U.S. patent application number 15/685846 was filed with the patent office on 2017-12-14 for boron-containing small molecules.
The applicant listed for this patent is Anacor Pharmaceuticals, Inc.. Invention is credited to Michael Richard Kevin Alley, Charles Ding, Eric Easom, Vincent S. Hernandez, Xianfeng Li, Jacob J. Plattner, Fernando Rock, Suoming Zhang, Ding Zhou.
Application Number | 20170355719 15/685846 |
Document ID | / |
Family ID | 44653589 |
Filed Date | 2017-12-14 |
United States Patent
Application |
20170355719 |
Kind Code |
A1 |
Hernandez; Vincent S. ; et
al. |
December 14, 2017 |
BORON-CONTAINING SMALL MOLECULES
Abstract
This invention relates to, among other items, benzoxaborole
compounds and their use for treating bacterial infections.
Inventors: |
Hernandez; Vincent S.;
(Watsonville, CA) ; Ding; Charles; (San Mateo,
CA) ; Plattner; Jacob J.; (Berkeley, CA) ;
Alley; Michael Richard Kevin; (Santa Clara, CA) ;
Rock; Fernando; (Los Altos, CA) ; Zhang; Suoming;
(San Mateo, CA) ; Easom; Eric; (Mountain View,
CA) ; Li; Xianfeng; (Cupertino, CA) ; Zhou;
Ding; (Shanghai, CN) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Anacor Pharmaceuticals, Inc. |
Palo Alto |
CA |
US |
|
|
Family ID: |
44653589 |
Appl. No.: |
15/685846 |
Filed: |
August 24, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14221637 |
Mar 21, 2014 |
9751898 |
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15685846 |
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13227444 |
Sep 7, 2011 |
8703742 |
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14221637 |
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61380596 |
Sep 7, 2010 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07F 5/025 20130101;
A61P 31/04 20180101; A61P 43/00 20180101; A61P 31/00 20180101; A61P
31/06 20180101 |
International
Class: |
C07F 5/02 20060101
C07F005/02 |
Claims
1. A compound having a structure which is: ##STR00117## wherein
R.sup.3 is --CH.sub.2NH.sub.2; R.sup.4 is selected from the group
consisting of halogen, methyl, ethyl, propyl, isopropyl, butyl,
isobutyl, sec-butyl, methoxy, ethoxy, propoxy, isopropoxy, butoxy,
isobutoxy, and sec-butoxy; Y is O; and R.sup.5 is ##STR00118##
wherein a is 2, 3 or 4; R.sup.10 and R.sup.11 is H; and R.sup.12 is
selected from the group consisting of H, OH, NH.sub.2, methyl,
ethyl, --NHS(O).sub.2CH.sub.3, cyano, --NHC(O)CH.sub.3,
--NHC(O)NHCH.sub.2CH.sub.3, --C(O)NH.sub.2, --C(O)OH,
4-(methoxy)phenyl, benzyl, benzoxy, --NHC(O)OCH.sub.2Ph,
--C(O)NHCH.sub.2CH.sub.2OH and --C(NH.sub.2)(NH); or a salt,
hydrate or solvate thereof.
2. The compound of claim 1 or a salt, hydrate or solvate thereof,
having a structure which is: ##STR00119## wherein C* is a carbon
atom stereocenter which has a configuration which is (R) or
(S).
3. The compound of claim 2 or a salt, hydrate or solvate thereof,
wherein the C* stereocenter is in a (S) configuration.
4. The compound of claim 1 or a salt, hydrate or solvate thereof,
wherein R.sup.4 is selected from the group consisting of fluorine,
chlorine, bromine, and iodine.
5. The compound of claim 1 or a salt, hydrate or solvate thereof,
wherein R.sup.4 is selected from the group consisting of methyl,
ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl.
6. A composition comprising: a) a first stereoisomer of the
compound of claim 3 or a salt, hydrate or solvate thereof, b) at
least one additional stereoisomer of the first stereoisomer;
wherein the first stereoisomer is present in an enantiomeric excess
of at least 80% relative to said at least one additional
stereoisomer.
7. A combination comprising the compound of claim 1, or a
pharmaceutically acceptable salt thereof, together with at least
one other therapeutically active agent.
8. A pharmaceutical formulation comprising: a) the compound of
claim 1, or a pharmaceutically acceptable salt thereof, and b) a
pharmaceutically acceptable excipient.
9. A method of inhibiting the editing domain of a t-RNA synthetase,
comprising: contacting the synthetase with an effective amount of a
compound of claim 1, or a pharmaceutically-acceptable salt thereof,
thereby inhibiting the synthetase.
10. The method of claim 9, wherein the t-RNA synthetase is
LeuRS.
11. A method of killing and/or preventing the growth of a
microorganism, comprising: contacting the microorganism with an
effective amount of the compound of claim 1, thereby killing and/or
preventing the growth of the microorganism.
12. The method of claim 11, wherein the microorganism is
Mycobacterium tuberculosis.
13. A method of treating and/or preventing a disease in an animal,
comprising: administering to the animal a therapeutically effective
amount of the compound of claim 1, or a pharmaceutically-acceptable
salt thereof, thereby treating and/or preventing the disease.
14. The method of claim 13, wherein the disease is tuberculosis
15. The method of claim 13, wherein the animal is a human.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a divisional application, which is a
continuation of U.S. application Ser. No. 14/221,637, filed Mar.
21, 2014, now U.S. Pat. No. 9,751,898, which is a continuation of
U.S. application Ser. No. 13/227,444, filed Sep. 7, 2011, now U.S.
Pat. No. 8,703,742, and claims the benefit of U.S. Provisional Pat.
App. No. 61/380,596, filed Sep. 7, 2010, which are incorporated by
reference in its entirety for all purposes.
REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY
[0002] The sequence listing contained in the file named
"064507-5066-US_ST25.txt", created on Jan. 19, 2012 and having a
size of 427 bytes, has been submitted electronically herewith via
EFS-Web, and the contents of the txt file are hereby incorporated
by reference in their entirety.
BACKGROUND OF THE INVENTION
[0003] The global rise of bacteria and other microorganisms
resistant to antibiotics and antimicrobials in general, poses a
major threat. Deployment of massive quantities of antimicrobial
agents into the ecosphere during the past 60 years has introduced a
powerful selective pressure for the emergence and spread of
antimicrobial-resistant pathogens. Thus, there is a need to
discover new broad spectrum antimicrobials, such as antibiotics,
useful in combating microorganisms, especially those with
multidrug-resistance.
[0004] Boron-containing molecules, such as
1-hydroxy-1,3-dihydro-benzo[c][1,2]oxaborole (also sometimes known
as 1-hydroxy-benzo[c][1,2]oxaborole or oxaboroles or cyclic boronic
esters), useful as antimicrobials have been described previously,
such as in U.S. patent application Ser. Nos. 12/142,692; 11/505,591
and 11/357,687. Generally speaking, a
1-hydroxy-1,3-dihydro-benzo[c][1,2]oxaborole has the following
structure and substituent numbering system:
##STR00001##
Surprisingly, it has now been discovered that certain classes of
1-hydroxy-1,3-dihydro-benzo[c][1,2]oxaboroles are effective
antibacterials. This, and other uses of these
1-hydroxy-1,3-dihydro-benzo[c][1,2]oxaboroles are described
herein.
SUMMARY OF THE INVENTION
[0005] In a first aspect, the invention provides a compound having
a structure according to the formula:
##STR00002##
wherein R.sup.3 is substituted or unsubstituted nitroalkyl or
substituted or unsubstituted aminoalkyl; R.sup.4 is selected from
the group consisting of halogen, unsubstituted alkyl and
unsubstituted phenyl; Y is O or S; R.sup.5 is selected from the
group consisting of substituted or unsubstituted alkyl and
substituted or unsubstituted heteroalkyl, or a salt, hydrate or
solvate thereof.
[0006] In a second aspect, the invention provides a combination
comprising: a) a compound of the invention, or a pharmaceutically
acceptable salt thereof, and b) a therapeutically active agent.
[0007] In a third aspect, the invention provides a pharmaceutical
formulation comprising: a) a compound of the invention, or a
pharmaceutically acceptable salt thereof, and b) a pharmaceutically
acceptable excipient.
[0008] In a fourth aspect, the invention provides a method of
killing or inhibiting the growth of a bacteria, said method
comprising: contacting said bacteria with an effective amount of a
compound of the invention or a combination of the invention, or a
pharmaceutically acceptable salt thereof, thereby killing or
inhibiting the growth of the bacteria.
[0009] In a fifth aspect, the invention provides a method of
treating a bacterial infection comprising: administering to an
animal suffering from said infection an effective amount of a
compound of the invention, or a pharmaceutically-acceptable salt
thereof, thereby treating the bacterial infection.
[0010] In a sixth aspect, the invention provides a method of
inhibiting the editing domain of a t-RNA synthetase, comprising:
contacting the synthetase with an effective amount of a compound of
the invention, or a pharmaceutically-acceptable salt thereof,
thereby inhibiting the synthetase.
BRIEF DESCRIPTION OF THE DRAWINGS
[0011] FIG. 1 displays biological data for exemplary compounds of
the invention.
DETAILED DESCRIPTION OF THE INVENTION
I. Definitions and Abbreviations
[0012] As used herein, the singular forms "a," "an", and "the"
include plural references unless the context clearly dictates
otherwise. For example, reference to "an active agent" includes a
single active agent as well as two or more different active agents
in combination. It is to be understood that present teaching is not
limited to the specific dosage forms, carriers, or the like,
disclosed herein and as such may vary.
[0013] The abbreviations used herein generally have their
conventional meaning within the chemical and biological arts.
[0014] The following abbreviations have been used: Ac is acetyl;
AcOH is acetic acid; ACTBr is cetyltrimethylammonium bromide; AIBN
is azobisisobutyronitrile or 2,2 azobisisobutyronitrile; aq. is
aqueous; Ar is aryl; B.sub.2pin.sub.2 is bis(pinacolato)diboron; Bn
is, in general, benzyl [see Cbz for one example of an exception];
(BnS).sub.2 is benzyl disulfide; BnSH is benzyl thiol or benzyl
mercaptan; BnBr is benzyl bromide; Boc is tert-butoxy carbonyl;
Boc.sub.2O is di-tert-butyl dicarbonate; Bz is, in general,
benzoyl; BzOOH is benzoyl peroxide; Cbz or Z is benzyloxycarbonyl
or carboxybenzyl; Cs.sub.2CO.sub.3 is cesium carbonate; CSA is
camphor sulfonic acid; CTAB is cetyltrimethylammonium bromide; Cy
is cyclohexyl; DABCO is 1,4-diazabicyclo[2.2.2]octane; DCM is
dichloromethane or methylene chloride; DHP is dihydropyran; DIAD is
diisopropyl azodicarboxylate; DIEA or DIPEA is
N,N-diisopropylethylamine; DMAP is 4-(dimethylamino)pyridine; DME
is 1,2-dimethoxyethane; DMF is N,N-dimethylformamide; DMSO is
dimethylsulfoxide; equiv or eq. is equivalent; EtOAc is ethyl
acetate; EtOH is ethanol; Et.sub.2O is diethyl ether; EDCI is
N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride; ELS
is evaporative light scattering; equiv or eq is equivalent; h is
hours; HATU is
O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate; HOBt is N-hydroxybenzotriazole; HCl is
hydrochloric acid; HPLC is high pressure liquid chromatography;
ISCO Companion is automated flash chromatography equipment with
fraction analysis by UV absorption available from Presearch; KOAc
or AcOK is potassium acetate; K.sub.2CO.sub.3 is potassium
carbonate; LiAlH.sub.4 or LAH is lithium aluminum hydride; LDA is
lithium diisopropylamide; LHMDS is lithium bis(trimethylsilyl)
amide; KHMDS is potassium bis(trimethylsilyl) amide; LiOH is
lithium hydroxide; m-CPBA is 3-chloroperoxybenzoic acid; MeCN or
ACN is methyl cyanide or cyanomethane or ethanenitrile or
acetonitrile which are all names for the same compound; MeOH is
methanol; MgSO.sub.4 is magnesium sulfate; mins or min is minutes;
Mp or MP is melting point; NaCNBH.sub.3 is sodium cyanoborohydride;
NaOH is sodium hydroxide; Na.sub.2SO.sub.4 is sodium sulfate; NBS
is N-bromosuccinimide; NH.sub.4Cl is ammonium chloride; NIS is
N-iodosuccinimide; N.sub.2 is nitrogen; NMM is N-methylmorpholine;
n-BuLi is n-butyllithium; overnight is O/N; PdCl.sub.2(pddf) is
1,1'-Bis(diphenylphosphino) ferrocene]dichloropalladium(II); Pd/C
is the catalyst known as palladium on carbon; Pd.sub.2(dba).sub.3
is an organometallic catalyst known as tris(dibenzylideneacetone)
dipalladium(0); Ra Ni or Raney Ni is Raney nickel; Ph is phenyl;
PMB is p-methoxybenzyl; PrOH is 1-propanol; iPrOH is 2-propanol;
POCl.sub.3 is phosphorus chloride oxide; PTSA is para-toluene
sulfonic acid; Pyr. or Pyr or Py as used herein means pyridine; RT
or rt or r.t. is room temperature; sat. is saturated; Si-amine or
Si--NH.sub.2 is amino-functionalized silica, available from
SiliCycle; Si-pyr is pyridyl-functionalized silica, available from
SiliCycle; TEA or Et.sub.3N is triethylamine; TFA is
trifluoroacetic acid; Tf.sub.2O is trifluoromethanesulfonic
anhydride; THF is tetrahydrofuran; TFAA is trifluoroacetic
anhydride; THP is tetrahydropyranyl; TMSI is trimethylsilyl iodide;
H.sub.2O is water; diNO.sub.2PhSO.sub.2Cl is dinitrophenyl sulfonyl
chloride; 3-F-4-NO.sub.2-PhSO.sub.2Cl is
3-fluoro-4-nitrophenylsulfonyl chloride;
2-MeO-4-NO.sub.2-PhSO.sub.2Cl is 2-methoxy-4-nitrophenylsulfonyl
chloride; and (EtO).sub.2POCH.sub.2COOEt is a triethylester of
phosphonoacetic acid known as triethyl phosphonoacetate.
[0015] "Compound of the invention," as used herein refers to the
compounds discussed herein, salts (e.g. pharmaceutically acceptable
salts), prodrugs, solvates and hydrates of these compounds.
[0016] The term "poly" as used herein means at least 2. For
example, a polyvalent metal ion is a metal ion having a valency of
at least 2.
[0017] "Moiety" refers to a radical of a molecule that is attached
to the remainder of the molecule.
[0018] The symbol , whether utilized as a bond or displayed
perpendicular to a bond, indicates the point at which the displayed
moiety is attached to the remainder of the molecule.
[0019] The term "alkyl," by itself or as part of another
substituent, means, unless otherwise stated, a straight or branched
chain, or cyclic hydrocarbon radical, or combination thereof, which
may be fully saturated, mono- or polyunsaturated and can include
di- and multivalent radicals, having the number of carbon atoms
designated (i.e. C.sub.1-C.sub.10 means one to ten carbons). In
some embodiments, the term "alkyl" means a straight or branched
chain, or combinations thereof, which may be fully saturated, mono-
or polyunsaturated and can include di- and multivalent radicals.
Examples of saturated hydrocarbon radicals include, but are not
limited to, groups such as methyl, ethyl, n-propyl, isopropyl,
n-butyl, t-butyl, isobutyl, sec-butyl, cyclohexyl,
(cyclohexyl)methyl, cyclopropylmethyl, homologs and isomers of, for
example, n-pentyl, n-hexyl, n-heptyl, n-octyl, and the like. An
unsaturated alkyl group is one having one or more double bonds or
triple bonds. Examples of unsaturated alkyl groups include, but are
not limited to, vinyl, 2-propenyl, crotyl, 2-isopentenyl,
2-(butadienyl), 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1-
and 3-propynyl, 3-butynyl, and the higher homologs and isomers.
[0020] The term "alkylene" by itself or as part of another
substituent means a divalent radical derived from an alkane, as
exemplified, but not limited, by
--CH.sub.2CH.sub.2CH.sub.2CH.sub.2--. Typically, an alkyl (or
alkylene) group will have from 1 to 24 carbon atoms, with those
groups having 10 or fewer carbon atoms being preferred in the
invention. A "lower alkyl" or "lower alkylene" is a shorter chain
alkyl or alkylene group, generally having eight or fewer carbon
atoms.
[0021] The term "alkenylene" by itself or as part of another
substituent means a divalent radical derived from an alkene.
[0022] The term "cycloalkylene" by itself or as part of another
substituent means a divalent radical derived from a cycloalkyl.
[0023] The term "heteroalkylene" by itself or as part of another
substituent means a divalent radical derived from an
heteroalkane.
[0024] The term "heterocycloalkylene" by itself or as part of
another substituent means a divalent radical derived from an
heterocycloalkane.
[0025] The term "arylene" by itself or as part of another
substituent means a divalent radical derived from an aryl.
[0026] The term "heteroarylene" by itself or as part of another
substituent means a divalent radical derived from heteroaryl.
[0027] The terms "alkoxy," "alkylamino" and "alkylthio" (or
thioalkoxy) are used in their conventional sense, and refer to
those alkyl groups attached to the remainder of the molecule via an
oxygen atom, an amino group, or a sulfur atom, respectively.
[0028] The term "heteroalkyl," by itself or in combination with
another term, means, unless otherwise stated, a stable straight or
branched chain, or cyclic hydrocarbon radical, or combinations
thereof, consisting of the stated number of carbon atoms and at
least one heteroatom. In some embodiments, the term "heteroalkyl,"
by itself or in combination with another term, means a stable
straight or branched chain, or combinations thereof, consisting of
the stated number of carbon atoms and at least one heteroatom. In
an exemplary embodiment, the heteroatoms can be selected from the
group consisting of B, O, N and S, and wherein the nitrogen and
sulfur atoms may optionally be oxidized and the nitrogen heteroatom
may optionally be quaternized. The heteroatom(s) B, O, N and S may
be placed at any interior position of the heteroalkyl group or at
the position at which the alkyl group is attached to the remainder
of the molecule. Examples include, but are not limited to,
--CH.sub.2--CH.sub.2--O--CH.sub.3,
--CH.sub.2--CH.sub.2--NH--CH.sub.3,
--CH.sub.2--CH.sub.2--N(CH.sub.3)--CH.sub.3,
--CH.sub.2--S--CH.sub.2--CH.sub.3, --CH.sub.2--CH.sub.2,
--S(O)--CH.sub.3, --CH.sub.2--CH.sub.2--S(O).sub.2--CH.sub.3,
--CH.dbd.CH--O--CH.sub.3, --CH.sub.2--CH.dbd.N--OCH.sub.3, and
--CH.dbd.CH--N(CH.sub.3)--CH.sub.3. Up to two heteroatoms may be
consecutive, such as, for example, --CH.sub.2--NH--OCH.sub.3.
Similarly, the term "heteroalkylene" by itself or as part of
another substituent means a divalent radical derived from
heteroalkyl, as exemplified, but not limited by,
--CH.sub.2--CH.sub.2--S--CH.sub.2--CH.sub.2-- and
--CH.sub.2--S--CH.sub.2--CH.sub.2--NH--CH.sub.2--. For
heteroalkylene groups, heteroatoms can also occupy either or both
of the chain termini (e.g., alkyleneoxy, alkylenedioxy,
alkyleneamino, alkylenediamino, and the like). Still further, for
alkylene and heteroalkylene linking groups, no orientation of the
linking group is implied by the direction in which the formula of
the linking group is written. For example, the formula
--C(O).sub.2R'-- represents both --C(O).sub.2R'-- and
--R'C(O).sub.2--.
[0029] The terms "cycloalkyl" and "heterocycloalkyl", by themselves
or in combination with other terms, represent, unless otherwise
stated, cyclic versions of "alkyl" and "heteroalkyl", respectively.
Additionally, for heterocycloalkyl, a heteroatom can occupy the
position at which the heterocycle is attached to the remainder of
the molecule. Examples of cycloalkyl include, but are not limited
to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl,
cycloheptyl, and the like. Examples of heterocycloalkyl include,
but are not limited to, 1-(1,2,5,6-tetrahydropyridyl),
1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl,
3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl,
tetrahydrothien-2-yl, tetrahydrothien-3-yl, 1-piperazinyl,
2-piperazinyl, and the like.
