by addition of methyl propargyl ether (Aldrich) (114 μL, 1.35 mmol, 5 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by column chromatography to give 13c (R =R2 =Me, R3=H, R9 =3-Methoxy-prop-l-ynyl) as a yellow syrup (105 mg, 92%).
[0665] 1H NMR (300 MHz, CDC13) δ 8.52-849 (m, 1), 8.20 (bs, 1), 8.16 (s, 1), 7.46 (dd, J= 1.7, 5.0, 1), 5.36 (d, J= 5.4, 1), 5.03 (d, J= 3, 1), 4.33 (s, 2), 4.27-4.10 (m, 2), 3.98 (d, J= 10.2, 1), 3.75 (t, J= 3.6, 1), 3.55-345 (m, 1), 3.44 (s, 3), 2.67 (d, J= 9.9, 1), 247-240 (m, 1), 2.38 (d, J= 5.1, 1), 2.17 (s, 3), 1.04 (d, J= 7.2, 3), 0.96 (d, J= 6.9, 3); MS (ESPOS): 425.6 [M+H]+ ; MS (ESNEG): 423.5 [M-H]".
[0666] To a mixture of 13c (R1=Me, R9 =3-Methoxy-prop-l-ynyl) (95 mg, 0.22 mmol) in MeOH (6 mL) and water (4 mL) were added platinum oxide (200 mg) and cone. HCl (26 μL). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filfrate was evaporated. The residue was purified by chromatography to give the title compound (8 mg, 8 %) as a white solid. [0667] 1H NMR (300 MHz, CD3OD) δ 5.23 (d, J= 5.7, 1), 4.17 (dd, J= 3.2, 10.1, 1), 4.10- 4.02 (m, 2), 3.80 (d, J= 3.3, 1), 3.53-343 (m, 2), 3.39 (t, J= 6.5, 2), 3.31 (s, 3), 3.27-3.18 (m, 1), 2.80-2.70 (m, 1), 2.21-2.11 (m, 1), 2.10 (s, 3), 2.06-1.98 (m, 1), 1.82-1.74 (m, 1), 1.65-1.54 (m, 3), 1.38-1.28 (m, 2), 1.23-1.08 (m, 2), 0.93-0.88 (m, 6); MS (ESPOS): 435.7 [M+H]+ ; MS (ESNEG): 433.6 [M-H]".
Example 92
Preparation of 4-(3-Ethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methyIsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0668] To a dry flask were added 13b (R1=R2 =Me, R3=H) made using method Q (130 mg, 0.27 mmol, 1 equiv), triphenylphosphine (45.3 mg, 0.17 mmol, 0.64 equiv), copper (I) iodide (32.9 mg, 0.17 mmol, 0.64 equiv), palladium acetate (19.4 mg, 0.086 mmol, 0.32 equiv) and friethylamine (1.5 mL). The mixture was deaerated with nitrogen, followed by addition of ethyl propargyl ether (113 mg, 1.35 mmol, 5 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 13c (R1=R2 =Me, R3=H, R9'=3-Ethoxy-ρroρ-l-ynyl) (78.1 mg, 66 %). MS (ESPOS): 439.6 [M+Hf; MS (ESNEG): 437.5 [M-H]".
[0669] To a mixture of the above syrup in MeOH (6 mL) and water (4 mL) were added platinum oxide (150 mg) and cone. HCl (14 μL). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filfration and the filtrate was evaporated. The residue was purified by chromatography to give the title compound (20 mg, 25 %) as a white solid.
[0670] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J= 5.7, 1), 4.21 (dd, J= 3, 9.9, 1), 4.11-4.04 (m, 2), 3.89-3.80 (m, 2), 3.54-3.38 (m, 6), 3.10-2.98 (m, 1), 2.25-2.12 (m, 2), 2.11 (s, 3), 2.00- 1.90 (m, 1), 1.78-1.56 (m, 2), 144-1.27 (m, 5), 1.17 (t, J= 6.9, 3), 0.94-0.88 (m, 6). MS (ESPOS): 449.6 [M+Hf; MS (ESNEG): 447.7 [M-H]". Example 93
Preparation of 4-(3-Propoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l- (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0671] To a dry flask were added 13b (R*=R2 =Me, R3=H) made using method Q (150 mg, 0.31 mmol, 1 equiv), triphenylphosphine (52.4 mg, 0.2 mmol, 0.64 equiv), copper (I) iodide (38.1 mg, 0.2 mmol, 0.64 equiv), palladium acetate (22.4 mg, 0.1 mmol, 0.32 equiv) and triethylamine (1.7 mL). The mixture was deaerated with nitrogen, followed by addition of propargyl ether (117 mg, 1.24 mmol, 4 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 13c (R1=R2 =Me, R3=H, R9 =3-prop-l-ynyl-oxy-prop-l-ynyl) (40.9 mg, 29 %). MS (ESPOS): 449.2 [M+Hf.
[0672] To a mixture of the above syrup in MeOH (6 mL) and water (4 mL) were added platinum oxide (100 mg) and 0.1037 N HCl (0.86 mL, 0.089 mmol, 0.98 equiv). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filtrate was evaporated. The residue was purified by chromatography to give a white solid, the title compound (11 mg, 26 %). [0673] 1HNMR (300 MHz, CD3OD) δ 5.24 (d, J= 5.7, 1), 4.17 (dd, J= 3.2, 10.1, 1), 4.10- 4.02 (m, 2), 3.79 (d, J= 3, 1), 3.53-3.32 (m, 6), 3.25-3.18 (m, 1), 2.80-2.68 (m, 1), 2.22-2.12 (m, 1), 2.10 (s, 3), 2.06-1.98 (m, 1), 1.83-1.74 (m, 1), 1.66-1.08 (m, 9), 0.97-0.86 (m, 9); MS (ESPOS): 463.4 [M+Hf. Example 94
Preparation of 4-(3-Cyclopropylmethoxy-propyl)-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0674] To a dry -flask were added 13b (R1=R2 =Me, R3=H) made using method Q (200 mg, 0.42 mmol, 1 equiv), triphenylphosphine (69.7 mg, 0.27 mmol, 0.64 equiv), copper (I) iodide (50.6 mg, 0.27 mmol, 0.64 equiv), palladium acetate (29.8 mg, 0.13 mmol, 0.32 equiv) and triethylamine (2.4 mL). The mixture was deaerated with nitrogen, followed by addition of prop- 2-ynyloxymethyl-cyclopropane (229 mg, 2.08 mmol, 5 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give a 13c (R1=R2 =Me, R3=H, R9 =3-Prop-2-ynyloxymethyl- cyclopropane) (113 mg, 59 %). MS (ESPOS): 465.1 [M+Hf.
[0675] To a mixture of 13c (R1=R2 =Me, R3=H, R9'=3-Prop-2-ynyloxymethyl-cyclopropane) in MeOH (6 mL) and water (4 mL) were added platinum oxide (230 mg) and cone. HCl (19.3 μL, 0.232 mmol, 0.95 equiv). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filfration and the filtrate was evaporated. The residue was purified by chromatography to give a white solid, the title compound (20 mg, 17 %).
[0676] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J= 5.7, 1), 4.21 (dd, J= 3.6, 9.9, 1), 4.11- 4.04 (m, 2), 3.87-3.78 (m, 2), 3.54-3.38 (m, 4), 3.08-2.98 (m, 1), 2.27-2.07 (m, 2), 2.11 (s, 3), 2.01-1.91 (m, 1), 1.80-1.25 (m, 9), 1.01-0.85 (m, 7), 0.56-047 (m, 2), 0.22-0.15 (m, 2); MS (ESPOS): 475.2 [M+Hf. Example 95
Preparation of 4-[3-(2-Fluoro-ethoxy)-propyl]-piperidine-2-carboxylic acid [2- m -amide
[0677] To a dry flask were added 13b (R1=R2 =Me, R3=H) made using method Q (230 mg, 048 mmol, 1 equiv), triphenylphosphine (80.1 mg, 0.31 mmol, 0.64 equiv), copper (I) iodide (58.2 mg, 0.31 mmol, 0.64 equiv), palladium acetate (34.2 mg, 0.15 mmol, 0.32 equiv) and triethylamine (2.7 mL). The mixture was deaerated with nitrogen, followed by addition of 3-(2- Fluoro-ethoxy)-propyne (244 mg, 2.39 mmol, 5 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was purified by chromatography to give 13c (R1=R2 =Me, R3=H, R9 =3-(2-Fluoro-ethoxy)-propyne) (159 mg, 73%).