[0030] The terms "halo" or "halogen," by themselves or as part of
another substituent, mean, unless otherwise stated, a fluorine,
chlorine, bromine, or iodine atom. Additionally, terms such as
"haloalkyl," are meant to include monohaloalkyl and polyhaloalkyl.
For example, the term "halo(C.sub.1-C.sub.4)alkyl" is mean to
include, but not be limited to, trifluoromethyl,
2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the
like.
[0031] The term "aryl" means, unless otherwise stated, a
polyunsaturated, aromatic, substituent that can be a single ring or
multiple rings (preferably from 1 or 2 or 3 rings), which are fused
together or linked covalently. The term "heteroaryl" refers to aryl
groups (or rings) that contain from one to four heteroatoms. In an
exemplary embodiment, the heteroatom is selected from B, N, O, and
S, wherein the nitrogen and sulfur atoms are optionally oxidized,
and the nitrogen atom(s) are optionally quaternized. A heteroaryl
group can be attached to the remainder of the molecule through a
heteroatom. Non-limiting examples of aryl and heteroaryl groups
include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl,
2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl,
pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl,
3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl,
5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyridyl,
3-pyridyl, 4-pyridyl, 2-pyrimidyl, 4-pyrimidyl, 5-benzothiazolyl,
purinyl, 2-benzimidazolyl, 5-indolyl, 1-isoquinolyl, 5-isoquinolyl,
2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyl, and 6-quinolyl.
Substituents for each of the above noted aryl and heteroaryl ring
systems are selected from the group of acceptable substituents
described below.
[0032] For brevity, the term "aryl" when used in combination with
other terms (e.g., aryloxy, arylthioxy, arylalkyl) includes those
radicals in which an aryl group is attached through the next moiety
to the rest of the molecule. Thus, the term "arylalkyl" is meant to
include those radicals in which an aryl group is attached to an
alkyl group (e.g., benzyl, 1-(3-nitrophenyl)ethyl and the like). A
substituent such as benzyl or 1-(3-nitrophenyl)ethyl can also be
represented by `substituted alkyl` wherein the ethyl radical is
substituted with a 3-nitrophenyl moiety. The term "aryloxy" is
meant to include those radicals in which an aryl group is attached
to an oxygen atom. The term "aryloxyalkyl" is meant to include
those radicals in which an aryl group is attached to an oxygen atom
which is then attached to an alkyl group (e.g., phenoxymethyl,
3-(1-naphthyloxy)propyl, and the like).
[0033] For brevity, the term "heteroaryl" when used in combination
with other terms (e.g., heteroaryloxy, heteroarylthioxy,
heteroarylalkyl) includes those radicals in which a heteroaryl
group is attached through the next moiety to the rest of the
molecule. Thus, the term "heteroarylalkyl" is meant to include
those radicals in which a heteroaryl group is attached to an alkyl
group (e.g., pyridylmethyl and the like). The term "heteroaryloxy"
is meant to include those radicals in which a heteroaryl group is
attached to an oxygen atom. The term "heteroaryloxyalkyl" is meant
to include those radicals in which an aryl group is attached to an
oxygen atom which is then attached to an alkyl group. (e.g.,
2-pyridyloxymethyl and the like).
[0034] Each of the above terms (e.g., "alkyl," "heteroalkyl,"
"aryl" and "heteroaryl") are meant to include both substituted and
unsubstituted forms of the indicated radical. Preferred
substituents for each type of radical are provided below.
[0035] Substituents for the alkyl and heteroalkyl radicals
(including those groups often referred to as alkylene, alkenyl,
heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl,
heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl) are
generically referred to as "alkyl group substituents," and they can
be one or more of a variety of groups selected from, but not
limited to: --R', --OR', .dbd.O, .dbd.NR', .dbd.N--OR', --NR'R'',
--SR', -halogen, --SiR'R''R''', --OC(O)R', --C(O)R', --CO.sub.2R',
--CONR'R'', --OC(O)NR'R'', --NR''C(O)R', --NR'--C(O)NR''R''',
--NR''C(O).sub.2R', --NR'''''--C(NR'R''R''').dbd.NR'''',
--NR''''--C(NR'R'').dbd.NR''', --S(O)R', --S(O).sub.2R',
--S(O).sub.2NR'R'', --NR''SO.sub.2R', --CN, --NO.sub.2, --N.sub.3,
--CH(Ph).sub.2, fluoro(C.sub.1-C.sub.4)alkoxy, and
fluoro(C.sub.1-C.sub.4)alkyl, in a number ranging from zero to
(2m'+1), where m' is the total number of carbon atoms in such
radical. R', R'', R''', R'''' and R''''' each preferably
independently refer to hydrogen, substituted or unsubstituted
heteroalkyl, substituted or unsubstituted aryl, e.g., aryl
substituted with 1 or 2 or 3 halogens, substituted or unsubstituted
alkyl, alkoxy or thioalkoxy groups, or arylalkyl groups. When a
compound of the invention includes more than one R group, for
example, each of the R groups is independently selected as are each
R', R'', R''', R'''' and R''''' groups when more than one of these
groups is present. When R' and R'' are attached to the same
nitrogen atom, they can be combined with the nitrogen atom to form
a 5-, 6-, or 7-membered ring. For example, --NR'R'' is meant to
include, but not be limited to, 1-pyrrolidinyl and 4-morpholinyl.
From the above discussion of substituents, one of skill in the art
will understand that the term "alkyl" is meant to include groups
including carbon atoms bound to groups other than hydrogen groups,
such as haloalkyl (e.g., --CF.sub.3 and --CH.sub.2CF.sub.3) and
acyl (e.g., --C(O)CH.sub.3, --C(O)CF.sub.3,
--C(O)CH.sub.2OCH.sub.3, and the like).
[0036] Similar to the substituents described for the alkyl radical,
substituents for the aryl and heteroaryl groups are generically
referred to as "aryl group substituents." The substituents are
selected from, for example: --R', --OR', .dbd.O, .dbd.NR',
.dbd.N--OR', --NR'R'', --SR', -halogen, --SiR'R''R''', --OC(O)R',
--C(O)R', --CO.sub.2R', --CONR'R'', --OC(O)NR'R'', --NR''C(O)R',
--NR'--C(O)NR''R''', --NR''C(O).sub.2R', --NR''
''-C(NR'R''R''').dbd.NR'''', --NR''''--C(NR'R'').dbd.NR''',
--S(O)R', --S(O).sub.2R', --S(O).sub.2NR'R'', --NR''SO.sub.2R',
--CN, --NO.sub.2, --N.sub.3, --CH(Ph).sub.2,
fluoro(C.sub.1-C.sub.4)alkoxy, and fluoro(C.sub.1-C.sub.4)alkyl, in
a number ranging from zero to the total number of open valences on
the aromatic ring system; and where R', R'', R''', R'''' and R'''''
are preferably independently selected from hydrogen, substituted or
unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted aryl and substituted or unsubstituted
heteroaryl. When a compound of the invention includes more than one
R group, for example, each of the R groups is independently
selected as are each R', R'', R''', R'''' and R''''' groups when
more than one of these groups is present.
[0037] Two of the substituents on adjacent atoms of the aryl or
heteroaryl ring may optionally be replaced with a substituent of
the formula -T-C(O)--(CRR').sub.q--U--, wherein T and U are
independently --NR--, --O--, --CRR'-- or a single bond, and q is an
integer from 0 to 3. Alternatively, two of the substituents on
adjacent atoms of the aryl or heteroaryl ring may optionally be
replaced with a substituent of the formula
-A-(CH.sub.2).sub.r--B--, wherein A and B are independently
--CRR'--, --O--, --NR--, --S--, --S(O)--, --S(O).sub.2--,
--S(O).sub.2NR'-- or a single bond, and r is an integer from 1 or 2
or 3 or 4. One of the single bonds of the new ring so formed may
optionally be replaced with a double bond. Alternatively, two of
the substituents on adjacent atoms of the aryl or heteroaryl ring
may optionally be replaced with a substituent of the formula
--(CRR').sub.s--X--(CR''R''').sub.d--, where s and d are
independently integers from 0 or 1 or 2 or 3, and X is --O--,
--NR'--, --S--, --S(O)--, --S(O).sub.2--, or --S(O).sub.2NR'--. The
substituents R, R', R'' and R''' are preferably independently
selected from hydrogen or substituted or unsubstituted (C.sub.1 or
C.sub.2 or C.sub.3 or C.sub.4 or C.sub.5 or C.sub.6)alkyl.
[0038] "Ring" as used herein, means a substituted or unsubstituted
cycloalkyl, substituted or unsubstituted heterocycloalkyl,
substituted or unsubstituted aryl, or substituted or unsubstituted
heteroaryl. A ring includes fused ring moieties. The number of
atoms in a ring is typically defined by the number of members in
the ring. For example, a "5- to 7-membered ring" means there are 5
or 6 or 7 atoms in the encircling arrangement. Unless otherwise
specified, the ring optionally includes a heteroatom. Thus, the
term "5- to 7-membered ring" includes, for example phenyl,
pyridinyl and piperidinyl. The term "5- to 7-membered
heterocycloalkyl ring", on the other hand, would include pyridinyl
and piperidinyl, but not phenyl. The term "ring" further includes a
ring system comprising more than one "ring", wherein each "ring" is
independently defined as above.
[0039] As used herein, the term "heteroatom" includes atoms other
than carbon (C) and hydrogen (H). Examples include oxygen (O),
nitrogen (N) sulfur (S), silicon (Si), and boron (B).
[0040] The term "leaving group" means a functional group or atom
which can be displaced by another functional group or atom in a
substitution reaction, such as a nucleophilic substitution
reaction. By way of example, representative leaving groups include
triflate, chloro, bromo and iodo groups; sulfonic ester groups,
such as mesylate, tosylate, brosylate, nosylate and the like; and
acyloxy groups, such as acetoxy, trifluoroacetoxy and the like.
[0041] The symbol "R" is a general abbreviation that represents a
substituent group that is selected from substituted or
unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted aryl, substituted or unsubstituted
heteroaryl, substituted or unsubstituted cycloalkyl and substituted
or unsubstituted heterocycloalkyl groups.
[0042] By "effective" amount of a drug, formulation, or permeant is
meant a sufficient amount of an active agent to provide the desired
local or systemic effect. A "Topically effective,"
"pharmaceutically effective," or "therapeutically effective" amount
refers to the amount of drug needed to effect the desired
result.
[0043] The term "pharmaceutically acceptable salt" is meant to
include a salt of a compound of the invention which is prepared
with relatively nontoxic acids or bases, depending on the
particular substituents found on the compounds described herein.
When compounds of the invention contain relatively acidic
functionalities, base addition salts can be obtained by contacting
the neutral form of such compounds with a sufficient amount of the
desired base, either neat or in a suitable inert solvent. Examples
of pharmaceutically acceptable base addition salts include sodium,
potassium, calcium, ammonium, organic amino (such as choline or
diethylamine or amino acids such as d-arginine, 1-arginine,
d-lysine or 1-lysine), or magnesium salt, or a similar salt. When
compounds of the invention contain relatively basic
functionalities, acid addition salts can be obtained by contacting
the neutral form of such compounds with a sufficient amount of the
desired acid, either neat or in a suitable inert solvent. Examples
of pharmaceutically acceptable acid addition salts include those
derived from inorganic acids like hydrochloric, hydrobromic,
nitric, carbonic, monohydrogencarbonic, phosphoric,
monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,
monohydrogensulfuric, hydriodic, or phosphorous acids and the like,
as well as the salts derived from relatively nontoxic organic acids
like acetic, propionic, isobutyric, maleic, malonic, benzoic,
succinic, suberic, fumaric, lactic, mandelic, phthalic,
benzenesulfonic, p-tolylsulfonic, citric, tartaric,
methanesulfonic, and the like. Also included are salts of amino
acids such as arginate and the like, and salts of organic acids
like glucuronic or galactunoric acids and the like (see, for
example, Berge et al., "Pharmaceutical Salts", Journal of
Pharmaceutical Science 66: 1-19 (1977)). Certain specific compounds
of the invention contain both basic and acidic functionalities that
allow the compounds to be converted into either base or acid
addition salts.
[0044] The neutral forms of the compounds are preferably
regenerated by contacting the salt with a base or acid and
isolating the parent compounds in the conventional manner. The
parent form of the compound differs from the various salt forms in
certain physical properties, such as solubility in polar
solvents.
[0045] In addition to salt forms, the invention provides compounds
which are in a prodrug form. Prodrugs of the compounds described
herein readily undergo chemical changes under physiological
conditions to provide the compounds of the invention. Additionally,
prodrugs can be converted to the compounds of the invention by
chemical or biochemical methods in an ex vivo environment.
[0046] Certain compounds of the invention can exist in unsolvated
forms as well as solvated forms, including hydrated forms. In
general, the solvated forms are equivalent to unsolvated forms and
are encompassed within the scope of the invention. Certain
compounds of the invention may exist in multiple crystalline or
amorphous forms.
[0047] Certain compounds of the invention possess asymmetric carbon
atoms (optical centers) or double bonds; the racemates,
diastereomers, geometric isomers and individual isomers are
encompassed within the scope of the invention. The graphic
representations of racemic, ambiscalemic and scalemic or
enantiomerically pure compounds used herein are taken from Maehr,
J. Chem. Ed. 1985, 62: 114-120. Solid and broken wedges are used to
denote the absolute configuration of a stereocenter unless
otherwise noted. When the compounds described herein contain
olefinic double bonds or other centers of geometric asymmetry, and
unless specified otherwise, it is intended that the compounds
include both E and Z geometric isomers. Likewise, all tautomeric
forms are included.
[0048] Compounds of the invention can exist in particular geometric
or stereoisomeric forms. The invention contemplates all such
compounds, including cis- and trans-isomers, (-)- and
(+)-enantiomers, (R)- and (S)-enantiomers, diastereomers,
(D)-isomers, (L)-isomers, the racemic mixtures thereof, and other
mixtures thereof, such as enantiomerically or diastereomerically
enriched mixtures, as falling within the scope of the invention.
Additional asymmetric carbon atoms can be present in a substituent
such as an alkyl group. All such isomers, as well as mixtures
thereof, are intended to be included in this invention.
[0049] Optically active (R)- and (S)-isomers and d and l isomers
can be prepared using chiral synthons or chiral reagents, or
resolved using conventional techniques. If, for instance, a
particular enantiomer of a compound of the invention is desired, it
can be prepared by asymmetric synthesis, or by derivatization with
a chiral auxiliary, where the resulting diastereomeric mixture is
separated and the auxiliary group cleaved to provide the pure
desired enantiomers. Alternatively, where the molecule contains a
basic functional group, such as an amino group, or an acidic
functional group, such as a carboxyl group, diastereomeric salts
can be formed with an appropriate optically active acid or base,
followed by resolution of the diastereomers thus formed by
fractional crystallization or chromatographic means known in the
art, and subsequent recovery of the pure enantiomers. In addition,
separation of enantiomers and diastereomers is frequently
accomplished using chromatography employing chiral, stationary
phases, optionally in combination with chemical derivatization
(e.g., formation of carbamates from amines).
[0050] The compounds of the invention may also contain unnatural
proportions of atomic isotopes at one or more of the atoms that
constitute such compounds. For example, the compounds may be
radiolabeled with radioactive isotopes, such as for example tritium
(.sup.3H), iodine-125 (.sup.125I) or carbon-14 (.sup.14C). All
isotopic variations of the compounds of the invention, whether
radioactive or not, are intended to be encompassed within the scope
of the invention.
[0051] The term "pharmaceutically acceptable carrier" or
"pharmaceutically acceptable vehicle" refers to any formulation or
carrier medium that provides the appropriate delivery of an
effective amount of an active agent as defined herein, does not
interfere with the effectiveness of the biological activity of the
active agent, and that is sufficiently non-toxic to the host or
patient. Representative carriers include water, oils, both
vegetable and mineral, cream bases, lotion bases, ointment bases
and the like. These bases include suspending agents, thickeners,
penetration enhancers, and the like. Their formulation is well
known to those in the art of cosmetics and topical pharmaceuticals.
Additional information concerning carriers can be found in
Remington: The Science and Practice of Pharmacy, 21st Ed.,
Lippincott, Williams & Wilkins (2005) which is incorporated
herein by reference.
[0052] The term "excipients" is conventionally known to mean
carriers, diluents and/or vehicles used in formulating drug
compositions effective for the desired use.
[0053] The term "microbial infection" or "infection by a
microorganism" refers to any infection of a host tissue by an
infectious agent including, but not limited to, bacteria or
protozoa (see, e.g., Harrison's Principles of Internal Medicine,
pp. 93-98 (Wilson et al., eds., 12th ed. 1991); Williams et al., J.
of Medicinal Chem. 42:1481-1485 (1999), herein each incorporated by
reference in their entirety).
[0054] "Biological medium," as used herein refers to both in vitro
and in vivo biological milieus. Exemplary in vitro "biological
media" include, but are not limited to, cell culture, tissue
culture, homogenates, plasma and blood. In vivo applications are
generally performed in mammals, preferably humans.
[0055] "Inhibiting" and "blocking," are used interchangeably herein
to refer to the partial or full blockade of enzyme. In an exemplary
embodiment, the enzyme is an editing domain of a tRNA
synthetase.
[0056] Boron is able to form additional covalent or dative bonds
with oxygen, sulfur or nitrogen under some circumstances in this
invention.
[0057] Embodiments of the invention also encompass compounds that
are poly- or multi-valent species, including, for example, species
such as dimers, trimers, tetramers and higher homologs of the
compounds of use in the invention or reactive analogues
thereof.
[0058] "Salt counterion", as used herein, refers to positively
charged ions that associate with a compound of the invention when
the boron is fully negatively or partially negatively charged.
Examples of salt counterions include H.sup.+, H.sub.3O.sup.+,
ammonium, potassium, calcium, magnesium (such as choline or
diethylamine or amino acids such as d-arginine, 1-arginine,
d-lysine or 1-lysine) and sodium.
[0059] The compounds comprising a boron bonded to a carbon and
three heteroatoms (such as three oxygens described in this section)
can optionally contain a fully negatively charged boron or
partially negatively charged boron. Due to the negative charge, a
positively charged counterion may associate with this compound,
thus forming a salt. Examples of salt counterions include H.sup.+,
H.sub.3O.sup.+, ammonium, potassium, calcium, magnesium (such as
choline or diethylamine or amino acids such as d-arginine,
1-arginine, d-lysine or 1-lysine) and sodium. The salts of the
compounds are implicitly contained in descriptions of these
compounds.
II. Introduction
[0060] The invention provides novel boron compounds and methods for
the preparation of these molecules. The invention further provides
methods of treating bacterial infections, killing or inhibiting the
growth of bacteria in part or wholly through the use of the
compounds described herein. In another aspect, the invention is a
combination of a compound of the invention and an antibiotic. In
another aspect, the invention is a pharmaceutical formulation
comprising a pharmaceutically acceptable excipient and a compound
of the invention. In another aspect, the invention is a
pharmaceutical formulation comprising a compound of the invention,
an antibiotic, and a pharmaceutically acceptable excipient.
III. Composition of Matter
III. a.) Compounds
[0061] In one aspect the invention provides a compound of the
invention. In an exemplary embodiment, the invention provides a
compound described herein, or a salt thereof. In an exemplary
embodiment, the salt of a compound described herein is a
pharmaceutically acceptable salt. In an exemplary embodiment, the
invention provides a compound described herein, or a
pharmaceutically acceptable salt thereof. In an exemplary
embodiment, the invention provides a compound described in a
formula provided herein. In an exemplary embodiment, the invention
provides a compound described herein.
[0062] In an aspect, the invention provides a compound having a
structure which is:
##STR00003##
wherein R.sup.3 is substituted or unsubstituted nitroalkyl or
substituted or unsubstituted aminoalkyl; R.sup.4 is selected from
the group consisting of halogen, unsubstituted alkyl unsubstituted
alkoxy and unsubstituted phenyl; Y is O or S; R.sup.5 is selected
from the group consisting of substituted or unsubstituted alkyl and
substituted or unsubstituted heteroalkyl; or a salt, hydrate or
solvate thereof.
[0063] In an aspect, the invention provides a compound having a
structure which is:
##STR00004##
wherein R.sup.3 is substituted or unsubstituted nitroalkyl or
substituted or unsubstituted aminoalkyl; R.sup.4 is selected from
the group consisting of halogen, unsubstituted alkyl, unsubstituted
alkoxy, and unsubstituted phenyl; Y is O or S; R.sup.5 is selected
from the group consisting of substituted or unsubstituted alkyl and
substituted or unsubstituted heteroalkyl; or a salt, hydrate or
solvate thereof.
[0064] In an aspect, the invention provides a compound having a
structure which is:
##STR00005##
wherein R.sup.3 is substituted or unsubstituted nitroalkyl or
substituted or unsubstituted aminoalkyl; R.sup.4 is selected from
the group consisting of halogen, unsubstituted alkyl, unsubstituted
alkoxy, and unsubstituted phenyl; Y is O or S; R.sup.5 is selected
from the group consisting of substituted or unsubstituted alkyl and
substituted or unsubstituted heteroalkyl; or a salt, hydrate or
solvate thereof.
[0065] In an exemplary embodiment, there is provided a compound
having a structure according to the following formula:
##STR00006##
wherein C* is a carbon atom stereocenter which has a configuration
which is (R) or (S). In an exemplary embodiment, the C*
stereocenter is in the (S) configuration.
[0066] In an exemplary embodiment, there is provided a compound
having a structure according to the following formula:
##STR00007##
wherein C* is a carbon atom stereocenter which has a configuration
which is (R) or (S). In an exemplary embodiment, the C*
stereocenter is in the (S) configuration.
[0067] In an exemplary embodiment, there is provided a compound
having a structure according to the following formula:
##STR00008##
wherein C* is a carbon atom stereocenter which has a configuration
which is (R) or (S). In an exemplary embodiment, the C*
stereocenter is in the (S) configuration.
[0068] In an exemplary embodiment, Y, R.sup.5 and R.sup.4 are as
described herein, R.sup.3 is
--(CR.sup.20R.sup.21).sub.nNR.sup.22R.sup.23 in which n is an
integer selected from 1 to 10; each R.sup.20 and each R.sup.21 is
independently selected from the group consisting of H, R.sup.26,
OR.sup.26, NR.sup.26R.sup.27, SR.sup.26, --S(O)R.sup.26,
--S(O).sub.2R.sup.26, --S(O).sub.2NR.sup.26R.sup.27,
--C(O)R.sup.27, --C(O)OR.sup.27, --C(O)NR.sup.26R.sup.27; R.sup.22
and R.sup.23 are independently selected from the group consisting
of H, --S(O)R.sup.28, --S(O).sub.2R.sup.28,
--S(O).sub.2NR.sup.28R.sup.29, --C(O)R.sup.28, --C(O)OR.sup.28,
--C(O)NR.sup.28R.sup.29, substituted or unsubstituted alkyl,
substituted or unsubstituted heteroalkyl, substituted or
unsubstituted cycloalkyl, substituted or unsubstituted
heterocycloalkyl, substituted or unsubstituted aryl, and
substituted or unsubstituted heteroaryl, wherein each R.sup.26,
each R.sup.27, each R.sup.28 and each R.sup.29 is independently
selected from the group consisting of H, substituted or
unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted cycloalkyl, substituted or
unsubstituted heterocycloalkyl, substituted or unsubstituted aryl,
and substituted or unsubstituted heteroaryl.
[0069] In an exemplary embodiment, Y, R.sup.5 and R.sup.4 are as
described herein, and R.sup.3 is --CH.sub.2NH.sub.2 or
--CH.sub.2NO.sub.2. In an exemplary embodiment, Y, R.sup.5 and
R.sup.4 are as described herein, and R.sup.3 is --CH.sub.2NH.sub.2.
In an exemplary embodiment, Y, R.sup.5 and R.sup.4 are as described
herein, R.sup.3 is --CH.sub.2NH.sub.2, and C* has a configuration
which is (S).
[0070] In an exemplary embodiment, Y, R.sup.5 and R.sup.3 are as
described herein, and R.sup.4 is selected from the group consisting
of methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and
sec-butyl. In an exemplary embodiment, Y, R.sup.5 and R.sup.3 are
as described herein, and R.sup.4 is selected from the group
consisting of fluorine, chlorine, bromine, and iodine. In an
exemplary embodiment, Y, R.sup.5 and R.sup.3 are as described
herein, and R.sup.4 is chlorine or bromine. In an exemplary
embodiment, Y, R.sup.5 and R.sup.3 are as described herein, and
R.sup.4 is chlorine.
[0071] In an exemplary embodiment, Y, R.sup.5 and R.sup.3 are as
described herein, and R.sup.4 is selected from the group consisting
of methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, and
sec-butoxy. In an exemplary embodiment, Y, R.sup.5 and R.sup.3 are
as described herein, and R.sup.4 is methoxy or ethoxy. In an
exemplary embodiment, Y, R.sup.5 and R.sup.3 are as described
herein, and R.sup.4 is methoxy.
[0072] In an exemplary embodiment, Y, R.sup.4 and R.sup.3 are as
described herein, and R.sup.5 is:
##STR00009##
wherein a is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; each R.sup.10 and
each R.sup.11 is independently selected from the group consisting
of H, substituted or unsubstituted alkyl, OH and NH.sub.2; R.sup.12
is selected from the group consisting of H, R.sup.7, halogen,
cyano, amidino, OR.sup.7, NR.sup.7R.sup.8, SR.sup.7,
--N(R.sup.7)S(O).sub.2R.sup.8, --C(O)R.sup.7, --C(O)OR.sup.7,
--C(O)NR.sup.7R.sup.8 wherein each R.sup.7 and each R.sup.8 is
independently selected from the group consisting of H, substituted
or unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted cycloalkyl, substituted or
unsubstituted heterocycloalkyl, substituted or unsubstituted aryl,
and substituted or unsubstituted heteroaryl. In an exemplary
embodiment, Y, R.sup.4, R.sup.3, R.sup.10, R.sup.11, and R.sup.12
are as described herein, and a is 1, 2, 3, 4, or 5. In an exemplary
embodiment, Y, R.sup.4, R.sup.3, R.sup.10, R.sup.11, and R.sup.12
are as described herein, and a is 2, 3, or 4. In an exemplary
embodiment, Y, R.sup.4, R.sup.3, R.sup.10, R.sup.11, and R.sup.12
are as described herein, and a is 3. In an exemplary embodiment, Y,
R.sup.4, R.sup.3, a, and R.sup.12 are as described herein, and each
R.sup.10 and each R.sup.11 is independently selected from the group
consisting of H, substituted or unsubstituted alkyl, OH, and
NH.sub.2. In an exemplary embodiment, Y, R.sup.4, R.sup.3, a, and
R.sup.12 are as described herein, and each R.sup.10 and each
R.sup.11 is H. In an exemplary embodiment, Y, R.sup.4, R.sup.3,
R.sup.10, R.sup.11, and a are as described herein, and R.sup.12 is
selected from the group consisting of H, OH, NH.sub.2, methyl,
ethyl, --NHS(O).sub.2CH.sub.3, cyano, --NHC(O)CH.sub.3,
--NHC(O)NHCH.sub.2CH.sub.3, --C(O)NH.sub.2, --C(O)OH,
4-(methoxy)phenyl, benzyl, benzoxy, --NHC(O)OCH.sub.2Ph,
--C(O)NHCH.sub.2CH.sub.2OH and --C(NH.sub.2)(NH).
[0073] In an exemplary embodiment, R.sup.4, R.sup.3, and R.sup.5
are as described herein, and Y is O. In an exemplary embodiment,
R.sup.4, R.sup.3, and Y are as described herein, and R.sup.5 is
unsubstituted alkyl. In an exemplary embodiment, R.sup.4, R.sup.3,
and Y are as described herein, and R.sup.5 is selected from the
group consisting of methyl, ethyl, propyl, isopropyl, butyl,
isobutyl, t-butyl, and sec-butyl.
[0074] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, isopropyl, butyl, isobutyl,
t-butyl, and sec-butyl. In an exemplary embodiment, R.sup.4 is
halogen, R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is
selected from the group consisting of methyl, ethyl, propyl, and
isopropyl. In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of butyl, isobutyl, t-butyl, and sec-butyl.
[0075] In an exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is chlorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0076] In an exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is fluorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0077] In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is bromine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0078] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is methyl. In an exemplary
embodiment, R.sup.4 is fluorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is methyl. In an exemplary embodiment, R.sup.4 is
chlorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is
methyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is methyl.
[0079] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is ethyl. In an exemplary
embodiment, R.sup.4 is fluorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is ethyl. In an exemplary embodiment, R.sup.4 is
chlorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is
ethyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is ethyl.
[0080] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is propyl. In an exemplary
embodiment, R.sup.4 is fluorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is propyl. In an exemplary embodiment, R.sup.4 is
chlorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is
propyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is propyl.
[0081] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is isopropyl. In an
exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is isopropyl. In an
exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is isopropyl. In an
exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is isopropyl.
[0082] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.4
alkyl. In an exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.4
alkyl. In an exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.4
alkyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.4
alkyl.
[0083] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.5
alkyl. In an exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.5
alkyl. In an exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.5
alkyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.5
alkyl.
[0084] In an exemplary embodiment, R.sup.4 is halogen, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.6
alkyl. In an exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.6
alkyl. In an exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.6
alkyl. In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is unsubstituted C.sub.6
alkyl.
[0085] In an exemplary embodiment, R.sup.4 is chlorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is chlorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0086] In an exemplary embodiment, R.sup.4 is fluorine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is fluorine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0087] In an exemplary embodiment, R.sup.4 is bromine, R.sup.3
is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is selected from the group
consisting of methyl, ethyl, propyl, and isopropyl. In an exemplary
embodiment, R.sup.4 is bromine, R.sup.3 is-CH.sub.2NH.sub.2; Y is
O; and R.sup.5 is selected from the group consisting of butyl,
isobutyl, t-butyl, and sec-butyl.
[0088] In an exemplary embodiment, R.sup.4 is as described herein,
R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is substituted or
unsubstituted alkyl. In an exemplary embodiment, Y and R.sup.5 are
as described herein, R.sup.3 is-CH.sub.2NH.sub.2; and R.sup.4 is
halogen. In an exemplary embodiment, Y is as described herein,
R.sup.4 is halogen; Y is O; and R.sup.5 is unsubstituted alkyl. In
an exemplary embodiment, R.sup.3 is-CH.sub.2NH.sub.2; R.sup.4 is
chlorine; Y is O; and R.sup.5 is substituted or unsubstituted
alkyl. In an exemplary embodiment, R.sup.4 is as described herein,
R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is ethyl.
[0089] In an exemplary embodiment, the compound has a structure
which is
##STR00010##
[0090] In an exemplary embodiment, the compound has a structure
which is
##STR00011##
wherein R.sup.4, Y and R.sup.5 are as described herein.
[0091] In an exemplary embodiment, the compound has a structure
which is
##STR00012##
wherein R.sup.4, Y and R.sup.5 are as described herein. In an
exemplary embodiment, Y is O, and R.sup.4 and R.sup.5 are as
described herein. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 are as described herein. In an exemplary
embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
unsubstituted alkyl. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 is methyl or ethyl or propyl or isopropyl. In
an exemplary embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
butyl or isobutyl or neobutyl or t-butyl.
[0092] In an exemplary embodiment, the compound has a structure
which is
##STR00013##
wherein R.sup.4, Y and R.sup.5 are as described herein.
[0093] In an exemplary embodiment, the compound has a structure
which is
##STR00014##
wherein R.sup.4, Y and R.sup.5 are as described herein. In an
exemplary embodiment, Y is O, and R.sup.4 and R.sup.5 are as
described herein. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 are as described herein. In an exemplary
embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
unsubstituted alkyl. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 is methyl or ethyl or propyl or isopropyl. In
an exemplary embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
butyl or isobutyl or neobutyl or t-butyl.
[0094] In an exemplary embodiment, the compound has a structure
which is
##STR00015##
wherein R.sup.4, Y and R.sup.5 are as described herein.
[0095] In an exemplary embodiment, the compound has a structure
which is
##STR00016##
wherein R.sup.4, Y and R.sup.5 are as described herein. In an
exemplary embodiment, Y is O, and R.sup.4 and R.sup.5 are as
described herein. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 are as described herein. In an exemplary
embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
unsubstituted alkyl. In an exemplary embodiment, Y is O, R.sup.4 is
halogen, and R.sup.5 is methyl or ethyl or propyl or isopropyl. In
an exemplary embodiment, Y is O, R.sup.4 is halogen, and R.sup.5 is
butyl or isobutyl or neobutyl or t-butyl.
[0096] In an exemplary embodiment, said alkyl is linear alkyl or
branched alkyl, In an exemplary embodiment, said heteroalkyl is
linear heteroalkyl or branched heteroalkyl.
[0097] In an exemplary embodiment, the invention provides poly- or
multi-valent species of the compounds of the invention, including a
dimer or a trimer. Another exemplary embodiment of the invention
provides an anhydride of the compounds of the invention. In another
exemplary embodiment, the invention provides poly- or multi-valent
species of the compounds of the invention. In an exemplary
embodiment, the invention provides a dimer of the compounds
described herein. In an exemplary embodiment, the invention
provides a dimer of the compounds described herein.
[0098] In an exemplary embodiment, the invention provides an
anhydride of the compounds described herein. In an exemplary
embodiment, the invention provides an anhydride of the compounds
described herein.
[0099] In an exemplary embodiment, the invention provides a trimer
of the compounds described herein. In an exemplary embodiment, the
invention provides a trimer of the compounds described herein.
[0100] The compounds of the invention can form a hydrate with
water, solvates with alcohols such as methanol, ethanol, propanol,
and the like; adducts with amino compounds, such as ammonia,
methylamine, ethylamine, and the like; adducts with acids, such as
formic acid, acetic acid and the like; complexes with ethanolamine,
quinoline, amino acids, and the like.
[0101] In an exemplary embodiment, the invention provides a
compound described herein, or a salt, hydrate or solvate thereof,
or a combination thereof. In an exemplary embodiment, the invention
provides a compound described herein, or a salt, hydrate or solvate
thereof. In an exemplary embodiment, the invention provides a
compound described herein, or a salt thereof. In an exemplary
embodiment, the salt is a pharmaceutically acceptable salt. In an
exemplary embodiment, the invention provides a compound described
herein, or a hydrate thereof. In an exemplary embodiment, the
invention provides a compound described herein, or a solvate
thereof. In an exemplary embodiment, the invention provides a
compound described herein, or a prodrug thereof. In an exemplary
embodiment, the invention provides a salt of a compound described
herein. In an exemplary embodiment, the invention provides a
pharmaceutically acceptable salt of a compound described herein. In
an exemplary embodiment, the invention provides a hydrate of a
compound described herein. In an exemplary embodiment, the
invention provides a solvate of a compound described herein. In an
exemplary embodiment, the invention provides a prodrug of a
compound described herein. In an exemplary embodiment, the
invention provides a compound as described in FIG. 1, or a salt
thereof. In an exemplary embodiment, the invention provides a
compound as described in FIG. 1, or a pharmaceutically acceptable
salt thereof.
[0102] In an exemplary embodiment, alkyl is linear alkyl. In
another exemplary embodiment, alkyl is branched alkyl.
[0103] In an exemplary embodiment, heteroalkyl is linear
heteroalkyl. In another exemplary embodiment, heteroalkyl is
branched heteroalkyl.
III. b) Combinations Comprising Additional Therapeutic Agents
[0104] The compounds of the invention may also be used in
combination with additional therapeutic agents. The invention thus
provides, in a further aspect, a combination comprising a compound
of the invention together with at least one additional therapeutic
agent, or a salt, prodrug, hydrate or solvate thereof. In an
exemplary embodiment, the compound of the invention is a compound
described herein, or a salt thereof. In an exemplary embodiment,
the additional therapeutic agent is a compound of the invention. In
an exemplary embodiment, the additional therapeutic agent includes
a boron atom. In an exemplary embodiment, the additional
therapeutic agent does not contain a boron atom. In an exemplary
embodiment, the additional therapeutic agent is a compound
described in sections III a) or b).
[0105] When a compound of the invention is used in combination with
a second therapeutic agent active against the same disease state,
the dose of each compound may differ from that when the compound is
used alone. Appropriate doses will be readily appreciated by those
skilled in the art. It will be appreciated that the amount of a
compound of the invention required for use in treatment will vary
with the nature of the condition being treated and the age and the
condition of the patient and will be ultimately at the discretion
of the attendant physician or veterinarian.
[0106] In an exemplary embodiment, the additional therapeutic agent
is an antibacterial agent. In an exemplary embodiment, the
additional therapeutic agent is an antituberculosis agent. In an
exemplary embodiment, the additional therapeutic agent is
rifampicin. In an exemplary embodiment, the additional therapeutic
agent is isoniazid. In an exemplary embodiment, the additional
therapeutic agent is pyrazinamide. In an exemplary embodiment, the
additional therapeutic agent is ethambutol. In an exemplary
embodiment, the additional therapeutic agent is isoniazid. In an
exemplary embodiment, the additional therapeutic agent is
streptomycin. In an exemplary embodiment, the additional
therapeutic agent is an aminoglycoside. In an exemplary embodiment,
the additional therapeutic agent is amikacin or kanamycin. In an
exemplary embodiment, the additional therapeutic agent is a
polypeptide. In an exemplary embodiment, the additional therapeutic
agent is selected from the group consisting of capreomycin,
viomycin, and enviomycin. In an exemplary embodiment, the
additional therapeutic agent is a fluoroquinolone. In an exemplary
embodiment, the additional therapeutic agent is selected from the
group consisting of ciprofloxacin, levofloxacin, and moxifloxacin.
In an exemplary embodiment, the additional therapeutic agent is a
thioamide. In an exemplary embodiment, the additional therapeutic
agent is ethionamide or prothionamide. In an exemplary embodiment,
the additional therapeutic agent is cycloserine. In an exemplary
embodiment, the additional therapeutic agent is p-aminosalicylic
acid. In an exemplary embodiment, the additional therapeutic agent
is selected from the group consisting of rifabutin, linezolid,
thioacetazone, thioridazine, arginine, vitamin D, and R207910. In
an exemplary embodiment, the additional therapeutic agent is a
macrolide.
[0107] The individual components of such combinations may be
administered either simultaneously or sequentially in a unit dosage
form. The unit dosage form may be a single or multiple unit dosage
forms. In an exemplary embodiment, the invention provides a
combination in a single unit dosage form. An example of a single
unit dosage form is a capsule wherein both the compound of the
invention and the additional therapeutic agent are contained within
the same capsule. In an exemplary embodiment, the invention
provides a combination in a two unit dosage form. An example of a
two unit dosage form is a first capsule which contains the compound
of the invention and a second capsule which contains the additional
therapeutic agent. Thus the term `single unit` or `two unit` or
`multiple unit` refers to the object which the animal (for example,
a human) ingests, not to the interior components of the object.
Appropriate doses of known therapeutic agents will be readily
appreciated by those skilled in the art.
[0108] The combinations referred to herein may conveniently be
presented for use in the form of a pharmaceutical formulation.
Thus, an exemplary embodiment of the invention is a pharmaceutical
formulation comprising a) a compound of the invention; b) an
additional therapeutic agent and c) a pharmaceutically acceptable
excipient. In an exemplary embodiment, the pharmaceutical
formulation is a unit dosage form. In an exemplary embodiment, the
pharmaceutical formulation is a single unit dosage form. In an
exemplary embodiment, the pharmaceutical formulation is a single
unit dosage form which includes a compound of the invention; an
antibiotic and a pharmaceutically acceptable excipient. In an
exemplary embodiment, the pharmaceutical formulation is a single
unit dosage form which includes a compound of the invention; an
antibiotic and at least one pharmaceutically acceptable excipient.
In an exemplary embodiment, the pharmaceutical formulation is a two
unit dosage form. In an exemplary embodiment, the pharmaceutical
formulation is a two unit dosage form comprising a first unit
dosage form and a second unit dosage form, wherein the first unit
dosage form includes a) a compound of the invention and b) a first
pharmaceutically acceptable excipient; and the second unit dosage
form includes c) an additional therapeutic agent and d) a second
pharmaceutically acceptable excipient. In an exemplary embodiment,
the pharmaceutical formulation is a two unit dosage form comprising
a first unit dosage form and a second unit dosage form, wherein the
first unit dosage form includes a) a compound of the invention and
b) a first pharmaceutically acceptable excipient; and the second
unit dosage form includes c) an antibiotic and d) a second
pharmaceutically acceptable excipient.