[0678] To a mixture of 13c (R*=R2 =Me, R3=H, R9'=3-(2-Fluoro-ethoxy)-propyne) (159 mg, 73 %). in MeOH (6 mL) and water (4 mL) were added platinum oxide (320 mg) and cone. HCl (27.7 μL, 0.33 mmol, 0.95 equiv). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filtrate was evaporated. The residue was purified by chromatography to give a white solid, the title compound (25 mg, 15%).
[0679] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J= 5.7, 1), 4.60-4.56 (m, 1), 444-4.39 (m, 1), 4.20 (dd, J= 3.3, 9.9, 1), 4.10-4.04 (m, 2), 3.81-3.75 (m, 2), 3.73-3.68 (m, 1), 3.63-3.58 (m, 1), 3.54-348 (m, 3), 343-3.34 (m, 1), 3.04-2.93 (m, 1), 2.23-2.12 (m, 2), 2.11 (s, 3), 1.98-1.88 (m, 1), 1.78-1.25 (m, 7), 0.95-0.87 (m, 6). MS (ESPOS): 467.2 [M+Hf. Example 96
Preparation of 4-[3-(3-Fluoro-propoxy)-propyl]-piperidine-2-carboxylic acid [2- m amide
[0680] To a dry flask were added 13b (R1=R2 =Me, R3=H) made using method Q (230 mg, 0.48 mmol, 1 equiv), triphenylphosphine (80.1 mg, 0.31 mmol, 0.64 equiv), copper (I) iodide (58.2 mg, 0.31 mmol, 0.64 equiv), palladium acetate (34.2 mg, 0.15 mmol, 0.32 equiv) and triethylamine (2.7 mL). The mixture was deaerated with nitrogen, followed by addition of 3-(3- Fluoro-propoxy)-propyne (277 mg, 2.39 mmol, 5 equiv). The mixture was stirred at 50 °C overnight. The solvent was removed under vacuum to give a dark residue. The residue was- purified by chromatography to give 13c (R1=R2 =Me, R3=H, R9 =3-(3-Fluoro-propoxy)- propyne) (137 mg, 61 %).
[0681] To a mixture of the 13c (R1=R2 =Me, R3=H, R9 =3-(3-Fluoro-propoxy)-propyne) in MeOH (6 mL) and water (4 mL) were added platinum oxide (280 mg) and cone. HCl (23.1 μL, 0.28 mmol, 0.95 equiv). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filtrate was evaporated. The residue was purified by chromatography to give a white solid, the title compound (23 mg, 16 %).
[0682] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J= 5.7, 1), 4.57 (t, J= 5.9, 1), 4.41 (t, J= 6, 1), 4.20 (dd, J= 3.2, 10.1, 1), 4.10-4.04 (m, 2), 3.81 (d, J= 3, 1), 3.74-3.67 (m, 1), 3.55-342 (m, 5), 3.38-3.32 (m, 1), 2.98-2.87 (m, 1), 2.23-2.08 (m, 2), 2.11 (s, 3), 2.00-1.83 (m, 3), 1.72-1.57 (m, 2), 142-1.20 (m, 5), 0.97-0.87 (m, 6). MS (ESPOS): 481.2 [M+Hf. Example 97
Preparation of 4-(4-Methoxy-butyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- -amide
[0683] To a mixture of NaH (60 %, 37.6 mg, 0.94 mmol, 1 equiv) in dry DMF (2 mL) at 0 °C was added a solution of 14c (R9=4-methoxybutyl) prepared by method R (197 mg, 0.94 mmol, 1 equiv) in DMF (0.5 mL) dropwise. The mixture was stirred at 0 °C for 20 min, then was cooled to -78° C. To the mixture was added iodomethane (134 mg, 0.94 mmol, 1 equiv). The mixture was stirred at 0°C for 2 hr, then at rt overnight. The reaction mixture was diluted with DCM, washed with brine, dried and concentrated. The residue was purified by preparative TLC to give 4-(4-Methoxy-butyl)^pyridine-2-carboxylic acid methyl ester (43 mg, 21%). [0684] 1H NMR (300 MHz, CDC13) δ 8.61-8.58 (m, 1), 7.97-7.95 (m, 1), 729-7.26 (m, 1), 3.98 (s, 3), 3.37 (t, J= 6, 2), 3.30 (s, 3), 2.70 (t, J= 7.7, 2), 1.78-1.54 (m, 4). [0685] To a mixture of 4-(4-Methoxy-butyl)-pyridine-2-carboxylic acid methyl ester ( (107 mg, 0.48 mmol, 1 equiv) in THF (1.5 mL) and water (0.5 mL) was added lithium hydroxide monohydrate (30.2 mg, 0.72 mmol, 1.5 equiv). The mixture was stirred at rt for 4 h and diluted with methanol (20 mL). Then H1" resin (0.4 g) was added and the mixture was shaken for 10 minutes. The resin was washed with methanol (lx), 1 : 1 acetonitrile/water (lx), and acetonitrile (lx). The product was eluted with 5% TEA in methanol (4x) and acetonitrile (lx). The combined organic solvents were evaporated and co-evaporated with toluene to give 4-(4- Methoxy-butyl)-pyridine-2-carboxylic acid (63.4 mg, 63 %): 1H NMR (300 MHz, CD3OD) δ 847-843 (m, 1), 7.98-7.94 (m, 1), 743-7.37 (m, 1), 3.41 (t, J= 6.3, 2), 3.30 (s, 3), 2.75 (t, J= 7.5, 2), 1.80-1.55 (m, 4).
[0686] To a solution of 4-(4-Methoxy-butyl)-pyridine-2-carboxylic acid (63.4 mg) in dry DMF (0.75 mL) at 0 °C were added a mixture of 7-Me MTL HCl salt 2b (R*=Me, R2=Me) (87.3 mg, 0.30 mmol, 1 equiv) and DIEA (78 mg, 0.61 mmol, 2 equiv) in DMF (0.75 mL), followed by the addition of HBTU (100 mg, 0.73 mmol, 1 equiv). The reaction mixture was stirred at rt for 3 h. The reaction mixture was evaporated under high vacuum to dryness. The residue was purified by chromatography to give a syrup (88 mg, 65 %).
[0687] MS (ESPOS): 443.0 [M+Hf; MS (ESNEG): 441.2 [M-H]".
[0688] To a mixture of the above syrup (88 mg, 0.20 mmol, 1 equiv) in MeOH (6 mL) and water (4 mL) were added cone. HCl (15.7 μL, 0.19 mmol, 0.95 equiv) and platinum oxide (180 mg). The mixture was purged and charged with hydrogen (65 psi) and shaken overnight. The platinum oxide was removed by filtration and the filfrate was evaporated to give a residue, which was purified by chromatography to provide the title compound (lower isomer, 20 mg,
22%) as a white solid.
[0689] 1H NMR (300 MHz, CD3OD) δ 5.22 (d, J= 5.7, 1), 4.15 (dd, J= 3.2, 10.1, 1), 4.09-
4.00 (m, 2), 3.78 (d, J= 2.7, 1), 3.49 (dd, J= 3.3, 10.5, 1), 3.38 (t, J= 6.3, 2), 3.30 (s, 3), 3.26-
3.22 (m, 1), 3.16-3.08 (m, 1), 2.66-2.55 (m, 1), 2.20-2.11 (m, 1), 2.09 (s, 3), 1.96-1.87 (m, 1),
1.73-1.64 (m, 1), 1.58-1.22 (m, 7), 1.11-0.97 (m, 2), 0.93-0.86 (m, 6); MS (ESPOS): 449.4
[M+Hf; MS (ESNEG): 447.2 [M-H]".