III. c) Preparation of Boron-Containing Compounds
[0109] Compounds of use in the invention can be prepared using
commercially available starting materials, known intermediates, or
by using the synthetic methods published in references described
and incorporated by reference herein, such as U.S. patent
application Ser. No. 12/142,692 and U.S. Pat. Pubs. US20060234981,
US20070155699 and US20070293457.
[0110] The following general procedures were used as indicated in
generating the examples and can be applied, using the knowledge of
one of skill in the art, to other appropriate compounds to obtain
additional analogues.
##STR00017##
[0111] Scheme 1 describes a synthesis for compounds of (I), wherein
R.sup.4 is fluorine or chlorine and Y and R.sup.5 are as described
herein. The fluoro or chloro compound of formula A, which may be
prepared or may be available commercially from Sigma-Aldrich, is
reacted with a strong base (such as n-BuLi, sec-BuLi, or t-BuLi, 2
equiv) followed by quenching with a formylating agent (such as DMF,
dimethylformamide, formanilide, N-formylmorpholine, large excess)
to give the compound of formula B. Treatment of compound B with a
demethylating agent (typically BBr.sub.3, 2 equiv) in a suitable
solvent (dichloromethane, THF) gives the phenol of formula C.
Compound C can react with a corresponding bromide or mesylate
(1-1.5 equiv) in the presence of a base (such as KOtBu,
K.sub.2CO.sub.3, or Cs.sub.2CO.sub.3, 1.5-2 equiv) in an aprotic
solvent such as DMF or DMSO to afford the compound of formula D.
Compound D may be converted to triflate E by the reaction with 1.2
equiv of trifluoromethanesulfonic anhydride and pyridine in
dichloromethane. The conversion of triflate E to boronate F can be
achieved by the reaction with bis(pinacolato)diborane (2 equiv),
KOAc (3 equiv) and catalytic amount of PdCl.sub.2(dppf) (4-8 mol
%). The reaction of compound F with nitromethane (3 equiv) in the
presence of sodium hydroxide (3 equiv) in water or THF gives the
nitro compound of formula G. The compound G can be converted to the
final product of formula H by the Raney-Ni reduction (Raney Ni, 2
equiv w/w, 2.0 M NH.sub.3 in EtOH, absolute EtOH).
##STR00018##
[0112] Scheme 2 describes a synthesis for compounds of (I), wherein
R.sup.4 is chlorine, Y and R.sup.5 are as described herein. The
phenol or thiophenol of formula I, which may be prepared or may be
available commercially from Sigma-Aldrich, react with a solution of
bromine and catalytic amount of iron powder in glacial acetic acid
to give the bromo substituted compound of formula J. The alkylation
of J can be achieved by reacting with a bromide in the presence of
a base such as potassium carbonate in solvents like DMF or
acetonitrile. The protection of aldehyde K may be achieved by
refluxing with ethylene glycol in toluene, in the presence of
catalytic amount of p-toluenesulfonic acid. The reaction of the
compound L with BuLi and triisopropyl borate, followed by treating
with hydrochloric acid yields boronic acid M. The reaction of
compound M with nitromethane in the presence of sodium hydroxide
gives the nitro compound of formula N. The treatment of N with 1
equivalent of sulfuryl chloride affords the chloro substituted
compound O. The Raney-Ni reduction of compound O in MeOH gives the
final product of formula P.
##STR00019##
[0113] Scheme 3 describes a synthesis for compounds of (I), wherein
R.sup.4 is bromine, Y and R.sup.5 are as described herein. The
compound of formula N, which may be prepared according to Scheme 2,
can be reduced to the amine of formula S, by hydrogenation in the
presence of palladium hydroxide or Raney-Ni reduction as described
above. The amine of formula S reacts with an N-Protecting reagent
such as Boc anhydride in the presence of base like triethylamine in
dichloromethane to give Boc-protected compound of formula T. The
treatment of T with N-bromosuccinimide and catalytic amount of AIBN
in acetonitrile gives the bromo substituted compound of formula U.
Deprotection of compound U in the presence of acid such as HCl in
dioxane will afford the final compound of formula U.
##STR00020##
[0114] Scheme 4 describes a synthesis for compounds of (I), wherein
R.sup.4 is an alkyl or aryl group, Y and R.sup.5 are as described
herein. The compound of formula N, which may be prepared according
to Scheme 2, can be brominated with N-bromosuccinimide and
catalytic amount of AIBN in a solvent such as acetonitrile to give
the bromide of formula W. Stille coupling of W with an organotin
compound such as tetramethylstannane or tributyl-phenyl-stannane in
the presence of catalytic Pd(Ph.sub.3P).sub.4 in DMF affords the
compound of formula X. Compound X can be reduced to the final
compound of formula Y by hydrogenation in the presence of palladium
on carbon or Raney-Ni reduction as described above. Alternatively,
Stille reaction of W with an organotin compound such as
vinyltributyltin in the presence of catalytic amount of
Pd(Ph.sub.3P).sub.4 in DMF affords the compound of formula Z. After
Raney-Ni reduction of compound Z, further hydrogenation in the
presence of palladium on carbon as described above will afford the
final compound of formula AB.
##STR00021##
[0115] Scheme 5 describes a method to separate compounds (I) into
their enantiomeric isomers, wherein R.sup.4, Y, and R.sup.5 are as
described herein. The compound of formula AC, which may be prepared
according to Scheme 1 or Scheme 2 or Scheme 3 or Scheme 4, can be
converted to the Boc-protected compound AD by the reaction with an
N-Protecting reagent such as Boc anhydride in the presence of base
like triethylamine in dichloromethane. Racemic compound AD can be
resolved via chiral HPLC using a chiral column such as ChiralPak
AD-H and SF CO.sub.2/methanol as eluent. Two compounds collected
are enantiomer AE and enantiomer AF. Analysis of the enantiomeric
purity of each isomer can be achieved using a chiral column such as
ChiralPak AD column. The Boc-protected compounds AE and AF can be
converted to the final chiral compounds AG and AH, by deprotection
using acid such as HCl in dioxane.
##STR00022##
[0116] Scheme 6 describes an alternative method to separate chiral
compounds AI into their enantiomeric isomers, wherein R.sup.4, Y,
and R.sup.5 are as described herein. The compound of formula AI may
be prepared according to Scheme 1 or Scheme 2 or Scheme 3 or Scheme
4. The separation of the two enantiomers was achieved by dissolving
the racemic material AI in a suitable solvent and applying to an
appropriate chiral column and eluent system. The collected
separated enantiomer samples were then concentrated and used in the
next step without further purification. Using this technique, it is
possible to achieve a range of enantiomeric excesses of the
separated enantiomers. The nitro compound AJ and AK can be
converted to the final chiral compounds AL and AM, respectively, by
Raney-Ni reduction (Raney Ni, 2 equiv w/w, 2.0 M NH.sub.3 in EtOH,
absolute EtOH).
IV. Assays
[0117] Art-recognized techniques of genetics and molecular biology
are of use to identify compounds that bind to and/or inhibit an
enzyme, such as a tRNA synthetase. Moreover, these techniques are
of use to distinguish whether a compound binds to and/or inhibits a
particular domain of the enzyme. For example, for leucyl tRNA
synthetase (LeuRS), these techniques can distinguish whether a
compound binds to and/or inhibits the synthetic domain, the editing
domain, or both the editing and synthetic domains. The
Mycobacterium tuberculosis leuS gene was synthesized by Genscript
(Piscataway, N.J.) using E. coli optimized codons and protein was
made using standard T7 RNA polymerase over-expression protocols and
standard purification protocols.
IV. a) LeuRS
[0118] In an exemplary assay, activity of a representative compound
against the editing domain was confirmed. To identify the target of
a novel boron-containing antibacterial compound, mutants in E. coli
showing resistance to the compound were isolated. Characterization
of mutants showed that they have an 32-256 fold increase in
resistance to the compound over wildtype. The mutants were
furthermore shown to be sensitive to various antibacterial agents
with known modes of action, suggesting that the cellular target of
the compound is distinct from the target of the other antibacterial
agents. The leuS gene from the mutants was cloned onto a plasmid
and their resistance was confirmed by MIC. The editing domain from
these mutants were sequenced and the mutations were all located in
the editing domain of this enzyme.
[0119] Assays to determine whether, and how effectively, a
particular compound binds to and/or inhibits the editing domain of
a selected tRNA synthetase are also set forth herein, and
additional assays are readily available to those of skill in the
art. Briefly, in an exemplary assay, an improperly charged tRNA and
a tRNA synthetase that is capable of editing the improperly charged
tRNA are combined. The resulting mixture is contacted with the
putative inhibitor and the degree of editing inhibition is
observed.
[0120] Another assay uses genetics to show that a drug works via
the editing domain. In this assay, the compound is first tested
against a strain of cells over-expressing copies of the tRNA
synthetase gene. The compound's effect on the over-expressing
strain is compared with a control strain to determine whether the
compound is active against the synthetase. If the minimum
inhibitory concentration (MIC) is 2-fold higher in the strain with
extra copies of the synthetase gene than the MIC of the inhibitor
against a wild type cell, a further genetic screen is conducted to
determine whether the increased resistance is due to mutations in
the editing domain. In this second screen, the control strain is
challenged against a high concentration of the inhibitor. The
colonies surviving the challenge are isolated and DNA from these
cells is isolated. The editing domain is amplified using a
proof-reading PCR enzyme and the appropriate primers. The PCR
product can be purified using standard procedures. The sequence
amplified mutant DNA is compared to wild-type. If the mutant DNA
bears mutations in the editing domain, such results would suggest
that the compound binds to the editing domain and affects the
editing function of the molecule through this domain.
[0121] Generally, the compounds to be tested are present in the
assays in ranges from about 1 pM to about 100 mM, preferably from
about 1 pM to about 1 .mu.M. Other compounds range from about 1 nM
to about 100 nM, preferably from about 1 nM to about 1 .mu.M.
[0122] The effects of the test compounds upon the function of the
enzymes can also be measured by any suitable physiological change.
When the functional consequences are determined using intact cells
or animals, one can also measure a variety of effects such as
transmitter release, hormone release, transcriptional changes to
both known and uncharacterized genetic markers, changes in cell
metabolism such as cell growth or pH changes, and changes in
intracellular second messengers such as Ca.sup.2+, or cyclic
nucleotides.
[0123] Utilizing the assays set forth herein and others readily
available in the art, those of skill in the art will be able to
readily and routinely determine other compounds and classes of
compounds that operate to bind to and/or inhibit the editing domain
of tRNA synthetases.
[0124] In another aspect, the invention provides a method for
identifying a compound which binds to an editing domain of a tRNA
synthetase comprising:
a) contacting said editing domain with a test compound under
conditions suitable for binding; and b) detecting binding of said
test compound to said editing domain. In an exemplary embodiment,
detecting binding of said compound comprises use of at least one
detectable element, isotope, or chemical label attached to said
compound. In an exemplary embodiment, the element, isotope or
chemical label is detected by a fluorescent, luminescent,
radioactive, or absorbance readout. In an exemplary embodiment, the
contacting of said test compound with said editing domain also
includes further contacting said test compound and said editing
domain with a member selected from AMP and a molecule with a
terminal adenosine. In an exemplary embodiment, the tRNA synthetase
is derived from leucyl tRNA synthetase. In an exemplary embodiment,
the tRNA synthetase is derived from a mutated tRNA synthetase,
wherein said mutated tRNA synthetase comprises amino acid mutations
in an editing domain. In another exemplary embodiment, wherein said
editing domain of a tRNA synthetase comprises the amino acid
sequence of a peptide sequence described herein.
[0125] In another aspect, the invention provides a method for
identifying a compound which binds to an editing domain of a tRNA
synthetase, said assay comprising: a) contacting said editing
domain of a tRNA synthetase with said compound under conditions
suitable for binding of said compound with said editing domain of a
tRNA synthetase; b) comparing a biological activity of said editing
domain of a tRNA synthetase contacting said compound to said
biological activity when not contacting said compound; and c)
identifying said compound as binding to said editing domain of a
tRNA synthetase if said biological activity of said editing domain
of a tRNA synthetase is reduced when contacting said compound. In
an exemplary embodiment, the biological activity is hydrolysis of
noncognate amino acid. In another exemplary embodiment, the
hydrolysis of said noncognate amino acid is detected through the
use of one or more labels. In another exemplary embodiment, the
labels include a radiolabel, a fluorescent marker, an antibody, or
a combination thereof. In another exemplary embodiment, said labels
can be detected using spectroscopy. In another exemplary
embodiment, said editing domain of a tRNA synthetase is derived
from leucyl tRNA synthetase.
[0126] In another aspect, the invention provides a method of
generating a tRNA molecule with a noncognate amino acid comprising:
a) creating or isolating a mutated tRNA synthetase with altered
amino acid editing domains; and b) contacting a tRNA molecule with
said mutated tRNA synthetase and a noncognate amino acid. In
another exemplary embodiment, the mutated tRNA synthetase contains
one or more amino acid mutations in an editing domain. In another
exemplary embodiment, the mutated tRNA synthetase is unable to bind
with a compound of the invention. In another exemplary embodiment,
the mutated tRNA synthetase is unable to bind with a compound
described herein, or a pharmaceutically acceptable salt thereof. In
another exemplary embodiment, the mutated tRNA synthetase is unable
to bind with a compound according to a formula described herein, or
a pharmaceutically acceptable salt thereof.
[0127] In another aspect, the invention provides a composition that
comprises one or more tRNA molecules attached to noncognate amino
acids, wherein said tRNA molecules are synthesized using one or
more mutated tRNA synthetases isolated from a microorganism or a
cell line derived from a microorganism. In an exemplary embodiment,
the microorganism is a bacteria. In an exemplary embodiment,
wherein said mutated tRNA synthetases contain amino acid mutations
in their editing domains.
V. Amino Acid and Nucleotide Sequences Used in Assays
[0128] Amino acid and nucleotide sequences of use in the invention
are published in references described and incorporated by reference
herein, such as U.S. Pat. No. 7,816,344 and U.S. Pat. Pubs.
US20060234981, US20070155699 and US20070293457. The sequence for
the codon optimized M. tuberculosis leuS gene is as follows:
TABLE-US-00001 (SEQ. ID. 1)
CATATGACCGAAAGCCCGACCGCAGGTCCGGGTGGTGTGCCGCGTGCG
GATGATGCAGATAGCGATGTGCCGCGTTATCGTTATACCGCGGAACTGG
CGGCGCGTCTGGAACGTACCTGGCAGGAAAACTGGGCGCGTCTGGGCAC
CTTTAACGTGCCGAACCCGGTGGGTAGCCTGGCACCGCCGGATGGTGCA
GCAGTGCCGGATGATAAACTGTTTGTGCAGGATATGTTTCCGTATCCGA
GCGGCGAAGGCCTGCATGTGGGCCATCCGCTGGGCTATATTGCGACCGA
TGTGTATGCGCGTTATTTTCGTATGGTGGGCCGTAACGTGCTGCATGCG
CTGGGCTTTGATGCGTTTGGTCTGCCGGCGGAACAGTATGCGGTGCAGA
CCGGCACCCATCCGCGTACCCGTACCGAAGCGAACGTGGTGAACTTTCG
TCGTCAGCTGGGCCGTCTGGGCTTTGGCCATGATAGCCGTCGTAGCTTT
AGCACCACCGATGTGGATTTTTATCGTTGGACCCAGTGGATTTTTCTGC
AGATTTATAACGCGTGGTTTGATACCACCGCGAACAAAGCGCGTCCGAT
TAGCGAACTGGTGGCGGAATTTGAAAGCGGTGCACGTTGCCTGGATGGT
GGTCGTGATTGGGCAAAACTGACCGCAGGTGAACGTGCGGATGTGATTG
ATGAATATCGTCTGGTGTATCGTGCGGATAGCCTGGTGAACTGGTGCCC
GGGTCTGGGTACCGTGCTGGCAAACGAAGAAGTGACCGCAGATGGCCGT
AGCGATCGTGGCAACTTTCCGGTGTTTCGTAAACGTCTGCGTCAGTGGA
TGATGCGTATTACCGCGTATGCGGATCGTCTGCTGGATGATCTGGATGT
GCTGGATTGGCCGGAACAGGTGAAAACCATGCAGCGTAACTGGATTGGC
CGTAGCACCGGCGCGGTGGCGCTGTTTAGCGCGCGTGCGGCGAGCGATG
ATGGCTTTGAAGTGGATATTGAAGTGTTTACCACCCGTCCGGATACCCT
GTTTGGCGCGACCTATCTGGTGCTGGCGCCGGAACATGATCTGGTGGAT
GAACTGGTGGCGGCAAGCTGGCCGGCAGGTGTGAACCCGCTGTGGACCT
ATGGCGGTGGTACCCCGGGTGAAGCAATTGCAGCATATCGTCGTGCGAT
TGCGGCGAAAAGCGATCTGGAACGTCAGGAAAGCCGTGAAAAAACCGGC
GTGTTTCTGGGCAGCTATGCGATTAACCCGGCGAACGGCGAACCGGTGC
CGATTTTTATTGCGGATTATGTGCTGGCGGGCTATGGCACCGGCGCGAT
TATGGCGGTGCCGGGCCATGATCAGCGTGATTGGGATTTTGCGCGTGCG
TTTGGCCTGCCGATTGTGGAAGTGATTGCAGGTGGAAACATTAGCGAAA
GCGCGTATACCGGCGATGGCATTCTGGTGAACAGCGATTATCTGAACGG
CATGAGCGTGCCGGCAGCAAAACGTGCAATTGTGGATCGTCTGGAAAGC
GCAGGTCGTGGTCGTGCACGTATTGAATTTAAACTGCGTGATTGGCTGT
TTGCGCGTCAGCGTTATTGGGGCGAACCGTTTCCGATTGTGTATGATAG
CGATGGCCGTCCGCATGCGCTGGATGAAGCGGCGCTGCCGGTGGAACTG
CCGGATGTGCCGGATTATAGCCCGGTGCTGTTTGATCCGGATGATGCGG
ATAGCGAACCGAGCCCGCCGCTGGCGAAAGCGACCGAATGGGTGCATGT
GGATCTGGATCTGGGCGATGGCCTGAAACCGTATAGCCGTGATACCAAC
GTGATGCCGCAGTGGGCGGGCAGCAGCTGGTATGAACTGCGTTATACCG
ATCCGCATAACAGCGAACGTTTTTGCGCGAAAGAAAACGAAGCGTATTG
GATGGGTCCGCGTCCGGCAGAACATGGTCCGGATGATCCGGGTGGTGTG
GATCTGTATGTGGGCGGCGCGGAACATGCGGTGCTGCATCTGCTGTATA
GCCGTTTTTGGCATAAAGTGCTGTATGATCTGGGCCATGTGAGCAGCCG
TGAACCGTATCGTCGTCTGGTGAACCAGGGCTATATTCAGGCGTATGCG
TATACCGATGCGCGTGGCAGCTATGTGCCGGCGGAACAAGTGATTGAAC
GTGGCGATCGTTTTGTGTATCCGGGCCCGGATGGCGAAGTGGAAGTGTT
TCAGGAATTTGGCAAAATTGGCAAAAGCCTGAAAAACAGCGTGAGCCCG
GATGAAATTTGCGATGCGTATGGCGCGGATACCCTGCGTGTGTATGAAA
TGAGCATGGGCCCGCTGGAAGCGAGCCGTCCGTGGGCGACCAAAGATGT
GGTGGGCGCGTATCGTTTTCTGCAGCGTGTGTGGCGTCTGGTGGTGGAT
GAACATACCGGCGAAACCCGTGTGGCGGATGGCGTGGAACTGGATATTG
ATACCCTGCGTGCGCTGCATCGTACCATTGTGGGCGTGAGCGAAGATTT
TGCGGCGCTGCGTAACAACACCGCGACCGCGAAACTGATTGAATATACC
AACCATCTGACCAAAAAACATCGTGATGCAGTGCCGCGTGCGGCAGTGG
AACCGCTGGTGCAGATGCTGGCACCGCTGGCACCGCATATTGCGGAAGA
ACTGTGGCTGCGTCTGGGCAACACCACCAGCCTGGCGCATGGCCCGTTT
CCGAAAGCGGATGCGGCGTATCTGGTGGATGAAACCGTGGAATATCCGG
TGCAGGTGAACGGCAAAGTGCGTGGTCGTGTGGTGGTGGCGGCGGATAC
CGATGAAGAAACCCTGAAAGCGGCGGTGCTGACCGATGAAAAAGTGCAG
GCGTTTCTGGCGGGCGCGACCCCGCGTAAAGTGATTGTGGTGGCGGGCC
GTCTGGTGAACCTGGTGATTTAACTCGAG
VI. Methods
[0129] In another aspect, the compounds of the invention can be
utilized to inhibit an enzyme. In another aspect, the compounds of
the invention and/or combinations of the invention exhibit potency
against microorganisms, such as bacteria, and therefore have the
potential to kill and/or inhibit the growth of microorganisms. In
another aspect, the compounds of the invention and/or combinations
of the invention exhibit potency against microorganisms, such as
bacteria, and therefore have the potential to achieve therapeutic
efficacy in the animals described herein.
VI. a) LeuRS
[0130] In an exemplary embodiment, the compounds of the invention
exhibit the ability of inhibiting the editing domain of tRNA
synthetases, such as leucyl tRNA synthetase, of microorganisms,
such as bacteria, and therefore have the potential to be used as
editing domain inhibitors of microorganism tRNA synthetases.
[0131] According to another aspect of the invention, a method for
binding to and/or inhibiting the editing domain of a tRNA
synthetase is provided which comprises contacting a tRNA synthetase
with a compound of the invention that inhibits the editing domain
under conditions in which the tRNA synthetase interacts with its
substrate to form an aminoacyl adenylate intermediate and,
preferably, to form a charged tRNA. Such conditions are known to
those skilled in the art. In an exemplary embodiment, the compound
has a structure according to a formula described herein. In an
exemplary embodiment, the compound is described herein, or a salt,
hydrate or solvate thereof, or a combination thereof. In an
exemplary embodiment, the invention provides a compound described
herein, or a salt, hydrate or solvate thereof. In an exemplary
embodiment, the invention provides a compound described herein, or
a salt thereof. In an exemplary embodiment, the invention provides
a compound described herein, or a salt thereof. The tRNA synthetase
is contacted with an amount of compound of the invention sufficient
to result in a detectable amount of tRNA synthetase inhibition.
This method can be performed on a tRNA synthetase that is contained
within an organism or which is outside an organism. In an exemplary
embodiment, the method is performed on a tRNA synthetase that is
contained within a microorganism or a microbial cell that is in, or
on the surface of, an animal. In an exemplary embodiment, the
animal is a human. The method results in a decrease in the amount
of charged tRNA produced by the tRNA synthetase that has an
inhibited editing domain. In an exemplary embodiment, the
inhibition takes place in a cell, such as a microorganism cell. In
another exemplary embodiment, the microorganism cell is a bacteria.
In another exemplary embodiment, the tRNA synthetase is leucyl tRNA
synthetase.
[0132] In an exemplary embodiment, the invention provides a method
of inhibiting conversion of a tRNA molecule into a charged tRNA
molecule. The method involves contacting a tRNA synthetase with a
compound of the invention effective to inhibit activity of an
editing domain of said tRNA synthetase, under conditions sufficient
to inhibit said activity, thereby inhibiting said conversion. In an
exemplary embodiment, the compound of the invention is a compound
described herein, or a pharmaceutically acceptable salt thereof. In
an exemplary embodiment, the inhibition occurs within a cell, and
the cell is a microorganism cell. In another exemplary embodiment,
the microorganism cell is a bacteria. In another exemplary
embodiment, the microorganism cell is a bacteria which is described
herein. In another exemplary embodiment, the enzyme is a leucyl
tRNA synthetase of a bacteria described herein. In another
exemplary embodiment, the tRNA synthetase is leucyl tRNA
synthetase. In another exemplary embodiment, the compound has a
K.sub.D, synthesis of greater than 100 pM against a synthetic
domain of said tRNA synthetase.
[0133] In certain embodiments, the mechanism of action of a
compound of the invention is to inhibit the conversion of a tRNA
molecule into a charged tRNA molecule by binding to and/or
inhibiting at least the editing domain of the synthetase. The
compounds of use in this method may also inhibit or otherwise
interact with the synthetic domain (e.g., the active site of the
synthetic domain). In a presently preferred embodiment, the editing
domain is inhibited selectively in the presence of the synthetic
domain. In a preferred embodiment, the synthetic domain is
essentially uninhibited, while the editing domain is inhibited at
least 50%, preferably at least 60%, more preferably at least 70%,
still more preferably, at least 80% and even still more preferably
at least 90% of the activity of the tRNA synthetase. In another
preferred embodiment, the synthetic domain is inhibited by at most
50%, preferably at most 30%, preferably at most 20%, 10%,
preferably at most 8%, more preferably at most 5%, still more
preferably, at most 3% and even still more preferably at most 1%.
Inhibition of the editing domain produces a decrease in the amount
of the properly charged tRNA which results in retardation or
cessation of cell growth and division.
[0134] In another exemplary embodiment, the ratio of a minimum
concentration of said compound inhibiting said editing domain to a
minimum concentration of said compound inhibiting said synthetic
domain of said tRNA synthetase, represented as K.sub.D,
edit/K.sub.D, synthesis, is less than one. In another exemplary
embodiment, the K.sub.D, edit/K.sub.D, synthesis of the compound is
a member selected from less than 0.5, less than 0.1 and less than
0.05.
VI. b) Inhibiting Microorganism Growth or Killing
Microorganisms
[0135] The compounds of the invention and/or combinations of the
invention exhibit potency against microorganisms, such as bacteria,
and therefore have the potential to treat, and/or prevent a
microorganism infection, or kill and/or inhibit the growth of
microorganisms.
[0136] In a further aspect, the invention provides a method of
treating and/or preventing a microorganism infection, or a method
of killing and/or inhibiting the growth of a microorganism, said
method comprising: contacting said microorganism with an effective
amount of a compound of the invention, thereby killing and/or
inhibiting the growth of the microorganism. In a further aspect,
the invention provides a method of treating and/or preventing a
microorganism infection, or a method of killing and/or inhibiting
the growth of a microorganism, said method comprising: contacting
said microorganism with an effective amount of a combination of the
invention, thereby killing and/or inhibiting the growth of the
microorganism.
[0137] In a further aspect, the invention provides a method of
treating a bacterial infection comprising administering to an
animal suffering from the infection an effective amount of a
compound of the invention or a combination of the invention, or a
pharmaceutically acceptable salt thereof, thereby treating the
bacterial infection. In an exemplary embodiment, the invention
provides a method of treating a bacterial infection comprising
administering to an animal suffering from the infection an
effective amount of a compound of the invention, or a
pharmaceutically acceptable salt thereof, and an effective amount
of an antibiotic, or a pharmaceutically acceptable salt thereof,
thereby treating the bacterial infection.
[0138] In a further aspect, the invention provides a method of
preventing a bacterial infection comprising administering to an
animal a prophylactic amount of a compound of the invention or a
combination of the invention, or a pharmaceutically acceptable salt
thereof, thereby treating the bacterial infection. In an exemplary
embodiment, the invention provides a method of preventing a
bacterial infection comprising administering to an animal a
prophylactic amount of a compound of the invention, or a
pharmaceutically acceptable salt thereof.
[0139] In an exemplary embodiment, the microorganism is a bacteria.
In an exemplary embodiment, the compound or combination is
described herein, or a salt, prodrug, hydrate or solvate thereof,
or a combination thereof. In an exemplary embodiment, the invention
provides a compound or combination described herein, or a salt,
hydrate or solvate thereof. In an exemplary embodiment, the
invention provides a compound or combination described herein, or a
prodrug thereof. In an exemplary embodiment, the invention provides
a compound or combination described herein, or a salt thereof. In
another exemplary embodiment, the compound or combination of the
invention is a compound or combination described herein, or a
pharmaceutically acceptable salt thereof. In another exemplary
embodiment, the compound or compound of the combination is
described by a formula listed herein, or a pharmaceutically
acceptable salt thereof. In an exemplary embodiment, the compound
is part of a combination described herein. In an exemplary
embodiment, the compound is part of a pharmaceutical formulation
described herein. In another exemplary embodiment, the contacting
occurs under conditions which permit entry of the compound into the
organism. Such conditions are known to one skilled in the art and
are described herein.
[0140] In another aspect, the microorganism is inside, or on the
surface of an animal. In another exemplary embodiment, the animal
is described herein. In another exemplary embodiment, the animal is
a human.
[0141] In an exemplary embodiment, the microorganism infection is
treated and/or prevented, or the microorganism is killed or its
growth is inhibited, through oral administration of the compound of
the invention and/or the combination of the invention. In an
exemplary embodiment, the microorganism infection is treated and/or
prevented, or the microorganism is killed or its growth is
inhibited through intravenous administration of the compound of the
invention and/or the combination of the invention.
[0142] In an exemplary embodiment, the microorganism is a
bacterium. In an exemplary embodiment, an infection is caused by
and/or associated with a microorganism, particularly a bacterium.
In an exemplary embodiment, the bacterium is a gram-positive
bacteria. In another exemplary embodiment, the gram-positive
bacterium is selected from the group consisting of Staphylococcus
species, Streptococcus species, Bacillus species, Mycobacterium
species, Corynebacterium species (Propionibacterium species),
Clostridium species, Actinomyces species, Enterococcus species and
Streptomyces species. In another exemplary embodiment, the
gram-positive bacterium is selected from the group consisting of
Propionibacterium acnes, Staphylococcus aureus, Staphylococcus
epidermidis, Staphylococcus saprophyticus, Staphylococcus
haemolyticus, Streptococcus pyogenes, Streptococcus agalactiae,
Streptococcus pneumoniae, Enterococcus faecalis, Enterococcus
faecium, Actinomyces israelii, Bacillus anthracis, Corynebacterium
diphtheria, Clostridium perfringens, Clostridium botulinum,
Clostridium tetani, and Clostridium difficile. In another exemplary
embodiment, the gram-positive bacterium is selected from the group
consisting of Staphylococcus aureus, Staphylococcus epidermidis,
Streptococcus pneumoniae, Streptococcus pyogenes, Enterococcus
faecalis, Enterococcus faecium, Clostridium difficile and
Propionibacter acnes. In another exemplary embodiment, the
bacterium is a gram-negative bacterium. In another exemplary
embodiment, the gram-negative bacterium is selected from the group
consisting of Acinetobacter species, Neisseria species, Pseudomonas
species, Brucella species, Agrobacterium species, Bordetella
species, Escherichia species, Shigella species, Yersinia species,
Salmonella species, Klebsiella species, Enterobacter species,
Haemophilus species, Pasteurella species, Streptobacillus species,
spirochetal species, Campylobacter species, Vibrio species,
Helicobacter species, Bacteroides species, Citrobacter species,
Proteus species, Providencia species, Serratia species,
Stenotrophomonas species and Burkholderia species. In another
exemplary embodiment, the gram-negative bacterium is selected from
the group consisting ofAcinetobacter species, Pseudomonas species,
Escherichia species, Klebsiella species, Enterobacter species,
Bacteroides species, Citrobacter species, Proteus species,
Providencia species, Serratia species, Stenotrophomonas species and
Burkholderia species. In another exemplary embodiment, the
gram-negative bacterium is selected from the group consisting of
Neisseria gonorrhoeae, Neisseria meningitidis, Pseudomonas
aeruginosa, Legionella pneumophila, Escherichia coli, Yersinia
pestis, Haemophilus influenzae, Helicobacter pylori, Campylobacter
fetus, Campylobacter jejuni, Vibrio cholerae, Vibrio
parahemolyticus, Treponema pallidum, Rickettsia prowazekii,
Rickettsia rickettsii, Chlamydia trachomatis, Chlamydia psittaci,
Brucella abortus, Agrobacterium tumefaciens, Francisella
tularensis, Klebsiella pneumoniae, Enterobacter cloacae,
Acinetobacter baumannii, Bacteroides fragilis, Citrobacter
freundii, Proteus mirabilis, Providencia stuartii, Serratia
marcescens, Stenotrophomonas maltophilia and Burkholderia cepacia.
In another exemplary embodiment, the gram-negative bacterium is
selected from the group consisting of Pseudomonas aeruginosa,
Escherichia coli, Haemophilus influenzae, Klebsiella pneumoniae,
Enterobacter cloacae, Acinetobacter baumannii, Bacteroides
fragilis, Citrobacter freundii, Proteus mirabilis, Providencia
stuartii, Serratia marcescens, Stenotrophomonas maltophilia and
Burkholderia cepacia. In another exemplary embodiment, the
gram-negative bacterium is selected from the group consisting of
Enterobacter aerogenes, Enterobacter cloacae, Enterobacter
sakazakii, Escherichia coli, Klebsiella pneumoniae, Proteus
mirabilis, Serratia marcescens and Citrobacter freundii. In another
exemplary embodiment, the gram-negative bacterium is a Providencia
spp.
[0143] In an exemplary embodiment, the microorganism is an
acid-fact bacteria. In another exemplary embodiment, the bacterium
is a Mycobacterium spp. In another exemplary embodiment, the
bacterium is Mycobacterium avium. In another exemplary embodiment,
the bacterium is Mycobacterium avium-intracellulare. In another
exemplary embodiment, the bacterium is Mycobacterium kansasii. In
another exemplary embodiment, the bacterium is Mycobacterium
leprae. In another exemplary embodiment, the bacterium is
Mycobacterium lepromatosis. In another exemplary embodiment, the
bacterium is Mycobacterium africanum. In another exemplary
embodiment, the bacterium is Mycobacterium canetti. In another
exemplary embodiment, the bacterium is Mycobacterium microti. In
another exemplary embodiment, the bacterium is Mycobacterium
tuberculosis. In another exemplary embodiment, the bacterium is
Mycobacterium tuberculosis which is multi-drug resistant. In
another exemplary embodiment, the bacterium is Mycobacterium
tuberculosis which is extensively drug resistant. In another
exemplary embodiment, the bacterium is Mycobacterium tuberculosis
which is resistant to rifampicin. In another exemplary embodiment,
the bacterium is Mycobacterium tuberculosis which is resistant to
isoniazid. In another exemplary embodiment, the bacterium is
Mycobacterium tuberculosis which is resistant to kanamycin. In
another exemplary embodiment, the bacterium is Mycobacterium
tuberculosis which is resistant to capreomycin. In another
exemplary embodiment, the bacterium is Mycobacterium tuberculosis
which is resistant to amikacin.
[0144] In another exemplary embodiment, the bacterium is a
Pseudomonas species. In another exemplary embodiment, the bacterium
is Pseudomonas aeruginosa. In another exemplary embodiment, the
bacterium is selected from the group consisting of Pseudomonas
aeruginosa, Acinetobacter baumannii, Stenotrophomonas maltophilia
and Burkholderia cepacia. In another exemplary embodiment, the
bacterium is Acinetobacter baumannii. In another exemplary
embodiment, the bacterium is Stenotrophomonas maltophilia. In
another exemplary embodiment, the bacterium is Burkholderia
cepacia. In another exemplary embodiment, the bacterium is
Acinetobacter species. In another exemplary embodiment, the
bacterium is Acinetobacter anitratus. In another exemplary
embodiment, the bacterium is selected from the group consisting of
Enterobacter aerogenes, Enterobacter cloacae, Enterobacter
sakazakii, E. coli, K. pneumoniae, P. mirabilis, Serratia
marcescens, Citrobacter freundii and Providencia spp. In another
exemplary embodiment, the bacterium is selected from the group
consisting of Enterobacter aerogenes, Enterobacter cloacae,
Enterobacter sakazakii, E. coli, K. pneumoniae, P. mirabilis,
Serratia marcescens, Citrobacter freundii, Providencia spp., S.
aureus, S. pneumonia, S. pyogenes, E. faecalis, and E. faecium. In
another exemplary embodiment, the bacterium is selected from the
group consisting of Pseudomonas aeruginosa, Acinetobacter
baumannii, Stenotrophomonas maltophilia, Burkholderia cepacia. In
another exemplary embodiment, the bacterium is selected from the
group consisting of S. aureus, S. pneumonia, S. pyogenes, E.
faecalis, and E. faecium. In another exemplary embodiment, the
bacterium is selected from the group consisting of Viridans group
Strep. In another exemplary embodiment, the bacterium is selected
from the group consisting of Strep. mitis, Strep. mutans, Strep.
oralis, Strep. sanguis, Strep. sobrinus and Strep. millari. In
another exemplary embodiment, the bacterium is S. pneumonia. In
another exemplary embodiment, the bacterium is H. influenzae. In
another exemplary embodiment, the bacterium is S. aureus. In
another exemplary embodiment, the bacterium is M. catarrhalis. In
another exemplary embodiment, the bacterium is M. pneumoniae. In
another exemplary embodiment, the bacterium is L. pneumoniae. In
another exemplary embodiment, the bacterium is C. pneumoniae. In
another exemplary embodiment, the bacterium is S. pyogenes. In
another exemplary embodiment, the bacterium is an anaerobe. In
another exemplary embodiment, the bacterium is an Alcaligenes
species. In another exemplary embodiment, the bacterium is a B.
cepacia. In another exemplary embodiment, the bacterium is selected
from the group consisting of Enterobacter cloacae, Escherichia
coli, Klebsiella pneumoniae, Proteus mirabilis, Providencia
stuartii, Serratia marcescens, and Citrobacter freundii. In another
exemplary embodiment, the bacterium is resistant to methicillin. In
another exemplary embodiment, the bacterium is
methicillin-resistant Staphylococcus aureus. In another exemplary
embodiment, the bacterium is selected from the group consisting of
Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus
aureus, Mycobacterium catarrhalis, Mycobacterium pneumoniae,
Legionella pneumophila and Chlamydia pneumoniae. In another
exemplary embodiment, the bacterium is selected from the group
consisting of Enterobacter cloacae, Escherichia coli, Klebsiella
pneumoniae, Proteus mirabilis, Serratia marcescens, Citrobacter
freundii, Providencia stuartii, Pseudomonas aeruginosa,
Acinetobacter baumannii, Stenotrophomonas maltophilia, Burkholderia
cepacia, Staphylococcus aureus, Streptococcus pneumoniae,
Streptococcus pyogenes, Enterococcus faecalis, and Enterococcus
faecium. In another exemplary embodiment, the bacterium is selected
from the group consisting of Staphylococcus aureus, Staphylococcus
epidermidis, Staphylococcus haemolyticus, Streptococcus pyogenes,
Streptococcus agalactiae and Streptococcus pneumoniae.
[0145] In an exemplary embodiment, the microorganism is a
bacterium, which is selected from the group consisting of bacilli,
including Bacillus species, Corynebacterium species (also
Propionibacterium) and Clostridium species; filamentous bacteria,
including Actinomyces species and Streptomyces species; bacilli,
such as Pseudomonas species, Brucella species, Agrobacterium
species, Bordetella species, Escherichia species, Shigella species,
Yersinia species, Salmonella species, Klebsiella species,
Enterobacter species, Haemophilus species, Pasteurella species, and
Streptobacillus species; spirochetal species, Campylobacter
species, Vibrio species; and intracellular bacteria including
Rickettsiae species and Chlamydia species.
VI. b) Microorganism Infection
[0146] The compounds of the invention and/or combinations of the
invention exhibit potency against microorganisms, such as bacteria,
and therefore have the potential to be used to treat and/or prevent
a microorganism infection, such as a bacterial infection.
[0147] In a further aspect, the invention provides a method of
treating a bacterial infection comprising administering to an
animal suffering from the infection an effective amount of a
compound of the invention, or a pharmaceutically acceptable salt
thereof, thereby treating the bacterial infection. In an exemplary
embodiment, the invention provides a method of treating a bacterial
infection comprising administering to an animal suffering from the
infection an effective amount of a compound of the invention, or a
pharmaceutically acceptable salt thereof, and an effective amount
of an antibiotic, or a pharmaceutically acceptable salt thereof,
thereby treating the bacterial infection.