Example 98
Preparation of 4-Propoxymethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0690] To (l-Oxy-pyridin-4-yl)-methanol (5g, 0.04 mol) in a solution of dichloromethane (10 mL) and pyridine (10 mL) was added acetic anhydride (12.2 mL, 0.12 mol) and the reaction mixture was stirred for 16 h at room temperature. The reaction mixture was then poured into water and extracted with dichloromethane (200 mL) and then washed with brine (100 mL). The crude product obtained (4.27 g, 64%) was taken without purification for the next reaction. To the crude product (4.27 g, 25.6 mmol) in DCM (25 mL) and trimethylsilyl cyanide (3.40 mL, 25.6 mmol), dimethylcarbamyl chloride (2.35 mL, 25.6 mmol) was added slowly and then the reaction mixture was stirred at room temperature over night. Aqueous potassium carbonate (100 mL, 10%) was added and stirred for 10 minutes. Extraction with ethyl acetate followed by removal of solvent resulted in crude product which was purified on silica gel column chromatography using 50% ethyl acetate in hexanes to obtain cyanopyridine 10a (R9 = acetoxymethylene) (2.37 g, 48%).
[0691] 1H NMR (300 MHz, CDC13) δ 8.68 (d, J= 5.7 Hz, 1), 7.65 (s, 1), 7.45 (d, J= 5.7 Hz, 1), 5.13 (s, 2), 2.16 (s, 3). MS (ESPOS): 199 [M+Naf.
[0692] To acetic acid 2-cyano-pyridin-4-ylmethyl ester 10a (R9 = acetoxymethylene) (2.37 g, 0.012 mol), aqueous hydrochloric acid (50 mL, 6N) was added and refluxed for 16 hr. Stripping off hydrochloric acid and water resulted in acid 10b (R9 = acetoxymethylene) (247 g, 100%).
[0693] 1H NMR (300 MHz, CD3OD) δ 8.83 (d, J= 5.7 Hz, 1), 8.56 (s, 1), 8.24 (d, J= 5.7 Hz, 1), 4.92 (s, 2 MS (ESNEG): 152 [M-H]".
[0694] To the 10b (R9 = acetoxymethylene) (2.40 g, 0.012 mol) in methanol (25 mL), water (15 mL) and acetic acid (15 mL), platinum dioxide (1 g) was added and hydrogenated at 50 psi for 16 h. The catalyst was filtered and the solvent was removed to obtain the hydrogenated product (2.50 g, 100%>) which was taken as such for the next reaction. To the crude product from previous reaction in dioxane (30 mL) and water (30 mL), sodium hydroxide (2.2 g, 0.05 mol) " and di t-butyldicarbonate (12 g, 0.05 mol) was added and stirred at room temperature for 16 h. The reaction mixture was then extracted with ethyl acetate (100 mL) to remove any excess Boc- anhydride. The aqueous phase was then carefully acidified with dil. HCl (IN) and extracted with ethyl acetate. The crude product obtained on removal of solvent was purified on silica gel column chromatography using ethyl acetate to obtain the 4-hydroxymethyl-piperidine- 1,2- dicarboxylic acid 1-tert-butyl ester (0.62 g, 20%).
[0695] 1H NMR (300 MHz, CD3OD) δ 4.27 (d, J= 6.3 Hz, 1), 3.37-3.65 (m, 4), 2.05 (m, 1), 1.70-1.84 (m, 4), 1.52 (s, 9). MS (ESNEG): 258 [M-H]".
[0696] To the 4-hydroxymethyl-piperidine- 1,2-dicarboxylic acid 1-tert-butyl ester (100 mg, 0.39 mmol) in DMF (5 mL), sodium hydride (30 mg, 0.67 mmol) was added and stirred at 0 °C for 10 minutes. Propyl bromide (0.5 mL, 4.0 mmol) (0.5 mL, 0.35 mmol) was added and stirred at room temperature for 30 minutes and then at 50-60 °C for 2 h. DMF was removed, water (50 mL) was added and extracted with ethyl acetate (50 mL). The product obtained on removal of solvent was purified on column chromatography using 50%» ethyl acetate in hexanes to provide 4-Butoxymethyl-piperidine- 1,2-dicarboxylic acid 2-butyl ester 1-tert-butyl ester (100 mg, 75%>). [0697] 1H NMR (300 MHz, CDC13) δ 4.10 (q, J= 6.6, 7.2 Hz, 4), 346-3.53 (m, 2), 3.25- 3.36 (m, 2), 2.05 (m, 2), 144-1.75 (m, 6), 1.45 (s, 9), 0.88-0.98 (m, 6). MS (ESPOS): 366
[0698] To the 4-Butoxymethyl-piperidine- 1,2-dicarboxylic acid 2-butyl ester 1-tert-butyl ester (100 mg, 0.29 mmol) in THF (5 mL), lithium hydroxide (122 mg, 2.90 mmol) in water (1 mL) was added and stirred at room temperature for 16 h. It was then poured into water and exfracted with ethyl acetate and discarded. The water layer was acidified and then exfracted with ethyl acetate (50 mL). Removal of solvent gave the acid which was taken as such for the next coupling reaction. To 4-Butoxymethyl-piperidine- 1,2-dicarboxylic acid 1-tert-butyl ester (88 mg, 0.29 mmol) in DMF (3 mL), the amine 2b (R1=Me, R2=Me) (100 mg, 0.34 mmol), HBTU (132 mg, 0.34 mmol) and DIEA (0.1 mL, 0.58 mmol) was added and stirred at room temperature for 3 h. DMF was removed and product was purified on silica gel column chromatography using 50% ethyl acetate in hexanes provided the Boc protected lincosamide (R9 = 4-Butoxymethyl) (77 mg, 49%).
[0699] 1H NMR (300 MHz, CD3OD) δ 5.25 (m, 1), 447-4.64 (m, 2), 4.41 (t, J= 6.0 Hz, 1), 4.26 (s, 1), 3.99-4.12 (m, 4), 3.85 (d, J= 3.3 Hz, 1), 3.50-3.54 (m, 2), 3.31-3.36 (m; 2), 2.14- 2.2.19 (m, 2), 1.81 (m, 3), 1.53 (m, 2), 1.45 (bs, 12), 0.88-0.98 (m, 9). MS (ESPOS): 535
[0700] To the above Boc protected lincosamide (R9 = 4-Butoxymethyl) (77 mg, 0.14 mmol), 30% TFA in DCE (30 mL), triethylsilane (1 mL) and water (1 mL) was added and stirred at room temperature for 1 hr. The residue obtained on removing the solvent was chromatographed using 20%) methanol in DCM to obtain the title compound (10 mg, 16%) as a white solid. [0701] 1H NMR (300 MHz, CD3OD) δ 5.25 (d, J= 5.7 Hz, 1), 4.05-4.19 (m, 3), 3.74-3.86 (m, 2), 3.50-3.54 (m, 2), 340-3.54 (m, 4), 2.14 (m, 2), 2.10 (s, 3), 2.00 (m, 1), 1.71-1.75 (m, 2), 1.54-1.61 (m, 2), 1.31 (m, 1), 0.89-0.94 (m, 9). MS (ESPOS): 435 [M+Hf.
Example 99
Preparation of 4-(3-Fluoro-propoxymethyl)-piperidine-2-carboxylic acid [2-methyl- l-(3,4,5-trihyd e
[0702] The title compound was prepared by the same reaction sequence described in example 98 starting from Boc protected 4-hydroxymethyl-piperidine- 1,2-dicarboxylic acid 1- tert-butyl ester substituting the reagent 3-fluoropropyl bromide in the alkylation. [0703] 1H NMR (300 MHz, CD3OD) δ 5.22 (d, J= 54 Hz, 1), 4.58 (q, J= 3.9, 6.9 Hz, 1), 4.22 (q, J= 5.1, 5.7 Hz, 1), 3.90-4.16 (m, 3), 3.82-3.86 (m, 1), 3.74 (d, J= 2.1 Hz, 1), 347-3.59 (m, 3), 3.35-3.39 (m, 1), 3.04 (m, 2), 2.07 (s, 3), 1.85-1.98 (m, 4), 147-1.56 (m, 2), 1.02-1.12 (m, 2), 0.90-0.96 (m, 6). MS (ESPOS): 453 [M+Hf.
Example 100
Preparation of l-(5-Methyl-2-oxo-[l,3]dioxol-4-ylmethyl)-4-propyl-piperidine-2- carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propyl] -amide
[0704] To the title compound from example 17 (50 mg, 0.123 mmol) in DMF (5 mL) at room temperature, sodium carbonate (25 mg, 0.246 mmol) and 4-bromomethyl-5-methyl- [l,3]dioxol-2-one, prepared as described in J. Alexander, etal. J. Med. Chem, 1996, 39, 480- 486. (47 mg, 0.246 mmol) were added and stirred over night at room temperature. DMF was removed and the crude material was loaded into a silica gel column and eluted with a mixture of methanol (5%) and dicliloromethane to obtain the title compound as a white solid (30 mg, 48%). [0705] TLC: Rf = 0.5 (EtOAc). MS (ESPOS):517 [M+Hf; 1H NMR (CD3OD, 200 MHz): δ 5.22 (d, J= 3.8 Hz, 1), 4.05-4.20 (m, 3), 3.86 (d, J= 2 Hz, 1), 3.51-3.60 (m, 2), 2.88-3.01 (m, 2), 2.17 (m, 2), 2.13 (s, 3), 2.10 (s, 3), 1.88 (d, J= 6.2 Hz, 1), 1.70 (d, J= 8.2 Hz, 1), 1.21-1.35 (m, 6), 0.90-0.96 (m, 9).
Example 101
Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyI-tetrahydro-pyran- 2-yl)-propyIcarbamoyl]-4-propyl-piperidine-l-carboxylic acid 5-methyl-2-oxo-[l,3]dioxol- 4-ylmethyl ester
[0706] To the title compound from example 17 (50 mg, 0.123 mmol) in DMF (5 mL) at room temperature, potassium bicarbonate (20 mg, 0.30 mmol) and carbonic acid 5-methyl-2- oxo-[l,3]dioxol-4-ylmethyl ester 4-nitro-phenyl ester, prepared as described in F. Sakamoto, etal, Chem. Pharm. Bull. 1984, 32 (6), 2241-2348. (36 mg, 0.123 mmol) were added and stirred over night at room temperature. DMF was removed and the crude material was loaded into a silica gel column and eluted with a mixture of methanol (5%) and dichloromethane to obtain the title compound (30 mg, 44%) as a white solid.
[0707] TLC: Rf = 0.5 (EtOAc). MS (ESPOS): 561 [M+H]+; 1H NMR (CD3OD, 200 MHz): δ 5.23 (d, J= 3.8 Hz, 1), 4.92 (m, 2), 4.21-4.24 (m, 1), 4.07-4.13 (m, 3), 3.98 (m, 1), 3.57 (d, J= 2.2 Hz, 1), 3.54 (d, J= 2.2 Hz, 1), 2.16 (s, 3), 2.08 (s, 3), 1.95-1.97 (m, 2), 1.76-1.85 (m, 2), 1.60 (m, 1), 1.32 (m, 4), 0.88-0.98 (m, 9). Example 102
Preparation of Hexadecanoic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester
[0708] The title compound, 5 (wherein R =R2 =Me, R3=H, R6=H, R9=4- -s-n-propyl, Rπ=CO(CH2) 14CH3, m=2) may be prepared employing method N by treatment of alcohol 18b (wherein R1=R2=Me, R3=H, R6=Boc, R9=4-cis-n-Pr, and m=2) with the acylating reagent palmitoyl chloride.
Example 103
Preparation of l-(l-Methyl-3-oxo-but-l-enyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0709] The title compound may be prepared employing the method described by Jensen et al, Journal of Medicinal Chemistry 1980, 23, by treatment of 1 (wherein R =R =Me, R =H R6=H, R9=4-cis-n-propyl, and m=2) with acetylacetone. Example 104
Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid 1-acetoxy-ethyl ester
[0710] The title compound may be prepared employing the method described by Alexander et al, Journal of Medicinal Chemistry 1988, 31, 318-322, by treatment of 1 (wherein R1=R2=Me, R =H, R =H, R =4-cis-n-propyl, and m=2) with the acylating reagent Acetic acid l-(4-nitro- phenoxycarbonyloxy)-ethyl ester.
Example 105
Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propylcarbamoyl]-4-propyl-piperidine-l-carboxy lie acid 2-amino-3-methyl- pentanoyloxymethyl ester
[0711] The title compound may be prepared employing the method described by Alexander et al, Journal of Medicinal Chemistry 1988, 31, 318-322, by treatment of 1 (wherein R*=R2=Me,
R3=H, R6=H, R9=4-cis-n-propyl, and m=2) with the appropriate acylating reagent (2-tert- butoxycarbonylamino-3-methyl-pentanoic acid 4-nitro-phenoxycarbonyloxymethyl ester), followed by TFA deprotection. Example 106
Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propylcarbamoyl]-4-propyl-piperidine-l-carboxy lie acid piperidine-4- carbonyloxymethyl ester
[0712] The title compound may be prepared employing the method described by Alexander et al, Journal of Medicinal Chemistry 1988, 31, 318-322, by treatment of 1 (wherein R^R^Me, R3=H, R6=H, R9=4-cis-n-propyl, and m=2) with the appropriate acylating reagent (piperidine- 1,4-dicarboxylic acid 1-tert-butyl ester 4-(4-nifro-phenoxycarbonyloxymethyl) ester), followed by TFA deprotection.
Example 107
Preparation of l-(Propionylamino-methyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0713] The title compound may be prepared from 1 (wherein R =R =Me, R =H, R =H, R9=4-cis-n-propyl, and m=2) employing the method described by Bundgaard et al, Journal of Pharmaceutical Sciences, 1980, 69 (1), 44-46. Example 108
Preparation of N-{2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro- pyran-2-yl)-propylcarbamoyl]-4-propyl-piperidin-l-ylmethyl}-nicotinamide
[0714] The title compound m
R9=4-cis-n-propyl, and m=2) employing the method described by Bundgaard et al, Journal of
Pharmaceutical Sciences, 1980, 69 (1), 44-46.
Example 109
Preparation of l-(2-Amino-propionyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0715] The title compound may be prepared employing the method described by Weiss et al,
Antimicrobial Agents and Chemotherapy, 1999, 43 (3), 460-464, by treatment of 1 (wherein R1=R2=Me, R3=H, R6=H, R9=4-cis-n-ρroρyl, and m=2) with Boc-alanine and HATU in the presence of DIEA in an appropriate solvent followed by TFA deprotection. Example 110
Preparation of l-(2-Amino-3-phenyl-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0716] The title compound may be prepared employing the method described by Weiss et al, Antimicrobial Agents and Chemotherapy, 1999, 43 (3), 460^464, by treatment of 1 (wherein R1=R2=Me, R3=H, R6=H, R9=4-cis-n-propyl, and m=2) with Boc-phenylalanine and HATU in the presence of DIEA in an appropriate solvent followed by TFA deprotection.
Example 111
Preparation of l-(2-Amino-3-methyl-pentanoyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0717] The title compound may be prepared employing the method described by Weiss et al,
Antimicrobial Agents and Chemotherapy, 1999, 43 (3), 460-464, by treatment of 1 (wherein
R1=R2=Me, R3=H, R6=H, R9=4-cis-n-propyl, and m=2) with Boc-isoleucine and HATU in the presence of DIEA in an appropriate solvent followed by TFA deprotection. Example 112
Preparation of l-(2-Amino-3-methyl-butyryl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0718] The title compound may be prepared employing the method described by Weiss et al,
Antimicrobial Agents and Chemotherapy, 1999, 43 (3), 460-464, by treatment of 1 (wherein
R1=R2=Me, R3=H, R6=H, R9=4-cis-n-propyl, and m=2) with with Boc-valine and HATU in the presence of DIEA in an appropriate solvent followed by TFA deprotection.
Example 113
Preparation of l-(l-Methyl-l,4-dihydro-pyridine-3-carbonyl)-4-propyl-piperidine- 2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyI-tetrahydro-pyran-2-yl)- propyl] -amide
[0719] The title compound m
R9=4 -cis-n-propyl, and m=2) employing the method described by Shek et al, Journal of
Medicinal Chemistry 1976, 19 (1), 108-112. Example 114
Preparation of 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran- 2-yl)-propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid l-methyl-l,2,3,6-tetrahydro- pyridin-4-yl ester
[0720] The title compound may be prepared from 1 (wherein R1=R2=Me, R3=H, R6=H,
R9=4-cis-n-propyl, and m=2) employing the method described by Flaherty et al, Journal of
Medicinal Chemistry 1996, 39, 4756-4761.
Example 115
4-Cyclohexylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0721] The intermediate 1 ying method P using 4- benzylpyridine as the starting material.
[0722] !H NMR (300 MHz, CD3OD) δ 8.78 (m, 1), 8.40 (s, 1), 8.03 (m, 1), 7.26-740 (m, 5),
4.39 (s, 2); MS (ESNEG): 212 [M - 1]".