[0148] In a further aspect, the invention provides a method of
preventing a bacterial infection comprising administering to an
animal a prophylactic amount of a compound of the invention, or a
pharmaceutically acceptable salt thereof, thereby treating the
bacterial infection. In an exemplary embodiment, the invention
provides a method of preventing a bacterial infection comprising
administering to an animal a prophylactic amount of a compound of
the invention, or a pharmaceutically acceptable salt thereof, and
an effective amount of an antibiotic, or a pharmaceutically
acceptable salt thereof, thereby treating the bacterial
infection.
VI. c) Diseases
[0149] The compounds of the invention and/or combinations of the
invention exhibit potency against microorganisms, such as bacteria,
and therefore have the potential to achieve therapeutic efficacy in
the animals described herein.
[0150] In another aspect, the invention provides a method of
treating and/or preventing a disease. In an exemplary embodiment,
the method includes administering to the animal a therapeutically
effective amount of a compound of the invention, thereby treating
and/or preventing the disease. In an exemplary embodiment, the
method includes administering to the animal a therapeutically
effective amount of a combination of the invention, thereby
treating and/or preventing the disease. In an exemplary embodiment,
the compound of the invention or the combination of the invention
can be used in human or veterinary medical therapy, particularly in
the treatment or prophylaxis of bacterial-associated disease. In an
exemplary embodiment, the compound is described herein, or a salt,
prodrug, hydrate or solvate thereof, or a combination thereof. In
an exemplary embodiment, the invention provides a compound
described herein, or a prodrug thereof. In an exemplary embodiment,
the invention provides a compound described herein, or a salt,
hydrate or solvate thereof. In an exemplary embodiment, the
invention provides a compound described herein, or a salt thereof.
In another exemplary embodiment, the compound of the invention is a
compound described herein, or a pharmaceutically acceptable salt
thereof. In an exemplary embodiment, the compound is a compound
described herein, or a pharmaceutically acceptable salt thereof. In
an exemplary embodiment, the compound is according to a formula
described herein, or a pharmaceutically acceptable salt thereof. In
an exemplary embodiment, the compound is part of a combination
described herein. In an exemplary embodiment, the compound is part
of a pharmaceutical formulation described herein. In another
exemplary embodiment, the disease is a systemic disease. In another
exemplary embodiment, the disease is a topical disease. In an
exemplary embodiment, the animal being administered the compound is
not otherwise in need of treatment with the compound.
[0151] In an exemplary embodiment, the disease is treated through
oral administration of a compound of the invention and/or a
combination of the invention. In an exemplary embodiment, the
disease is treated through intravenous administration of a compound
of the invention and/or a combination of the invention. In an
exemplary embodiment, the disease is treated through subcutaneous
administration of a compound of the invention and/or a combination
of the invention.
[0152] Systemic Diseases
[0153] In another aspect, the invention provides a method of
treating a systemic disease. The method involves contacting an
animal with a compound of the invention and/or a combination of the
invention.
[0154] In another exemplary embodiment, the disease is associated
with a bacteria described herein. In another exemplary embodiment,
the disease is associated with infection by a Gram-positive
bacteria. In an exemplary embodiment, the disease is associated
with a Staphylococcus species. In another exemplary embodiment, the
disease is selected from the group consisting of pneumonia,
gastroenteritis, toxic shock syndrome, community acquired pneumonia
(CAP), meningitis, septic arthritis, urinary tract infection,
bacteremia, endocarditis, osteomylitis, skin and skin-structure
infection. In an exemplary embodiment, the disease is associated
with a Streptococcus species. In another exemplary embodiment, the
disease is selected from the group consisting of strep throat, skin
infections, necrotizing fasciitis, toxic shock syndrome, pneumonia,
otitis media and sinusitis. In an exemplary embodiment, the disease
is associated with an Actinomyces species. In another exemplary
embodiment, the disease is actinomycosis. In an exemplary
embodiment, the disease is associated with a Norcardia species. In
another exemplary embodiment, the disease is pneumonia. In an
exemplary embodiment, the disease is associated with a
Corynebacterium species. In another exemplary embodiment, the
disease is diphtheria. In an exemplary embodiment, the disease is
associated with a Listeria species. In another exemplary
embodiment, the disease is meningitis. In an exemplary embodiment,
the disease is associated with a Bacillus species. In another
exemplary embodiment, the disease is anthrax or food poisoning. In
an exemplary embodiment, the disease is associated with a
Clostridium species. In another exemplary embodiment, the disease
is selected from the group consisting of botulism, tetanus, gas
gangrene and diarrhea.
[0155] In an exemplary embodiment, the disease is associated with
aMycobacterium species. In an exemplary embodiment, the disease is
associated with Mycobacterium tuberculosis. In an exemplary
embodiment, the disease is associated with Mycobacterium kansasii.
In an exemplary embodiment, the disease is associated with
Mycobacterium avium-intracellulare. In another exemplary
embodiment, the disease is leprosy. In another exemplary
embodiment, the disease is tuberculosis. In another exemplary
embodiment, the disease is pulmonary tuberculosis. In another
exemplary embodiment, the disease is extrapulmonary tuberculosis.
In another exemplary embodiment, the disease is associated with
multi-drug resistant tuberculosis. In another exemplary embodiment,
the disease is associated with extensively drug resistant
tuberculosis.
[0156] In another exemplary embodiment, the disease is associated
with infection by a Gram-negative bacteria. In an exemplary
embodiment, the disease is associated with a Neisseria species. In
another exemplary embodiment, the disease is selected from the
group consisting of meningitis, gonorrhea, otitis extema and
folliculitis. In an exemplary embodiment, the disease is associated
with an Escherichia species. In another exemplary embodiment, the
disease is selected from the group consisting of diarrhea, urinary
tract infections, meningitis, sepsis and HAP. In an exemplary
embodiment, the disease is associated with a Shigella species. In
another exemplary embodiment, the disease is selected from the
group consisting of diarrhea, bacteremia, endocarditis, meningitis
and gastroenteritis. In an exemplary embodiment, the disease is
associated with a Salmonella species. In another exemplary
embodiment, the disease is selected from the group consisting of
Typhoid fever, sepsis, gastroenteritis, endocarditis, sinusitis and
meningitis. In an exemplary embodiment, the disease is associated
with a Yersinia species. In another exemplary embodiment, the
disease is selected from the group consisting of Typhoid fever,
bubonic plague, enteric fever and gastroenteritis. In an exemplary
embodiment, the disease is associated with a Klebsiella species. In
another exemplary embodiment, the disease is sepsis or urinary
tract infection. In an exemplary embodiment, the disease is
associated with a Proteus species. In another exemplary embodiment,
the disease is an urinary tract infection. In an exemplary
embodiment, the disease is associated with an Enterobacter species.
In another exemplary embodiment, the disease is a hospital-acquired
infection. In an exemplary embodiment, the disease is associated
with a Serratia species. In another exemplary embodiment, the
disease is selected from the group consisting of a urinary tract
infection, skin and skin-structure infection and pneumonia. In an
exemplary embodiment, the disease is associated with a Vibrio
species. In another exemplary embodiment, the disease is cholera or
gastroenteritis. In an exemplary embodiment, the disease is
associated with a Campylobacter species. In another exemplary
embodiment, the disease is gastroenteritis. In an exemplary
embodiment, the disease is associated with a Helicobacter species.
In another exemplary embodiment, the disease is chronic gastritis.
In an exemplary embodiment, the disease is associated with a
Pseudomonas species. In another exemplary embodiment, the disease
is selected from the group consisting of pneumonia, osteomylitis,
burn-wound infections, sepsis, UTIs, endocarditis, otitis and
corneal infections. In an exemplary embodiment, the disease is
associated with a Bacteroides species. In another exemplary
embodiment, the disease is periodontal disease or aspiration
pneumonia. In an exemplary embodiment, the disease is associated
with a Haemophilus species. In another exemplary embodiment, the
disease is selected from the group consisting of meningitis,
epiglottitis, septic arthritis, sepsis, chancroid and vaginitis. In
an exemplary embodiment, the disease is associated with a
Bordetella species. In another exemplary embodiment, the disease is
Whooping cough. In an exemplary embodiment, the disease is
associated with a Legionella species. In another exemplary
embodiment, the disease is pneumonia or pontiac fever. In an
exemplary embodiment, the disease is associated with a Francisella
species. In another exemplary embodiment, the disease is tularemia.
In an exemplary embodiment, the disease is associated with a
Brucella species. In another exemplary embodiment, the disease is
brucellosis. In an exemplary embodiment, the disease is associated
with a Pasteurella species. In another exemplary embodiment, the
disease is a skin infection. In an exemplary embodiment, the
disease is associated with a Gardnerella species. In another
exemplary embodiment, the disease is vaginitis. In an exemplary
embodiment, the disease is associated with a Spirochetes species.
In another exemplary embodiment, the disease is syphilis or Lyme
disease. In an exemplary embodiment, the disease is associated with
a Chlamydia species. In another exemplary embodiment, the disease
is chlamydia. In an exemplary embodiment, the disease is associated
with aRickettsiae species. In another exemplary embodiment, the
disease is Rocky Mountain spotted fever or typhus.
[0157] In an exemplary embodiment, the disease is associated with
Mycoplasma pneumoniae. In another exemplary embodiment, the disease
is tracheobronchitis or walking pneumonia. In an exemplary
embodiment, the disease is associated with Ureaplasma urealyticum.
In another exemplary embodiment, the disease is urethritis. In
another exemplary embodiment, the disease is pyelonephritis. In
another exemplary embodiment, the disease is an intra-abdominal
infection. In another exemplary embodiment, the disease is febrile
neutropenia. In another exemplary embodiment, the disease is a
pelvic infection. In another exemplary embodiment, the disease is
bacteraemia. In another exemplary embodiment, the disease is
septicaemia.
[0158] In an exemplary embodiment, the disease is an acute
exacerbation of chronic obstructive pulmonary disease. In an
exemplary embodiment, the disease is chronic obstructive pulmonary
disease. In an exemplary embodiment, the disease is pharyngitis. In
an exemplary embodiment, the disease is tonsillitis. In an
exemplary embodiment, the disease is Acute Exacerbation of Chronic
Bronchitis (AECB). In an exemplary embodiment, the disease is
cervicitis. In an exemplary embodiment, the disease is genital
ulcer disease.
[0159] In an exemplary embodiment, for any of the methods described
herein, the animal is selected from the group consisting of human,
cattle, deer, reindeer, goat, honey bee, pig, sheep, horse, cow,
bull, dog, guinea pig, gerbil, rabbit, cat, camel, yak, elephant,
ostrich, otter, chicken, duck, goose, guinea fowl, pigeon, swan,
and turkey. In another exemplary embodiment, for any of the methods
described herein, the animal is selected from the group consisting
of a human, cattle, goat, pig, sheep, horse, cow, bull, dog, guinea
pig, gerbil, rabbit, cat, chicken and turkey. In another exemplary
embodiment, for any of the methods described herein, the animal is
a human.
[0160] In an exemplary embodiment, for any of the methods described
herein, a compound of the invention, a combination of the
invention, a compound described herein or a pharmaceutically
acceptable salt thereof, or combination described herein, and/or a
pharmaceutical formulation described herein can be used.
VII. Pharmaceutical Formulation
[0161] In another aspect, the invention provides a pharmaceutical
formulation comprising: a) a compound of the invention; and b) a
pharmaceutically acceptable excipient. In another aspect, the
invention provides a pharmaceutical formulation comprising: a) a
combination of the invention; and b) a pharmaceutically acceptable
excipient. In an exemplary embodiment, the compound is according to
a formula described herein. In an exemplary embodiment, the
compound is according to an example described herein. In an
exemplary embodiment, the compound of the invention or combination
of the invention is a compound described herein or combination
described herein, or a pharmaceutically acceptable salt thereof. In
an exemplary embodiment, the compound of the invention is a
compound described herein.
[0162] In an exemplary embodiment, the compound of the invention is
present in the pharmaceutical formulation in an amount of between
about 0.0001% to about 60% (w/w). In an exemplary embodiment, the
amount is between about 0.01% to about 10% (w/w). In an exemplary
embodiment, the amount is between about 0.1% to about 10% (w/w). In
an exemplary embodiment, the amount is between about 0.25% to about
6% (w/w). In an exemplary embodiment, the amount is between about
0.5% to about 5% (w/w). In an exemplary embodiment, the amount is
between about 0.1% and about 1.0% (w/w). In an exemplary
embodiment, the amount is between about 1.0% and about 2.0% (w/w).
In an exemplary embodiment, the amount is between about 2.0% and
about 3.0% (w/w). In an exemplary embodiment, the amount is between
about 3.0% and about 4.0% (w/w). In an exemplary embodiment, the
amount is between about 4.0% and about 5.0% (w/w).
[0163] The pharmaceutical formulations of the invention can take a
variety of forms adapted to the chosen route of administration.
Those skilled in the art will recognize various synthetic
methodologies that may be employed to prepare non-toxic
pharmaceutical formulations incorporating the compounds described
herein. Those skilled in the art will recognize a wide variety of
non-toxic pharmaceutically acceptable solvents that may be used to
prepare solvates of the compounds of the invention, such as water,
ethanol, propylene glycol, mineral oil, vegetable oil and
dimethylsulfoxide (DMSO).
[0164] The compositions of the invention may be administered
orally, topically, parenterally, by inhalation or spray or rectally
in dosage unit formulations containing conventional non-toxic
pharmaceutically acceptable carriers, adjuvants and vehicles. It is
further understood that the best method of administration may be a
combination of methods. Oral administration in the form of a pill,
capsule, elixir, syrup, lozenge, troche, or the like is
particularly preferred. The term parenteral as used herein includes
subcutaneous injections, intradermal, intravascular (e.g.,
intravenous), intramuscular, spinal, intrathecal injection or like
injection or infusion techniques.
[0165] The pharmaceutical formulations containing compounds of the
invention are preferably in a form suitable for oral use, for
example, as tablets, troches, lozenges, aqueous or oily
suspensions, dispersible powders or granules, emulsion, hard or
soft capsules, or syrups or elixirs.
[0166] Compositions intended for oral use may be prepared according
to any method known in the art for the manufacture of
pharmaceutical formulations, and such compositions may contain one
or more agents selected from the group consisting of sweetening
agents, flavoring agents, coloring agents and preserving agents in
order to provide pharmaceutically elegant and palatable
preparations. Tablets may contain the active ingredient in
admixture with non-toxic pharmaceutically acceptable excipients
that are suitable for the manufacture of tablets. These excipients
may be for example, inert diluents, such as calcium carbonate,
sodium carbonate, lactose, calcium phosphate or sodium phosphate;
granulating and disintegrating agents, for example, corn starch, or
alginic acid; binding agents, for example starch, gelatin or
acacia; lubricating agents, for example magnesium stearate, stearic
acid or talc; and extenders and bulking agents, such as
microcrystalline cellulose. The tablets may be uncoated or they may
be coated by known techniques to delay disintegration and
absorption in the gastrointestinal tract and thereby provide a
sustained action over a longer period. For example, a time delay
material such as glyceryl monostearate or glyceryl distearate may
be employed.
[0167] Formulations for oral use may also be presented as hard
gelatin capsules wherein the active ingredient is mixed with an
inert solid diluent, for example, calcium carbonate, calcium
phosphate or kaolin, or as soft gelatin capsules wherein the active
ingredient is mixed with water or an oil medium, for example peanut
oil, liquid paraffin or olive oil.
[0168] Aqueous suspensions contain the active materials in
admixture with excipients suitable for the manufacture of aqueous
suspensions. Such excipients are suspending agents, for example
sodium carboxymethylcellulose, methylcellulose,
hydroxypropylmethylcellulose, sodium alginate,
polyvinylpyrrolidone, gum tragacanth and gum acacia; and dispersing
or wetting agents, which may be a naturally-occurring phosphatide,
for example, lecithin, or condensation products of an alkylene
oxide with fatty acids, for example polyoxyethylene stearate, or
condensation products of ethylene oxide with long chain aliphatic
alcohols, for example heptadecaethyleneoxycetanol, or condensation
products of ethylene oxide with partial esters derived from fatty
acids and a hexitol such as polyoxyethylene sorbitol monooleate, or
condensation products of ethylene oxide with partial esters derived
from fatty acids and hexitol anhydrides, for example polyethylene
sorbitan monooleate. The aqueous suspensions may also contain one
or more preservatives, for example ethyl, or n-propyl
p-hydroxybenzoate, one or more coloring agents, one or more
flavoring agents, and one or more sweetening agents, such as
sucrose or saccharin.
[0169] Oily suspensions may be formulated by suspending the active
ingredients in a vegetable oil, for example arachis oil, olive oil,
sesame oil or coconut oil, or in a mineral oil such as liquid
paraffin. The oily suspensions may contain a thickening agent, for
example beeswax, hard paraffin or cetyl alcohol. Sweetening agents
such as those set forth above, and flavoring agents may be added to
provide palatable oral preparations. These compositions may be
preserved by the addition of an anti-oxidant such as ascorbic
acid.
[0170] Dispersible powders and granules suitable for preparation of
an aqueous suspension by the addition of water provide the active
ingredient in admixture with a dispersing or wetting agent,
suspending agent and one or more preservatives. Suitable dispersing
or wetting agents and suspending agents are exemplified by those
already mentioned above. Other dispersing agents include
hydrophilic polymers, electrolytes, Tween.TM. 60 or 80, PEG,
polyvinylpyrrolidone (PVP; commercially known as Plasdone.TM.), and
the carbohydrate-based dispersing agents such as, for example,
hydroxypropylcellulose and hydroxypropylcellulose ethers (e.g.,
HPC, HPC-SL, and HPC-L), hydroxypropylmethylcellulose and
hydroxypropylmethylcellulose ethers (e.g. HPMC K100, HPMC K4M, HPMC
K15M, and HPMC K100M), carboxymethylcellulose sodium,
methylcellulose, hydroxyethylcellulose,
hydroxypropylmethylcellulose phthalate,
hydroxypropylmethylcellulose acetate stearate, noncrystalline
cellulose, magnesium aluminum silicate, triethanolamine, polyvinyl
alcohol (PVA), polyvinylpyrrolidone/vinyl acetate copolymer
(Plasdone.TM., e.g., S-630), 4-(1,1,3,3-tetramethylbutyl)-phenol
polymer with ethylene oxide and formaldehyde (also known as
tyloxapol), poloxamers (e.g., Pluronics F68.TM., F88.TM., and
F108.TM., which are block copolymers of ethylene oxide and
propylene oxide); and poloxamines (e.g., Tetronic 9080, also known
as Poloxamine 9080, which is a tetrafunctional block copolymer
derived from sequential addition of propylene oxide and ethylene
oxide to ethylenediamine (BASF Corporation, Parsippany, N.J.)).
Additional excipients, for example sweetening, flavoring and
coloring agents, may also be present.
[0171] Pharmaceutical formulations of the invention may also be in
the form of oil-in-water emulsions and water-in-oil emulsions. The
oily phase may be a vegetable oil, for example olive oil or arachis
oil, or a mineral oil, for example liquid paraffin or mixtures of
these. Suitable emulsifying agents may be naturally-occurring gums,
for example gum acacia or gum tragacanth; naturally-occurring
phosphatides, for example soy bean, lecithin, and esters or partial
esters derived from fatty acids and hexitol; anhydrides, for
example sorbitan monooleate; and condensation products of the said
partial esters with ethylene oxide, for example polyoxyethylene
sorbitan monooleate. The emulsions may also contain sweetening and
flavoring agents.
[0172] Syrups and elixirs may be formulated with sweetening agents,
for example glycerol, propylene glycol, sorbitol or sucrose. Such
formulations may also contain a demulcent, a preservative, and
flavoring and coloring agents. The pharmaceutical formulations may
be in the form of a sterile injectable aqueous or oleaginous
suspension. This suspension may be formulated according to the
known art using those suitable dispersing or wetting agents and
suspending agents, which have been mentioned above. The sterile
injectable preparation may also be a sterile injectable solution or
suspension in a non-toxic parenterally acceptable diluent or
solvent, for example as a solution in 1,3-butanediol. Among the
acceptable vehicles and solvents that may be employed are water,
Ringer's solution and isotonic sodium chloride solution. In
addition, sterile, fixed oils are conventionally employed as a
solvent or suspending medium. For this purpose any bland fixed oil
may be employed including synthetic mono- or diglycerides. In
addition, fatty acids such as oleic acid find use in the
preparation of injectables.