[0723] Lincosamine 2b (R1=Me, R2=Me) was coupled to 4-Benzylpyridine-2-carboxylic acid 10b (R9=4-benzyl) as depicted in general coupling scheme 11 to provide intermediate lib
(R1=Me, R2=Me, R3=H, R9=benzyl, P1=H), which was reduced by catalytic hydrogenation to the title compound. [0724] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 54, 1), 4.03-4.19 (m, 4), 3.78 (m, 1), 348-3.52 (m, 2), 3.21-3.24 (m, 2), 2.77 (t, J = 12.6, 1), 2.17 (m, 2), 2.15 (s, 3), 1.98-2.02 (m 1), 1.70-1.74 (m, 60), 1.12-1.37 (m, 8), 0.91 (d, J = 6.9, 6); MS [ESPOS]: 459 [M + H]+.
Example 116
4-(2-Propyloxyethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide
[0725] To a solution of 4-(2-hydroxyethyl)-pyridine (2.15 g, 17.48 mmol, 1 equiv) in glacial acetic acid (11.7 mL) was added 30 % hydrogen peroxide (1.98 mL, 17.48 mmol, 1 equiv). The reaction mixture was refluxed overnight and concentrated. The residue was dissolved in DCM, and dried (MgSQ4). Solvent was evaporated under vacuum and then again co-evaporated with toluene to give the N-oxide intermediate as a syrup.
[0726] A solution of the above syrup in DCM (17 mL) was added to trimethylsilyl cyanide (2.95 mL, 22.14, 1.27 equiv) at r.t. and then cooled to 0 °C. To the mixture was added dropwise a solution of dimethylcarbamyl chloride in DCM (4.4 mL) over 10 min. The reaction mixture was stirred at r.t. overnight. A solution of 10% aqueous potassium carbonate (17.5 mL) was added dropwise and the mixture was stirred for 10 min. The organic layer was separated and the aqueous layer was extracted with DCM (2x). The combined organic layers were dried and concentrated. The residue was purified by chromatography to give the nitrile intermediate 10a (R9=2-hydroxyethyl) (0.82 g, 25 %) as a yellow oil.
[0727] 1HNMR (300 MHz, CDC13) δ 8.61 (d, J = 5.1, 1), 7.56 (d, J = 0.6, 1), 7.39-7.34 (m, 1), 4.31 (t, J = 6.5, 2), 2.99(t, J = 6.3, 2), 2.02 (s, 3).
[0728] A solution of above nitrile 10a (R9=2-hydroxyethyl) (0.82 g, 4.32 mmol) in 6 N HCl (2 mL) was refluxed overnight. The reaction mixture was concentrated under vacuum. MeOH (18 mL) and water (18 mL) were added, followed by cone. aq. HCl (2.2 mL) and platinum oxide (450 mg). The mixture was purged and charged with hydrogen (1 atm) and stirred overnight. The platinum oxide was removed by filtration and the filtrate evaporated under vacuum to afford the intermediate pipecolic acid intermediate.
[0729] The above pipecolic acid intermediate was taken in 2N NaOH (5 mL) and t-butanol (5 mL), and di-t-butyl dicarbonate (1.33 g, 6.1 mmol) was added with stirring. The mixture was stirred at r.t. overnight. The solvent was removed under vacuum. The residue was diluted with water and organic layer washed with ether. The aqueous layer was acidified with 2N HCl to pH = 2.0, and exfracted with ethyl acetate (twice). The combined organic layers were dried (MgS04) and concentrated under vacuum. The crude N-Boc pipecolinic acid intermediate was purified by chromatography to give 4-(2-hydroxyethyl)-piperidine-l,2-carboxylic acid- 1-tert- butyl ester (210 mg, 18 %) as a clear syrup.
[0730] 1H NMR (300 MHz, CDC13) δ 445-4.38 (m, IH), 3.64 (t, J = 7.1, 2), 3.63-3.56 (m, 1), 3.34-3.24 (m, 1), 1.99-1.94 (m, 2), 1.85-1.65 (m, 2), 1.62-1.53 (m, 2), 148-143 (m, 1), 1.42 (s, 9).
[0731] Sodium hydride (60% in oil, 28.9 mg, 0.72 mmol, 2 equiv) was washed with hexane (2x), and dried under high vacuum. A solution of 4-(2-hydroxyethyl)-piperidine-l,2-carboxylic acid- 1-tert-butyl ester (98.6 mg, 0.36, 1 equiv) in DMF (0.8 mL) was added to a mixture of the sodium hydride in DMF (0.7 mL) at 0 °C. The mixture was stirred at 0°C for 5 min, and then at r.t. for 20 min. 1-Bromopropane (0.33 mL, 3.6 mmol, 10 equiv) was added and the mixture stirred at r.t. for 3h and then quenched with water. The mixture was diluted with ethyl acetate, washed with water (lx), brine (lx), dried and concentrated to give a clear oil, which was purified by chromatography to give the ether intermediate 4-(2-propyloxyethyl)-piperidine-l,2- carboxylic acid- 1-tert-butyl ester-2-propyl ester (35.8 mg, 28 %>).
[0732] 1H NMR (300 MHz, CDC13) δ 4.36-4.29 (m, 1), 4.05 (t, J = 6.8, 2), 3.65-3.53 (m, 1), 3.39 (t, J = 6.5, 2), 3.32 (t, J = 6.8, 2), 3.33-3.25 (m, 1), 2.03-1.92 (m, 1), 1.87-1.70 (m, 3), 1.68- 1.48 (m, 6), 1.41 (s, 9), 1.41-1.34 (m, 1), 0.97-0.84 (m, 6); MS (ESPOS): 258.2 [M - Boc + H]+ . [0733] To a solution of ether intermediate 4-(2-propyloxyethyl)-piperidine- 1 ,2-carboxylic acid- 1-tert-butyl ester-2-propyl ester (73 mg, 0.20 mmol, 1 equiv) in dioxane (1 mL) and water (0.8 mL) was added lithium hydroxide monohydrate (84 mg, 2 mmol, 10 equiv). The reaction mixture was stirred at r.t. overnight. The solvent was removed under vacuum. The residue was diluted with water, washed with ether. The aqueous layer was taken up in ethyl acetate, and partitioned with 10% citric acid. The organic layer was washed with water (1 x), brine (1 x), dried and concentrated to give the acid intermediate 4-(2-propyloxyethyl)-piperidine-l,2- carboxylic acid- 1-tert-butyl ester (61.1 mg, 97%>) as a syrup. [0734] 1H NMR (300 MHz, CDC13 ) δ 4.37-4.30 (m, 1), 3.61-3.50 (m, 1), 3.42 (t, J = 6.5, 2),
3.34 (t, J = 6.8, 2), 342-3.35 (m, 1), 2.06-1.95 (m,l), 1.88-1.70 (m, 3), 1.65-1.50 (m, 4), 1.43 (s,
9), 142-1.34 (m, 1), 0.88 (t, J = 7.5, 3).
[0735] A mixture of lincosamine 2b hydrochloride (R1 = R2 = Me; 55.8 mg, 0.19 mmol, 1 equiv) and DIEA (50 mg, 0.39 mmol, 2 equiv) in DMF (1 mL), was added to above acid intermediate (61.1 mg, 0.19 mmol,l equiv), followed by the addition of HBTU (80 mg, 0.21 mmol, 1.1 equiv). The reaction mixture was stirred at r.t. for 3 h. The reaction mixture was evaporated under high vacuum to dryness. The residue was diluted with ethyl acetate, washed with 1:1 10% citric acid/brine (lx), sat. aqueous sodium bicarbonate (lx), brine (lx), dried and concentrated. The residue was purified by chromatography to give the N-Boc lincosamide intermediate (60 mg, 56 %).
[0736] MS (ESPOS): 549.5 [M + H]+. MS (ESNEG): 547.2 [M - H] ".
[0737] To a solution of above N-Boc intermediate in DCM (9 mL) with methyl sulfide (0.2 mL) were added trifluoroacetic acid (3 mL) and water (0.2 mL). The reaction mixture was stirred at r.t. for 1 h. The solvent was removed under vacuum and co-evaporated with toluene twice. The residue was purified by chromatography to provide the final product (lower Rf isomer, 12 mg, 25%) as a white solid.
[0738] 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J = 5.7, 1), 4.20 (dd, J = 3.3, 9.9, 1), 4.11-
4.04 (m, 2), 3.83-3.76 (m, 2), 3.55-348 (m, 3), 345-3.37 (m, 1), 3.38 (t, J = 6.6, 2), 3.07-2.94
(m, 1), 2.29-2.12 (m, 2), 2.11 (s, 3), 1.98-1.83 (m, 2), 1.65-1.51 (m, 4), 145-1.33 (m, 2), 0.96-
0.86 (m, 9); MS (ESPOS): 449.4 [M + H]+; MS (ESNEG): 447.2 [M - H]\
Example 117
4-Cyclopropylmethosy-piperidine-2-carbosylic acid [2-methyl-l-(3,4,5-trihydro∑y- 6-ϊMethyϊsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
[0739] The title compound was made using the synthetic sequence found in method S starting from 4-hydroxypyridine-2-carboxylic acid, substituting cyclopropylmethylbromide as the alkylating agent.
[0740] Compound 15a (R9 =cyclopropylmethyl): 1H NMR (300 MHz, CD3OD) δ 8.41 (d,
J=4.8, 1), 7.65 (d, J=24, 1), 7.20 (m, 1), 4.09 (d, J=6.9, 2), 1.32 (m, 1), 0.66 (m, 2), 0.43 (m, 2);
MS (ESNEG): 192 [M - H]\
[0741] Compound 15b (R1=Me, R2=Me, R3=H, R9'=cyclopropylmethyl): 1H NMR (300
MHz, CD3OD) δ 8.41 (d, J=5.7, 1), 7.61 (d, J=2.7, 1), 7.07 (dd, J=2.7, 5.7, 1), 5.27 (d, J=54, 1),
4.87 (m, 2), 4.12 (dd, J=3.0, 10.2, 1), 3.98 (d, J=6.9, 1), 3.84 (d, J=3.0, 1), 3.56 (dd, J=3.3, 9.6, 1), 3.33 (m, 1), 2.85 (m, 1), 2.25 (m, 1), 2.11 (s, 3), 1.29 (m, 1), 0.98 (t, J=5.7, 6), 0.66 (m, 2), 0.39 (m, 2); MS (ESPOS): 427 [M + H]+.
[0742] Title compound (38 mg, 18%): 1H NMR (300 MHz, CD3OD) δ 5.24 (d, J=54, 1),
4.88 (dd, J=6.3, 9.6, 1), 4.04 (m, 2), 4.21 (d, J=2.7, 1), 3.50 (m, 3), 3.33 (m, 2), 2.78 (m, 1), 2.30 (m, 1), 2.16 (m, 1), 2.10 (s, 3) 1.38 (m, 2), 1.02 (m, 9), 0.53 (m, 2), 0.22 (m, 2); MS (ESPOS): 433 [M + Hf . f
Example 118
4-Fluoro-4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
$] The title compound was made using the synthetic sequence found in method Y (general scheme 21) starting from 4-oxo-piperidine- 1,2-dicarboxylic acid 1-tert-butyl ester 21a (m=2, P=H, P2=Boc), substituting butyl lithium as the R9 carbon nucleophile. [0744] Lincosamine 2b (R1=Me, R2=Me) was coupled to carbamate 21d (P2=Boc, m=2, R9 = n-butyl) as depicted in general coupling scheme 11 to provide intermediate Ila (R!=Me, R2=Me, R3=H, R9=butyl/fluoro, P1=H, P2=carboxylic acid-t-butyl ester, m=2) which was deprotected under acidic conditions to provide the title compound. [0745] MS (ESPOS): 437 [M + H]+. Examples 119-122
[0746] Examples 119-124 are prepared in an analogous manner as in Example 118 (Method
Y, scheme 21), substituting the appropriate R9 carbon nucleophile.
Example 119
4-Fluoro-4-ethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Example 120
4-Fluoro-4-(3-fluoropropyl)-piperidinβ-2-carboxylie acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Example 121
4-Fluoro-4-(3,3-difluoropropyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide
Example 122
4-Fluoro-4-(2,2-difluoroethoxymethyl)-piperidine-2-carboxylic acid [2-methyI-l-
[0747] The following Examples may be used to test compounds of this invention. Example A Susceptibility Testing
[0748] Compounds were tested following the microdilution method of NCCLS (National Committee for Clinical Laboratory Standards. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; Approved standard - fifth edition. NCCLS document M7-A5, NCCLS, Wayne, PA. 2000; National Committee for Clinical Laboratory Standards. Methods for antimicrobial susceptibility testing of anaerobic bacteria; Approved standard - fifth edition. NCCLS document Ml 1-A4, NCCLS, Wayne, PA. 2001). Assays were performed in sterile plastic 96-well microtiter trays with round bottom wells (Greiner).
Compound Preparation
[0749] Stock solutions of test compounds and control antibiotics are prepared at lOmg/mL in DMSO. Serial 2-fold dilutions of each drug are performed in a microtiter plate across each row using DMSO as solvent at 100- fold the desired final concentration. Wells in columns #1- 11 contain drug and column #12 was kept as a growth control for the organism with no drug. Each well in the mother plate is diluted with sterile deionized water, mixed, and volumes of 10 μL distributed to each well in the resulting assay plates.
Preparation of Inoculum
[0750] Stock cultures were prepared using the Microbank™ method (Pro-Lab Diagnostics) and stored at -80°C. To propagate aerobic strains, one bead was removed from the frozen vial and aseptically streaked onto Trypticase Soy Agar (Difco), Chocolate Agar (Remel) or Blood Agar (Remel) which were incubated at 35°C overnight. Anaerobes were cultivated in Brucella Agar (Remel) supplemented with hemin and vitamin K and incubated in anaerobiosis using an Anaerobic Jar (Mitsubishi) at 35°C for 24 to 48 h. Standardized inocula were prepared using the direct colony suspension method according to NCCLS guidelines (National Committee for Clinical Laboratory Standards. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; Approved standard - fifth edition. NCCLS document M7-A5, NCCLS, Wayne, PA. 2000; National Committee for Clinical Laboratory Standards. Methods for antimicrobial susceptibility testing of anaerobic bacteria; Approved standard - fifth edition. NCCLS document Ml 1-A4, NCCLS, Wayne, PA. 2001). Isolated colonies were selected from an 18-24 h agar plate and resuspended in 0.9%) sterile saline to match a 0.5 McFarland turbidity standard. The suspension was used within 15 min. of preparation. Streptococcus pneumoniae NSPN1001 Streptococcus pneumoniae ATCC 49619 Streptococcus pneumoniae NSPN3026 Streptococcus pneumoniae R6x Streptococcus pneumoniae NSPN4054 Streptococcus pneumoniae 488K Streptococcus pneumoniae NSPN4021 Streptococcus pneumoniae 9 Staphylococcus aureus VSAU1017 Staphylococcus aureus Smith Staphylococcus aureus NSAU1003 Staphylococcus aureus ATCC 25923 Staphylococcus aureus NSAU4020 Staphylococcus aureus!25 Staphylococcus aureus NSAU4048 Staphylococcus aureus 85-EPI Staphylococcus aureus NSAU4065 Staphylococcus aureus NSAU4065 Staphylococcus epidermidis NSEP1001 Staphylococcus epidermidis ATCC 12228 Enterococcus faecalis NEFL1003 Enterococcus faecalis ATCC 51299 Enterococcus faecium NEFA1005 Enterococcus faecium BM4147.1 Haemophilus infuenzae NHIΝ1003 Haemophilus infuenzae ATCC 49766 Haemophilus infuenzae VHIN1004 Haemophilus infuenzae ATCC 31517 Haemophilus infuenzae VHIN1005 acr Haemophilus infuenzae LS-2 Moraxella catarrhalis NMCA1001 Moraxella catarrhalis ATCC 25238 Escherichia coli VECO2096 Escherichia coli MG1655 Escherichia coli NEC02526 tolC Escherichia coli MG1655 tolC Bacteroides fragilis NBFR1001 Bacteroides fragilis ATCC 25285 Bacteroides thetaiotaomicron VBTH 1001 Bacteroides thetaiotaomicron ATCC #29741 Clostridium difficile NCDI1001 Clostridium difficile ATCC 9689
Preparation of Assay Plates for MICs Preparation of Assay Plates for MICs [0751] Media were prepared at 1.1 x concentration. Mueller-Hinton Broth MHB (Difco) supplemented with Ca++ and Mg++ as recommended by ΝCCLS, MHB supplemented with 5% horse lysed blood, HTM Broth (Remel), or Brucella broth (Remel) supplemented with hemin and vitamin K. For each organism, the standardized suspension was diluted into appropriate growth medium in a sterile reservoir. After mixing, wells in the drug-containing assay plates were inoculated with a volume of 90μl. Thus, for each MIC determination, each well contains a final volume of 100 μL with an inoculum size of approximately 5 * 105 cfu/mL and no more than 1% DMSO.