[0173] The composition of the invention may also be administered in
the form of suppositories, e.g., for rectal administration of the
drug. These compositions can be prepared by mixing the drug with a
suitable non-irritating excipient that is solid at ordinary
temperatures but liquid at the rectal temperature and will
therefore melt in the rectum to release the drug. Such materials
are cocoa butter and polyethylene glycols.
[0174] Alternatively, the compositions can be administered
parenterally in a sterile medium. The drug, depending on the
vehicle and concentration used, can either be suspended or
dissolved in the vehicle. Advantageously, adjuvants such as local
anesthetics, preservatives and buffering agents can be dissolved in
the vehicle.
[0175] For administration to non-human animals, the composition
containing the therapeutic compound may be added to the animal's
feed or drinking water. Also, it will be convenient to formulate
animal feed and drinking water products so that the animal takes in
an appropriate quantity of the compound in its diet. It will
further be convenient to present the compound in a composition as a
premix for addition to the feed or drinking water. The composition
can also added as a food or drink supplement for humans.
[0176] Dosage levels of the order of from about 5 mg to about 250
mg per kilogram of body weight per day and more preferably from
about 25 mg to about 150 mg per kilogram of body weight per day,
are useful in the treatment of the above-indicated conditions. The
amount of active ingredient that may be combined with the carrier
materials to produce a single dosage form will vary depending upon
the condition being treated and the particular mode of
administration. Dosage unit forms will generally contain between
from about 1 mg to about 500 mg of an active ingredient.
[0177] Frequency of dosage may also vary depending on the compound
used and the particular disease treated. However, for treatment of
most disorders, a dosage regimen of 4 times daily or less is
preferred. It will be understood, however, that the specific dose
level for any particular patient will depend upon a variety of
factors including the activity of the specific compound employed,
the age, body weight, general health, sex, diet, time of
administration, route of administration and rate of excretion, drug
combination and the severity of the particular disease undergoing
therapy.
[0178] Preferred compounds of the invention will have desirable
pharmacological properties that include, but are not limited to,
oral bioavailability, low toxicity, low serum protein binding and
desirable in vitro and in vivo half-lives. Penetration of the blood
brain barrier for compounds used to treat CNS disorders is
necessary, while low brain levels of compounds used to treat
peripheral disorders are often preferred.
[0179] The amount of the composition required for use in treatment
will vary not only with the particular compound selected but also
with the route of administration, the nature of the condition being
treated and the age and condition of the patient and will
ultimately be at the discretion of the attendant physician or
clinician.
[0180] In an exemplary embodiment, the pharmaceutical composition
described herein includes an additional active ingredient. In
another exemplary embodiment, the additional active ingredient is a
compound that has been approved for human use by the United States
Food and Drug Administration. In another exemplary embodiment, the
additional active ingredient is an immunosuppressive agent. In
still another exemplary embodiment, the additional active
ingredient is selected from the group consisting of
corticosteroids, aminosalicylates, azathioprine (6-mercaptopurine),
methotrexate and cyclosporine, etanercept, infliximab, adalimumab,
alefacept, efalizumab and anakinra.
[0181] In still another exemplary embodiment, the additional active
ingredient is selected from the group consisting of betamethasone,
tacrolimus and pimecrolimus. In still another exemplary embodiment,
the additional active ingredient is selected from the group
consisting of an activated vitamin D analog and an arotinoid (an
aromatic retinoic acid analog). In still another exemplary
embodiment, the additional active ingredient is carcipotriol, such
as Tazorac (tazarotene).
VII. A) Topical Formulations
[0182] In a preferred embodiment, the methods of the invention can
be employed through the topical application of the compounds
described herein. Topical administration includes for example,
transmucosal, transdermal, ungual and transungual routes of
administration. The topical compositions useful in the subject
invention can be made into a wide variety of product types. These
include, but are not limited to, lotions, creams, gels, sticks,
sprays, ointments, pastes, foams, mousses, masks, eye ointments,
eye or ear drops, impregnated dressings, wipes, cleansers including
soaps, body washes and shampoos, and make-up products, such as
bases, blushes, lipsticks, and eye shadows, among others. These
product types can comprise several types of carrier systems
including, but not limited to particles, nanoparticles, and
liposomes. If desired, disintegrating agents can be added, such as
the cross-linked polyvinyl pyrrolidone, agar or alginic acid or a
salt thereof such as sodium alginate. Techniques for formulation
and administration can be found in Remington: The Science and
Practice of Pharmacy, supra. The formulation can be selected to
maximize delivery to a desired target site in the body. The
formulations can also include various conventional colorants,
fragrances, thickeners, preservatives, humectants, emollients,
demulcents, solubilizing excipients, dispersants, penetration
enhancers, plasticizing agents, preservatives, stabilizers,
demulsifiers, wetting agents, sunscreens, emulsifiers,
moisturizers, astringents, deodorants, and the like, which can be
added to provide additional benefits such as, for example,
improving the feel and/or appearance of the topical
preparation.
[0183] Lotions, which are preparations that are to be applied to
the skin, nail, hair, claw or hoof surface without friction, are
typically liquid or semi-liquid preparations in which finely
divided solid, waxy, or liquid are dispersed. Lotions will
typically contain suspending agents to produce better dispersions
as well as compounds useful for localizing and holding the active
agent in contact with the skin, nail, hair, claw or hoof, e.g.,
methylcellulose, sodium carboxymethyl-cellulose, or the like.
[0184] Creams containing the active agent for delivery according to
the invention are viscous liquid or semisolid emulsions, either
oil-in-water or water-in-oil. Cream bases are water-washable, and
contain an oil phase, an emulsifier and an aqueous phase. The oil
phase is generally comprised of petrolatum or a fatty alcohol, such
as cetyl- or stearyl alcohol; the aqueous phase usually, although
not necessarily, exceeds the oil phase in volume, and generally
contains a humectant. The emulsifier in a cream formulation, as
explained in Remington: The Science and Practice of Pharmacy,
supra, is generally a nonionic, anionic, cationic or amphoteric
surfactant.
[0185] Ointments, which are semisolid preparations, are typically
based on petrolatum or other petroleum derivatives. As will be
appreciated by the ordinarily skilled artisan, the specific
ointment base to be used is one that provides for optimum delivery
for the active agent chosen for a given formulation, and,
preferably, provides for other desired characteristics as well,
e.g., emolliency or the like. As with other carriers or vehicles,
an ointment base should be inert, stable, nonirritating and
non-sensitizing. As explained in Remington: The Science and
Practice of Pharmacy, 19th Ed. (Easton, Pa.: Mack Publishing Co.,
1995), at pages 1399-1404, ointment bases may be grouped in four
classes: oleaginous bases; emulsifiable bases; emulsion bases; and
water-soluble bases. Oleaginous ointment bases include, for
example, vegetable oils, fats obtained from animals, and semisolid
hydrocarbons obtained from petroleum. Preferred water-soluble
ointment bases are prepared from polyethylene glycols of varying
molecular weight; again, reference may be had to Remington: The
Science and Practice of Pharmacy, supra, for further
information.
[0186] Useful formulations of the invention also encompass sprays
and aerosols. Sprays generally provide the active agent in an
aqueous and/or alcoholic solution which can be misted onto the
skin, nail, hair, claw or hoof for delivery. Such sprays include
those formulated to provide for concentration of the active agent
solution at the site of administration following delivery, e.g.,
the spray solution can be primarily composed of alcohol or other
like volatile liquid in which the drug or active agent can be
dissolved. Upon delivery to the skin, nail, hair, claw or hoof, the
carrier evaporates, leaving concentrated active agent at the site
of administration. Examples of aerosol technology are disclosed in
U.S. Pat. Nos. 6,682,716; 6,716,415; 6,716,417; 6,783,753;
7,029,658; and 7,033,575.
[0187] Examples of solubilizing excipients include polyethoxylated
fatty acids, PEG-fatty acid diesters, PEG-fatty acid mono-ester and
di-ester mixtures, polyethylene glycol glycerol fatty acid esters,
alcohol-oil transesterification products, polyglycerized fatty
acids, propylene glycol fatty acid esters, mixtures of propylene
glycol esters-glycerol esters, mono- and diglycerides, sterol and
sterol derivatives, polyethylene glycol sorbitan fatty acid esters,
polyethylene glycol alkyl ethers, sugar esters, polyethylene glycol
alkyl phenols, polyoxyethylene-polyoxypropylene block copolymers,
sorbitan fatty acid esters, lower alcohol fatty acid esters, ionic
surfactants, tocopherol esters, and sterol esters.
[0188] Exemplary embodiments are summarized herein below.
[0189] In an exemplary embodiment, the invention provides a
compound having a structure according to a formula which is:
##STR00023##
wherein R.sup.3 is substituted or unsubstituted nitroalkyl or
substituted or unsubstituted aminoalkyl; R.sup.4 is selected from
the group consisting of halogen, unsubstituted alkyl, unsubstituted
alkoxy, and unsubstituted phenyl; Y is O or S; and R.sup.5 is
selected from the group consisting of substituted or unsubstituted
alkyl and substituted or unsubstituted heteroalkyl; or a salt,
hydrate or solvate thereof.
[0190] In an exemplary embodiment, according to the above
paragraph, having a structure which is
##STR00024##
wherein C* is a carbon atom stereocenter which has a configuration
which is (R) or (S).
[0191] In an exemplary embodiment, according to any of the above
paragraphs, wherein C* stereocenter is in a (S) configuration.
[0192] In an exemplary embodiment, according to any of the above
paragraphs, wherein R.sup.3 is
--(CR.sup.20R.sup.21).sub.nNR.sup.22R.sup.23 in which n is 1, 2, 3,
4, 5, 6, 7, 8, 9, or 10;
each R.sup.20 and each R.sup.21 is independently selected from the
group consisting of H, R.sup.26, OR.sup.26, NR.sup.26R.sup.27,
SR.sup.26, --S(O)R.sup.26, --S(O).sub.2R.sup.26,
--S(O).sub.2NR.sup.26R.sup.27, --C(O)R.sup.27, --C(O)OR.sup.27, and
--C(O)NR.sup.26R.sup.27 R.sup.22 and R.sup.23 are independently
selected from the group consisting of H, --S(O)R.sup.28,
--S(O).sub.2R.sup.28, --S(O).sub.2NR.sup.28R.sup.29,
--C(O)R.sup.28, --C(O)OR.sup.28, --C(O)NR.sup.28R.sup.29,
substituted or unsubstituted alkyl, substituted or unsubstituted
heteroalkyl, substituted or unsubstituted cycloalkyl, substituted
or unsubstituted heterocycloalkyl, substituted or unsubstituted
aryl, and substituted or unsubstituted heteroaryl; wherein each
R.sup.26, each R.sup.27, each R.sup.28 and each R.sup.29 is
independently selected from the group consisting of H, substituted
or unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted cycloalkyl, substituted or
unsubstituted heterocycloalkyl, substituted or unsubstituted aryl,
and substituted or unsubstituted heteroaryl.
[0193] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is --CH.sub.2NH.sub.2.
[0194] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is --CH.sub.2NH.sub.2, and C* has a
configuration which is (S).
[0195] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is selected from the group consisting of
methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and
sec-butyl.
[0196] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is selected from the group consisting of
fluorine, chlorine, bromine, and iodine.
[0197] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is fluorine.
[0198] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is chlorine.
[0199] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is bromine.
[0200] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.5 is:
##STR00025##
wherein a is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; each R.sup.10 and
each R.sup.11 is independently selected from the group consisting
of H, substituted or unsubstituted alkyl, OH and NH.sub.2; R.sup.12
is selected from the group consisting of H, R.sup.7, halogen,
cyano, amidino, OR.sup.7, NR.sup.7R.sup.8, SR.sup.7,
--N(R.sup.7)S(O).sub.2R.sup.8, --C(O)R.sup.7, --C(O)OR.sup.7,
--C(O)NR.sup.7R.sup.8 wherein each R.sup.7 and each R.sup.8 is
independently selected from the group consisting of H, substituted
or unsubstituted alkyl, substituted or unsubstituted heteroalkyl,
substituted or unsubstituted cycloalkyl, substituted or
unsubstituted heterocycloalkyl, substituted or unsubstituted aryl,
and substituted or unsubstituted heteroaryl.
[0201] In an exemplary embodiment, according to any of the above
paragraphs, a is 1, 2, 3, 4, or 5.
[0202] In an exemplary embodiment, according to any of the above
paragraphs, a is 2, 3, or 4.
[0203] In an exemplary embodiment, according to any of the above
paragraphs, each R.sup.10 and each R.sup.11 is independently
selected from the group consisting of H, substituted or
unsubstituted alkyl, OH, and NH.sub.2.
[0204] In an exemplary embodiment, according to any of the above
paragraphs, each R.sup.10 and each R.sup.11 is H.
[0205] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.12 is selected from the group consisting of H,
OH, NH.sub.2, methyl, ethyl, --NHS(O).sub.2CH.sub.3, cyano,
--NHC(O)CH.sub.3, --NHC(O)NHCH.sub.2CH.sub.3, --C(O)NH.sub.2,
--C(O)OH, 4-(methoxy)phenyl, benzyl, benzoxy, --NHC(O)OCH.sub.2Ph,
--C(O)NHCH.sub.2CH.sub.2OH and --C(NH.sub.2)(NH).
[0206] In an exemplary embodiment, according to any of the above
paragraphs, Y is O.
[0207] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.5 is unsubstituted alkyl.
[0208] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.5 is selected from the group consisting of
methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, and
sec-butyl.
[0209] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is-CH.sub.2NH.sub.2; and Y is O; and R.sup.5 is
substituted or unsubstituted alkyl.
[0210] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is-CH.sub.2NH.sub.2; and R.sup.4 is
halogen.
[0211] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.4 is halogen; Y is O; and R.sup.5 is
unsubstituted alkyl.
[0212] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is --CH.sub.2NH.sub.2, and C* has a
configuration which is (S) and R.sup.4 is halogen.
[0213] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is --CH.sub.2NH.sub.2, and C* has a
configuration which is (S), R.sup.5 is unsubstituted alkyl and
R.sup.4 is halogen.
[0214] In an exemplary embodiment, according to any of the above
paragraphs, Y is O, R.sup.3 is --CH.sub.2NH.sub.2, and C* has a
configuration which is (S), R.sup.5 is unsubstituted C.sub.1 or
C.sub.2 or C.sub.3 or C.sub.4 alkyl and R.sup.4 is halogen.
[0215] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is-CH.sub.2NH.sub.2; R.sup.4 is chlorine; Y is
O; and R.sup.5 is substituted or unsubstituted alkyl.
[0216] In an exemplary embodiment, according to any of the above
paragraphs, R.sup.3 is-CH.sub.2NH.sub.2; Y is O; and R.sup.5 is
ethyl.
[0217] In an exemplary embodiment, according to any of the above
paragraphs, the compound has a structure which is
##STR00026##
[0218] In an exemplary embodiment, the invention provides a
composition comprising: (a) a) a first stereoisomer of the compound
according to any of the above paragraphs; b) at least one
additional stereoisomer of the first stereoisomer; wherein the
first stereoisomer is present in an enantiomeric excess of at least
80% relative to said at least one additional stereoisomer.
[0219] In an exemplary embodiment, according to any of the above
paragraphs, wherein said enantiomeric excess is at least 92%.
[0220] In an exemplary embodiment, according to any of the above
paragraphs, wherein the C* stereocenter of the first stereoisomer
is in a (S) configuration.
[0221] In an exemplary embodiment, according to any of the above
paragraphs, wherein R.sup.3 is --CH.sub.2NH.sub.2.
[0222] In an exemplary embodiment, the invention provides a
composition according to any of the above paragraphs, wherein the
C* stereocenter is in a (S) configuration, and said composition is
substantially free of the (R) enantiomer of the compound.
[0223] In an exemplary embodiment, the invention provides a
combination comprising the compound according to any of the above
paragraphs, or a pharmaceutically acceptable salt thereof, together
with at least one other therapeutically active agent.
[0224] In an exemplary embodiment, the invention provides a
pharmaceutical formulation comprising: (a) a compound according to
any of the above paragraphs, or a pharmaceutically acceptable salt
thereof; and (b) a pharmaceutically acceptable excipient.
[0225] In an exemplary embodiment, according to any of the above
paragraphs, the formulation is in a unit dosage form.
[0226] In an exemplary embodiment, according to any of the above
paragraphs, the formulation is for oral or topical use.
[0227] In an exemplary embodiment, the invention provides a method
of inhibiting an enzyme, comprising: contacting the enzyme with the
compound according to any of the above paragraphs, thereby
inhibiting the enzyme.
[0228] In an exemplary embodiment, according to any of the above
paragraphs, the enzyme is a t-RNA synthetase which comprises an
editing domain.
[0229] In an exemplary embodiment, according to any of the above
paragraphs, the enzyme is a leucyl t-RNA synthetase.
[0230] In an exemplary embodiment, the invention provides a method
of killing and/or preventing the growth of a microorganism,
comprising: contacting the microorganism with an effective amount
of a compound according to any of the above paragraphs, or a
pharmaceutically acceptable salt thereof, thereby killing and/or
preventing the growth of the microorganism.
[0231] In an exemplary embodiment, according to any of the above
paragraphs, the microorganism is a bacterium.
[0232] In an exemplary embodiment, according to any of the above
paragraphs, the microorganism is Mycobacterium tuberculosis.
[0233] In an exemplary embodiment, the invention provides a method
of treating and/or preventing a disease in an animal, comprising:
administering to the animal a therapeutically effective amount of
compound according to any of the above paragraphs, or a
pharmaceutically-acceptable salt thereof, thereby treating and/or
preventing the disease.
[0234] In an exemplary embodiment, according to any of the above
paragraphs, the disease is tuberculosis.
[0235] In an exemplary embodiment, according to any of the above
paragraphs, the animal is a human.
[0236] In an exemplary embodiment, the invention provides a method
of inhibiting the editing domain of a t-RNA synthetase, comprising:
contacting the synthetase with an effective amount of a compound
according to any of the above paragraphs, or a
pharmaceutically-acceptable salt thereof, thereby inhibiting the
synthetase.
[0237] In an exemplary embodiment, according to any of the above
paragraphs, the synthetase is a leucyl t-RNA synthetase.
[0238] In an exemplary embodiment, according to any of the above
paragraphs, the synthetase is a Mycobacterium tuberculosis leucyl
t-RNA synthetase.
[0239] In an exemplary embodiment, the invention provides the use
of a compound according to any of the above paragraphs or a
combination according to any of the above paragraphs or a
pharmaceutically acceptable salt thereof, in the manufacture of a
medicament for the treatment and/or prophylaxis of bacterial
infection.
[0240] It is to be understood that the present invention covers all
combinations of aspects and/or embodiments, as well as suitable,
convenient and preferred groups described herein.
[0241] The invention is further illustrated by the Examples that
follow. The Examples are not intended to define or limit the scope
of the invention.
EXAMPLES
[0242] Proton NMR are recorded on Varian AS 300 spectrometer and
chemical shifts are reported as .delta. (ppm) down field from
tetramethylsilane. Mass spectra are determined on Micromass Quattro
II.
[0243] The M. tuberculosis LeuRS gene (DNA sequence listed herein)
was prepared by GenScript and cloned into the T7 expression vector
pET28a(+) at the NdeI-XhoI sites. Over-expression of M.
tuberculosis LeuRS from this construct generated a version of M.
tuberculosis LeuRS with an N-terminal his-tag, which will use the
standard procedures for purification of his-tagged proteins.
Example 1
A.
3-Aminomethyl-4-fluoro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,2]oxaborol-1-
-ol; bis trifluoroacetic salt
6-Fluoro-2, 3-dimethoxy-benzaldehyde
##STR00027##
[0245] To a solution of 4-fluoro-1,2-dimethoxy-benzene (20.0 g,
128.07 mmol) in anhydrous THF (200 mL) under nitrogen at
-78.degree. C. was added dropwise a 2.5M solution in hexane of
n-BuLi (102.4 mL, 256.14 mmol) for duration 30 min and the reaction
mixture was further stirred at the same temperature for 3 h. The
reaction mixture was quenched carefully with DMF (100 mL) at
-65.degree. C. to -40.degree. C. and left overnight. 2N HCl (300
mL) was added dropwise at -60.degree. C. and the mixture stirred
for 30 min. The two layers were separated and the aqueous layer
extracted with EtOAc. The combined organic layers were washed with
water and brine, dried over MgSO.sub.4, filtered and concentrated
in vacuo. Purification was completed by flash column chromatography
(20% EtOAc/hexane). Yield 18.0 g (85%). .sup.1H NMR (400 MHz,
CHLOROFORM-d) .delta. ppm 10.39 (s, 1H), 7.09 (dd, J=9.4, 5.1 Hz,
1H), 6.84 (t, J=9.6 Hz, 1H), 3.98 (s, 3H), 3.88 (s, 3H). .sup.19F
NMR (376 MHz, CHLOROFORM-d): -126 ppm.