Interpretation of MIC
[0752] The completed microtiter plates were incubated 16-20 h at 35°C in ambient air for aerobes, and at 35 °C for 46-48 h or in an anaerobe jar (Mitsubishi) for anaerobes. Optical density of each well was determined at 600 nm using a VersaMax Microplate reader (Molecular Devices, Sunnyvale, CA). The MIC was defined as the lowest drug concentration causing complete suppression of visible bacterial growth.
Example B Efficacy in Murine S. aureus Septicemia
[0753] Efficacy studies were performed in an S. aureus murine septicemia model according to models published elsewhere (Goldstein, B. P., G. Candiani, T. M. Arain, G. Romano, I. Ciciliato, M. Berti, M. Abbondi, R. Scotti, M. Mainini, F. Ripamonti, and et al. 1995. Antimicrobial activity of MDL 63,246, a new semisynthetic glycopeptide antibiotic Antimicrob Agents Chemother. 39:1580-1588.; Misiek, M., T. A. Pursiano, F. Leitner, and K. E. Price 1973. Microbiological properties of anew cephalosporin, BL-S 339: 7-(phenylacetimidoyl- aminoacetamido)-3 -(2-methyl-l ,3,4-thiadiazol-5-ylthio methyl)ceph-3-em-4-carboxylic acid Antimicrob Agents Chemother. 3:40-48).
Compound Preparation
[0754] Compounds were dissolved in 2% Tween 80 for oral dosing or 0.9% NaCl solution for intravenous dosing. Compounds were administered at 1 hour after bacterial inoculation. Vancomycin or ampicillin were used as controls.
Efficacy model
[0755] Male or female ICR mice weighing 22±2 g from MDS Pharma Services were used for the evaluation. Food and water was given ad libitum. Groups of 6 mice weighing 22 ± g were used for the experiment. Mice were inoculated intraperitoneally with Staphylococcus aureus Smith at 4 104 CFU in 0.5 mL of Brain Heart Infusion Broth (Difco) containing 5% mucin (Sigma). Mortality was recorded once daily for 7 days following bacterial inoculation. [0756] While the invention has been described and illustrated herein by references to various specific material, procedures and examples, it is understood that the invention is not restricted to the particular material combinations of material, and procedures selected for that purpose. Numerous variations of such details can be implied as will be appreciated by those skilled in the art. Example C In vivo animal model
[0757] In vivo activity of various compounds of the subject invention was evaluated in a standard Staphylococcus aureus septicemia model (MDS Pha-rma Services, Bothell, WA). [0758] Male ICR-derived mice (ICR is a strain of out-bred mice) provided by MDS Pharma Services animal breeding center were inoculated intraperitonealy with LDgo-ϊoo of Staphylococcus aureus (Smith; ATCC19636) in 0.5mL BHI broth containing 5% mucin (Sigma). Compounds were formulated in 2% Tween 80 (Sigma) and single doses were administered orally one hour after bacterial inoculation. Mortality was monitored daily for seven days.
[0759] In previous studies, the oral ED5o (i.e., concentration that protected 50% of the mice) was determined to be 19.9 mg/kg for clindamycin, a commercially available lincosamide (Sigma). To screen the compounds of this invention that were tested, compounds were administered at 10 mg/kg to a group of eight ICR mice and the number of survivors at that concentration was compared to clindamycin. Results are presented in the table below. Compound Tested No. of surviving mice (Indicated by at 10 mg/kg Example No.) Clindamycin 6 17 1 34 2 35 2 36 0 100 7 101 7
It is assumed that when compound from example 36 is tested at a higher dose, the number of surviving mice would increase.

Claims

CLAIMS WHAT IS CLAIMED IS: 1. A compound of formula (I) :
wherein: R1 is alkyl; R2 and R3 are independently H, alkyl, hydroxy, fluoro, or cyanoalkyl or one 9 ^ 7 9 ofR and R is =NOR and the other is absent, or one of R and R is =CH2 and the 9 ' ' "X 9 ^ other is absent, with the provisos that both R and R are not H; when one of R and R ' ' 9 ^ is fluoro, the other is not hydrogen or hydroxy; and when one of R and R is hydroxy, the other is not fluoro, hydrogen, or hydroxy; R is selected from the group consisting of H, alkyl, hydroxyalkyl, -C(0)0-alkylene-cycloalkyl, -C(0)0-alkylene-substituted cycloalkyl, -C(0)0-alkyl, -C(0)0-substituted alkyl, -C(Q)0-aryl, -C(0)0-substiτuted aryl, -C(0)0-heteroaryl, -C(0)0-substituted heteroaryl, -[C(0)0]p-alkylene-heterocycle, -[C(0)0]p-alkylene- substituted heterocycle, wherein p is 0 or 1; R7 is H or alkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH2)„-OH, -(CH2)n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 0, 1, 2 or 3; and prodrugs, tautomers or pharmaceutically acceptable salts thereof; with the proviso that the compound of formula I has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile.
2. A compound of formula (II)
wherein: R1 is alkyl; R2 and R3 are independently H, alkyl, or cyanoalkyl, with the proviso that both R2 and R3 are not H; R6 is H, alkyl, or hydroxyalkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH )n-OH, - (CH )n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R and R5 are H or alkyl; and m is 1 or 2; and prodrugs and pharmaceutically acceptable salts thereof; with the proviso that the compound of formula II has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile.
3. A compound of formula (III) :
(III)
wherein: R1 is alkyl; 9 "X R and R are fluoro; R is H, alkyl, or hydroxyalkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH2)n-OH, - (CH2)n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 1 or 2; and prodrugs and pharmaceutically acceptable salts thereof, with the proviso that the compound of formula III has a minimum inhibition concenfration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile.
4. A compound of formula (IN):
wherein: R1 is alkyl; R2 and R3 are independently H, or alkyl, hydroxy, fluoro, or cyanoalkyl or one of R2 and R3 is =ΝOR7 and the other is absent, or one of R2 and R3 is =CH2 and the other is absent, with the provisos that both R2 and R3 are not H; when one of R2 and R3 is fluoro, the other is not hydrogen or hydroxy; and when one of R2 and R3 is hydroxy, the other is not fluoro, hydrogen, or hydroxy; R6 is selected from the group consisting of -C(0)0-alkylene-cycloalkyl, -C(0)0-alkylene-substituted cycloalkyl, -C(0)0-alkyl, -C(0)0-substituted alkyl, -C(0)0-aryl, -C(0)0-substixuted aryl, -C(0)0-heteroaryl, -C(0)0-substituted heteroaryl, -[C(0)0]p-alkylene-heterocycle, -[C(0)0]p-alkylene-substituted heterocycle, wherein p is 0 or 1 with the proviso that -C(0)0-substituted alkyl does not include the following:
R7 is H or alkyl; R9, which can be singly or multiply substituted in the ring on the same or different carbons, is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxyalkoxy, cycloalkyl, substituted cycloalkyl, alkoxyalkoxy, substituted oxygen, substituted nitrogen, halogen, phenyl, substituted phenyl, -(CH2)n-OH, -(CH2)n-NR4R5, -alkylene-Ra where Ra is selected from monofluorophenyl and monochlorophenyl, and branched chain isomers thereof wherein n is an integer of from 1 to 8 inclusive and R4 and R5 are H or alkyl; and m is 1 or 2; and prodrugs, tautomers or pharmaceutically acceptable salts thereof; with the proviso that the compound of formula I has a minimum inhibition concentration of 32 μg/mL or less against at least one of the organisms selected from the group consisting of Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Bacteroides fragilis, Bacteroides thetaiotaomicron, and Clostridium difficile.
5. A compound of claim 1, wherein m is 1 or 2.
6. A compound of claim 1 , wherein R1 is methyl.
7. A compound of claim 1, wherein R6 is H, alkyl, or hydroxyalkyl.
8. A compound of claim 1, wherein each R9 is independently H, alkyl, substituted alkyl, alkoxy, substituted alkoxy, or cycloalkyl.
9. A compound selected from the group consisting of: 4-(2-methoxyethoxy)-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Butyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3, 4,5-frihy droxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4,4-Difluoro-pentyl)-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Fluoro-propyl)-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-frihy droxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-propyl-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Fluoroethoxy)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-ρropyl]-amide; 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclobutylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6-methylsulfanyl- tefrahydro-pyran-2-yl)-propyl] -amide; 3 -Cyclopropylmethyl-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-Propyl-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 3-Butyl-l-(2-hydroxy-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl] -amide; 3-Pentyl-azetidine-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Methyl-butyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Cyclobutyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(2-Cyclobutyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(2-Cyclopropyl-ethyl)-azetidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3-(3-Cyclopropyl-propyl)-azetidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 3 -Butyl- 1 -methyl-azetidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethyl-pyrrolidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclobutyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Cyclopropyl-ethyl)-pyrrolidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 5-Propyl-azepane-2-carboxylic acid [2-methyl-l -(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Cyclopentyl-propyl)-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-trihydroxy-6- methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Methoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Ethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-frihydroxy-6- meihylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Propoxy-propyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Cyclopropylmethoxy-propyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5- trihydroxy-6-methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 4- [3 -(2-Fluoro-ethoxy)-propyl] -piperidine-2-carboxylic acid [2-methyl- 1 -(3 , 4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-[3-(3-Fluoro-propoxy)-propyl]-piperidine-2-carboxylic acid [2-methyl-l -(3,4,5- trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(4-Methoxy-butyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Propoxymethyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(3-Fluoro-propoxymethyl)-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-frihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclohexylmethyl-piperidine-2-carboxylic acid [2-methyl-l -(3 ,4,5-frihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-(2-Propyloxyethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Cyclopropylmethoxy-piperidme-2-carboxylic acid [2-methyl- 1 -(3 ,4, 5 -trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-butyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-ethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3-fluoropropyl)-piperidine-2-carboxylic acid [2-methyl-l -(3, 4,5-trihydroxy- 6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(3 ,3 -difluoropropyl)-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 - frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 4-Fluoro-4-(2,2-difluoroethoxymethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; and prodrugs, tautomers and pharmaceutically acceptable salts thereof.
10. A compound selected from the group consisting of: Phosphoric acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine-2-carbonyl)- amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl) ester; Succinic acid mono-(4,5 -dihydroxy-6- { 2-methyl- 1 - [(4-propyl-piperidine-2-carbonyl)- amino]-propyl}-2-methylsulfanyl-tefrahydro-pyran-3-yl) ester; N-(2-Morpholin-4-yl-ethyl)-succinamic acid 4,5-dihydroxy-6-{2-methyl-l-[(4-propyl- piperidine-2-carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; Dimethylamino-acetic acid 4,5-dihydroxy-6- {2-methyl-l -[(4-propyl-piperidine-2- carbonyl)-amino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; 1 -(5-Methyl-2-oxo-[ 1 ,3]dioxol-4-ylmethyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid 5-methyl-2-oxo-[l,3]dioxol-4-ylmethyl ester; Hexadecanoic acid 4,5-dihydroxy-6-{2-methyl-l -[(4-propyl-piperidine-2-carbonyl)- -mιino]-propyl}-2-methylsulfanyl-tetrahydro-pyran-3-yl ester; 1 -(1 -Methyl-3 -oxo-but- 1 -enyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1-carboxylic acid 1-acetoxy-ethyl ester; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahy hO-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid 2-amino-3-methyl-pentanoyloxymethyl ester; 2- [2-Methyl- 1 -(3 ,4, 5 -trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine- 1-carboxylic acid piperidine-4-carbonyloxymethyl ester; l-(Propionylamino-methyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5- frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; N-{2-[2-Methyl-l-(3,4,5-1rihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piρeridin- 1 -ylmethyl } -nicotinamide; 1 -(2-Amino-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5- trihydroxy-6-methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -phenyl-propionyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulf-myl-tetr--ώydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -methyl-pentanoyl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 - (3,4,5-trihydroxy-6-methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide; 1 -(2- Amino-3 -methyl-butyryl)-4-propyl-piperidine-2-carboxylic acid [2-methyl- 1 -(3 ,4,5 - frihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 1 -( 1 -Methyl- 1 ,4-dihydro-pyridine-3 -carbonyl)-4-propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide; 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl] -4-propyl-piperidine- 1 -carboxylic acid 1 -methyl- 1 ,2,3 ,6-tetrahydro-pyridin-4- yl ester; and tautomers and pharmaceutically acceptable salts thereof.
11. A compound according to claim 9, wherein the compound is: 4-Fluoro-4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide.
12. A compound according to claim 9, wherein the compound is: 4-(2-Cyclopropyl-ethyl)-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide.
13. A compound according to claim 9, wherein the compound is: 4-Cyclopropylmethyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide.
14. A compound according to claim 10, wherein the compound is: 5-Propyl-azepane-2-carboxylic acid [2-methyl- 1 -(3 ,4,5-trihydroxy-6-methylsulfanyl- tetrahydro-pyran-2-yl)-propyl]-amide.
15. A compound according to claim 10, wherein the compound is: Phosphoric acid mono-(4,5-dihydroxy-6-{2-methyl-l-[(4-propyl-piperidine-2-carbonyl)- amino] -propyl } -2-methylsulfanyl-tetrahydro-pyran-3 -yl) ester.
16. A compound according to claim 10, wherein the compound is: l-(5-Methyl-2-oxo-[l ,3]dioxol-4-ylmethyl)-4 -propyl-piperidine-2-carboxylic acid [2- methyl-l-(3,4,5-frihydroxy-6-methylsulfanyl-tefrahydro-pyran-2-yl)-propyl]-amide.
17. A compound according to claim 10, wherein the compound is: 2-[2-Methyl-l-(3,4,5-trihydroxy-6-methylsulfanyl-tetrahydro-pyran-2-yl)- propylcarbamoyl]-4-propyl-piperidine-l-carboxylic acid 5-methyl-2-oxo-[l,3]dioxol-4-ylmethyl ester.
18. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim 1.
19. A method for the treatment of a microbial infection in a mammal comprising administering to the mammal a therapeutically effective amount of a compound of claim 1.
20. The method according to claim 19, wherein the compound is administered to the mammal orally, parenterally, transdermally, topically, rectally, or intranasaUy in a pharmaceutical composition.
21. The method according to claim 19, wherein the compound is administered in an amount of from about 0.1 to about 100 mg/kg of body weight/day.
22. The compound 4-propyl-piperidine-2-carboxylic acid [2-methyl-l-(3,4,5-trihydroxy-6- methylsulfanyl-tetrahydro-pyran-2-yl)-propyl]-amide.
EP04785949A 2003-06-17 2004-06-17 Lincomycin derivatives possessing antibacterial activity Withdrawn EP1654268A2 (en)

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US10/777,455 US7199105B2 (en) 2002-08-15 2004-02-11 Lincomycin derivatives possessing antibacterial activity
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US7256177B2 (en) 2003-06-17 2007-08-14 Vicuron Pharmaceuticals, Inc. Lincomycin derivatives possessing antibacterial activity
US7199106B2 (en) 2003-06-17 2007-04-03 Vicuron Pharmaceuticals, Inc. Lincomycin derivatives possessing antimicrobial activity
US7361743B2 (en) * 2004-02-11 2008-04-22 Pfizer Inc Lincomycin derivatives possessing antibacterial activity
WO2008146919A1 (en) * 2007-05-31 2008-12-04 Meiji Seika Kaisha, Ltd. Lincosamide derivative, and antibacterial agent comprising the same as active ingredient
EP2151447A4 (en) * 2007-05-31 2012-09-05 Meiji Seika Kaisha Lincomycin derivatives and antibacterial agents containing the same as the active ingredient
CN109627299B (en) * 2018-11-03 2022-02-18 北京工商大学 Bacteriocin Gr17 with broad-spectrum antibacterial activity and application thereof

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