6-Fluoro-2, 3-dihydroxy-benzaldehyde
##STR00028##
[0247] To a solution of 6-fluoro-2,3-dimethoxy-benzaldehyde (18.0
g, 97.74 mmol) in anhydrous dichloromethane (100 mL) under nitrogen
at -60.degree. C. was added dropwise BBr.sub.3.OEt.sub.2 (195 mL,
195.48 mmol) over 30 min and the solution was allowed to warm up to
r.t and stirred for 4 h. The reaction mixture was cooled to
-60.degree. C. and 2N HCl (250 mL) was carefully added drop wise.
The mixture was stirred at r.t overnight. The two layers were
separated and the aqueous layer extracted with DCM. The combined
organic layers were washed with water, sat. NaHCO.sub.3 solution,
water and brine and dried over MgSO.sub.4. The solvent was removed
in vacuo to provide the title compound which was used in the next
step without further purification. Yield 11.15 g (74%). .sup.1H NMR
(400 MHz, CHLOROFORM-d) .delta. ppm 11.53 (s, 1H), 10.23 (s, 1H),
7.11 (dd, J=8.8, 5.3 Hz, 1H), 6.57 (t, J=9.6 Hz, 1H), 5.48 (s, 1H).
.sup.19F NMR (376 MHz, CHLOROFORM-d): -132 ppm.
3-(3-Benzyloxy-propoxy)-6-fluoro-2-hydroxy-benzaldehyde
##STR00029##
[0249] To a solution of 6-fluoro-2,3-dihydroxy-benzaldehyde (11.15
g, 71.42 mmol) in anhydrous DMSO (88 mL) under nitrogen were added
sequentially sodium t-butoxide (13.72 g, 142.84 mmol) and
benzyl-3-bromopropyl ether (17.29 g, 78.56 mmol) and the reaction
mixture stirred at rt for 18 h. The mixture was diluted with water
(400 mL) and extracted with EtOAc (4.times.100 mL). The combined
organic layer was washed with water and brine, dried over
MgSO.sub.4, filtered and concentrated in vacuo. Purification was
completed by flash chromatography (20% EtOAc/hexane). Yield 11.5 g
(77%). H NMR (400 MHz, CHLOROFORM-d) .delta. ppm 11.62 (s, 1H),
10.23 (s, 1H), 7.42-7.16 (m, 5H), 7.05 (dd, J=8.8, 5.3 Hz, 1H),
6.54 (t, J=9.6 Hz, 1H), 4.52 (s, 2H), 4.13 (t, J=6.3 Hz, 2H), 3.68
(t, J=6.1 Hz, 2H), 2.12 (quin, J=6.2 Hz, 2H). .sup.19F NMR (376
MHz, CHLOROFORM-d): -132 ppm.
Trifluoro-methanesulfonic acid
6-(3-benzyloxy-propoxy)-3-fluoro-2-formyl-phenyl ester
##STR00030##
[0251] Trifluoromethanesulfonic anhydride (1.11 g, 3.94 mmol) was
added dropwise to a solution of pyridine (389 mg, 4.92 mmol) and
3-(3-benzyloxy-propoxy)-6-fluoro-2-hydroxy-benzaldehyde (1 g, 3.28
mmol) in CH.sub.2Cl.sub.2 (5 mL) at 0.degree. C. (bath temp). The
reaction mixture was then allowed to warm to rt and was stirred
until complete consumption of starting material (as determined by
TLC). Et.sub.2O and 2 N HCl were then added. The organic layer was
separated and washed with sat. NaHCO.sub.3 then brine. The organic
layer was dried (Na.sub.2SO.sub.4) and filtered through a short
silica gel plug, washing with Et.sub.2O. The filtrate was
concentrated in vacuo to give 1.10 g of the desired triflate (yield
76%) that was used directly without further purification. .sup.1H
NMR (400 MHz, CHLOROFORM-d) .delta. ppm 10.32 (s, 1H), 7.44-7.21
(m, 7H), 4.51 (s, 2H), 4.18 (t, J=6.1 Hz, 2H), 3.68 (t, J=5.7 Hz,
2H), 2.14 (quin, J=5.8 Hz, 2H).
3-(3-Benzyloxy-propoxy)-6-fluoro-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
##STR00031##
[0253] A solution of trifluoro-methanesulfonic acid
6-(3-benzyloxy-propoxy)-3-fluoro-2-formyl-phenyl ester (1.092 g,
2.50 mmol) in anhydrous 1,4-dioxane (10 mL) was added
bis(pinacolato)diborane (953 mg, 3.75 mmol) and KOAc (736 mg, 7.50
mmol) at rt, then degassed with N.sub.2 for 20 min.
PdCl.sub.2(dppf) (46 mg, 8 mol %) was added and the resulting
solution was stirred at 100.degree. C. until the reaction was
complete. The solution was cooled to rt, filtered through
Celite.RTM. or silica gel and concentrated in vacuo. The residue
was taken up in EtOAc. The organic layer was then washed with
H.sub.2O then brine, dried (Na.sub.2SO.sub.4), filtered, and
concentrated in vacuo. The product was purified by flash
chromatography (20% EtOAc/hexane) to give 0.5 g of the title
compound along with detriflated by product ratio .about.1:1 by H
NMR spectrum. .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta. ppm 10.33
(s, 1H), 7.41-7.23 (m, 6H), 7.19 (d, J=9.4 Hz, 1H), 4.57-4.42 (m,
2H), 4.06 (t, J=6.3 Hz, 2H), 3.69-3.64 (m, 2H), 2.20-2.00 (m, 2H),
1.44 (s, 12H); .sup.19F NMR (376 MHz, CHLOROFORM-d) .delta. ppm
-73.72.
7-(3-Benzyloxy-propoxy)-4-fluoro-3-nitromethyl-3H-benzo[C][1,2]oxaborol-1--
ol
##STR00032##
[0255] NaOH (48 mg, 1.20 mmol) was added to
3-(3-benzyloxy-propoxy)-6-fluoro-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborol-
an-2-yl)-benzaldehyde (500 mg, 1.2 mmol) in H.sub.2O (3 mL) at rt,
and the reaction mixture was stirred at rt for 5 min. MeNO.sub.2
(219 mg, 3.6 mmol) was added dropwise and the mixture was stirred
at rt for 16 h. The reaction mixture was acidified with 2 N HCl and
extracted with EtOAc. The organic fraction was washed with H.sub.2O
then brine, dried (MgSO.sub.4), and concentrated in vacuo.
Purification was accomplished by flash chromatography (10-40%
EtOAc/hexane) to give 120 mg of the title compound by 1H NMR
spectrum. .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta. ppm 7.40-7.22
(m, 5H), 7.11 (t, J=8.8 Hz, 1H), 6.81 (t, J=8.6 Hz, 1H), 5.93 (dd,
J=9.0, 2.3 Hz, 1H), 4.99 (dd, J=13.1, 2.5 Hz, 1H), 4.62-4.53 (m,
2H), 4.43 (dd, J=12.9, 9.0 Hz, 1H), 4.19-4.01 (m, 2H), 3.66 (dt,
J=15.9, 5.7 Hz, 2H), 2.18-1.94 (m, 2H); .sup.19F NMR (376 MHz,
CHLOROFORM-d) .delta. ppm -72.81 (s, 1F); MS (ESI) m/z=374 (M-1,
negative).
3-Aminomethyl-4-fluoro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,2]oxaborol-1-ol
bis-trifluoroacetic acid salt
##STR00033##
[0257] A mixture of
7-(3-benzyloxy-propoxy)-4-fluoro-3-nitromethyl-3H-benzo[C][1,2]oxaborol-1-
-ol (120 mg, 0.32 mmol) and 20% Pd(OH).sub.2 (120 mg, 1:1 w/w
substrate to catalyst) in AcOH (10 mL) was shaken under an
atmosphere of H.sub.2 (45-50 psi) in a Parr shaker. Once the
reaction was complete, the mixture was filtered through
Celite.RTM.. The filtrate was concentrated in vacuo to give a gummy
material. Remaining AcOH was removed by co-evaporation with toluene
(3.times.) to give the amine. Purification by preparative HPLC
(0.1% aq CF.sub.3CO.sub.2H/CH.sub.3CN) produced 12 mg of the title
compound as a white solid (yield 8.4%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. ppm 9.14 (br. s., 1H), 8.02 (br. s., 3H),
7.25 (d, J=7.8 Hz, 1H), 6.93 (d, J=7.4 Hz, 1H), 5.43 (br. s., 1H),
4.55 (br. s., 1H), 4.07 (br. s., 2H), 3.56 (br. s., 2H), 3.42-3.37
(m, 1H), 2.95 (br. s., 1H), 1.86 (br. s., 2H); .sup.19F NMR (376
MHz, DMSO-d.sub.6) .delta. ppm -73.90 (s, 6F), -131.51 (s, 1F); MS
(ESI) m/z=256 (M+1, positive); HPLC purity: 95.65% (MaxPlot 200-400
nm), 96.63% (220 nm).
B.
3-(Aminomethyl)-4-chloro-7-(3-hydroxypropoxy)benzo[c][1,2]oxaborol-1(3H-
)-ol hydrochloride
3-(3-Benzyloxy-propoxy)-2-hydroxy-benzaldehyde
##STR00034##
[0259] NaH (2.95 g, 72.4 mmol) was added to an ice-cold solution of
2,3-dihydroxybenzaldehyde (5.0 g, 36 mmol) in anhydrous DMSO (45
mL). Benzyl-3-bromopropyl ether (6.45 mL, 36.2 mmol) was then added
and the mixture was stirred at rt for 12 h. The mixture was
neutralized using 1 N HCl and then extracted with EtOAc. The
organic fraction was washed with H.sub.2O and concentrated in
vacuo. The residue was purified by flash chromatography (8:2
hexane/EtOAc) to give the title compound as a brown oil: yield 8.40
g (81%). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. (ppm): 9.93 (s,
1H), 7.36-7.23 (m, 6H), 7.20-7.16 (m, 2H), 6.98-6.91 (m, 1H), 4.53
(s, 2H), 4.19 (t, J=6.2 Hz, 2H), 3.70 (t, J=6.1 Hz, 2H), 2.19-2.16
(m, 2H).
3-(3-Benzyloxy-propoxy)-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-b-
enzaldehyde
##STR00035##
[0261] Trifluoromethanesulfonic anhydride (4.60 mL, 27.9 mmol) was
added dropwise to a solution of pyridine (3.42 mL, 42.5 mmol) and
3-(3-benzyloxy-propoxy)-2-hydroxy-benzaldehyde (7.6 g, 26 mmol) in
CH.sub.2Cl.sub.2 (200 mL) at 0.degree. C. (bath temp). The reaction
mixture was then allowed to warm to rt and was stirred until
complete consumption of starting material (as determined by TLC).
Et.sub.2O and 2 N HCl were then added. The organic layer was
separated and washed with sat. NaHCO.sub.3 then brine. The organic
layer was dried (Na.sub.2SO.sub.4) and filtered through a short
silica gel plug, washing with Et.sub.2O. The filtrate was
concentrated in vacuo to give 8.60 g of the desired triflate (yield
77%) that was used directly without further purification. .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. (ppm): 10.23 (s, 1H), 7.54-7.47
(m, 1H), 7.43 (t, J=8.0 Hz, 1H), 7.36-7.22 (m, 6H), 4.52 (s, 2H),
4.23 (t, J=6.3 Hz, 2H), 3.71 (t, J=6.1 Hz, 2H), 2.21-2.17 (m,
2H).
[0262] A solution of trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-6-formyl-phenyl ester (8.0 g, 19 mmol) in
anhydrous 1,4-dioxane (160 mL) was added bis(pinacolato)diborane
(9.71 g, 38.2 mmol) and KOAc (5.71 g, 57.4 mmol) at rt, then
degassed with N.sub.2 for 20 min. PdCl.sub.2(dppf) (1.39 g, 1.89
mmol) was added and the resulting solution was stirred at
100.degree. C. until the reaction was complete. The solution was
cooled to rt, filtered through Celite.RTM. or silica gel and
concentrated in vacuo. The residue was taken up in EtOAc. The
organic layer was then washed with H.sub.2O then brine, dried
(Na.sub.2SO.sub.4), filtered, and concentrated in vacuo. The
product was purified by flash chromatography (9:1 hexane/EtOAc) to
give 4.80 g of the title compound (yield 43%) along with some
pinacol contamination and was used without further purification.
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. (ppm): 9.93 (s, 1H), 7.46
(t, J=7.8 Hz, 1H), 7.41-7.36 (m, 1H), 7.35-7.24 (m, 5H), 7.08 (d,
J=7.8 Hz, 1H), 4.50 (s, 2H), 4.10 (t, J=6.3 Hz, 2H), 3.67 (t, J=6.3
Hz, 2H), 2.11 (quin, J=6.2 Hz, 2H), 1.43 (s, 12H).
7-(3-Benzyloxy-propoxy)-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
##STR00036##
[0264] NaOH aq. (NaOH (3.64 g, 83 mmol) was added to
3-(3-benzyloxy-propoxy)-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)--
benzaldehyde (36 g, 91 mmol) in H.sub.2O (180 mL), and THF (50 mL)
at rt, and the reaction mixture was stirred at rt for 5 min.
MeNO.sub.2 (16.6 g, 273 mmol) was added dropwise and the mixture
was stirred at rt for 16 h. The reaction mixture was acidified with
2 N HCl and extracted with EtOAc. The organic fraction was washed
with H.sub.2O then brine, dried (MgSO.sub.4), and concentrated in
vacuo. Purification was accomplished by flash chromatography (1:1
hexane/EtOAc) to give 15.9 g of the title compound as a light
yellow oil (yield 50%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
.quadrature..quadrature.ppm: 9.05 (s, 1H), 7.44 (t, J=7.8 Hz, 1H),
7.35-7.20 (m, 5H), 7.06 (d, J=7.4 Hz, 1H), 6.88 (d, J=8.2 Hz, 1H),
5.70 (dd, J=9.4, 2.3 Hz, 1H), 5.29 (dd, J=13.7, 2.7 Hz, 1H), 4.53
(dd, J=13.3, 9.4 Hz, 1H), 4.45 (s, 2H), 4.11 (t, J=6.1 Hz, 2H),
3.60 (t, J=6.3 Hz, 2H), 2.04-1.91 (m, 2H); MS (ESI): m/z=356 (M-1,
negative); HPLC purity: 99.35% (MaxPlot 200-400 nm), 97.32% (220
nm).
4-Chloro-7-(3-hydroxy-propoxy)-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-ol
##STR00037##
[0266] To
7-(3-benzyloxy-propoxy)-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-
-ol (1.1 g, 3.0 mmol) in glacial AcOH (10 mL) in cold water bath
was added SO.sub.2Cl.sub.2 (0.7 mE, 9.07 mmol) dropwise over 5
minutes period. The resulting solution was stirred for 30 minutes
at the same temperature then 1.5 h at room temperature. The
solution was quenched with crushed ice and then diluted with EtOAc
(100 mL). The organic layer was washed with water, dried over
Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure.
To the crude residue in MeOH (20 mL) was added Pd(OH).sub.2 (10%
w/w on carbon, 0.7 g), conc HCl was added until pH was 1, and the
reaction vessel was pressurized to 40 psi with hydrogen for 30
minutes at room temperature. The resulting mixture was filtered
through a pad of Celite.RTM. and washed with EtOAc. The filtrate
was concentrated in vacuo, then the residue was purified by silica
gel column chromatography (EtOAc:Hex, 1:1) providing the title
compound (0.2 g, 24% in 2 steps). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. ppm 9.31 (br. s., 1H), 7.49 (d, J=8.2 Hz,
1H), 6.98 (d, J=8.6 Hz, 1H), 5.76 (dd, J=8.2, 2.7 Hz, 1H), 5.33
(dd, J=13.2, 2.3 Hz, 1H), 4.70 (dd, J=13.0, 8.4 Hz, 1H), 4.55 (br.
s., 1H), 4.15-4.05 (m, 2H), 3.61-3.55 (m, 2H), 1.95-1.77 (m,
2H).
3-Aminomethyl-4-chloro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,
2]oxaborol-1-ol hydrogen chloride
##STR00038##
[0268]
4-Chloro-7-(3-hydroxy-propoxy)-3-nitromethyl-3H-benzo[c][1,2]oxabor-
ol-1-ol (105 mg, 0.35 mmol) in methanolic ammonia solution (2 M, 20
mL) was added Ra/Ni (0.15 g, 2800 Nickel slurry in water) and the
reaction vessel was pressurized to 40 psi with hydrogen overnight
at room temperature. The resultant mixture was filtered through a
pad of Celite.RTM. and washed with EtOAc. The filtrate was
concentrated in vacuo and the residue was added water (1 mL),
followed by conc HCl to pH 1. The heterogeneous mixture was
lyophilized provide the title compound (130 mg, quantitative) as a
white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. ppm 9.14
(br. s., 1H), 8.36 (br. s., 3H), 7.49 (d, J=8.2 Hz, 1H), 6.99 (d,
J=8.2 Hz, 1H), 5.40 (d, J=7.0 Hz, 1H), 4.40 (br. s., 1H), 4.10 (br.
s., 2H), 3.59 (br. s., 2H), -3.30 (hidden, 1H), 2.89 (br. s., 1H),
1.89 (br.s, 2H); MS (ESI) m/z=272 (M+1, positive); HPLC purity:
96.92% (MaxPlot 200-400 nm), 97.96% (220 nm).
C. 3-Aminomethyl-7-ethoxy-4-fluoro-3H-benzo[c][1,2]-oxaborol-1-ol;
hydrochloride
6-Fluoro-2, 3-dimethoxy-benzaldehyde
##STR00039##
[0270] To a cold (-78.degree. C.) solution of
4-fluoro-1,2-dimethoxybenzene (15.00 g, 96.05 mmol) in anhydrous
THF (150 mL) was added n-BuLi (84.5 mL, 211.32 mmol, 2.5 M solution
in hexanes) under nitrogen and stirred it for 3 h at -78.degree. C.
Quenched the reaction with DMF (75 mL) at -65.degree. C., added 2N
HCl (300 mL) dropwise and further stirred for 30 min. Two layers
separation was observed. Aqueous layer was extracted with EtOAc.
Combined organic layers were washed with water, brine and dried
over MgSO.sub.4. Filtered the ethyl acetate layer and concentrated
it in vacuo. The title compound was purified by flash column
chromatography using pure hexanes then 10 and 20% EtOAc in hexanes
which provided 14.40 g (78.19 mmol, 82%) of the title compound as a
white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. ppm 10.23
(s, 1H), 7.36 (dd, J=9.0, 5.1 Hz, 1H), 7.03 (t, J=9.6 Hz, 1H), 3.87
(s, 3H), 3.83 (s, 3H); .sup.19F NMR (376 MHz, DMSO-d.sub.6) .delta.
ppm -131.68--131.66 (m, 1F).
6-Fluoro-2, 3-dihydroxy-benzaldehyde
##STR00040##
[0272] To a cold (-78.degree. C.) solution of
6-fluoro-2,3-dimethoxy-benzaldehyde (4.50 g, 24.43 mmol) in
anhydrous dichloromethane (30 mL) was added BBr.sub.3 (1M in DCM,
48.8 mL, 48.87 mmol) dropwise (duration 30 min). The reaction was
warmed to room temperature and stirred for 4 h. Again cooled it to
-78.degree. C. and added 2N HCl (60 mL) to it dropwise. The
reaction was stirred for overnight at room temperature and
extracted with DCM. Combined organic layers were washed with water,
sat. NaHCO.sub.3 solution, brine and dried over MgSO.sub.4.
Filtration and removal of solvent provided 2.42 g (15.50 mmol, 64%)
of the title compound as a yellow solid. This was used in the next
step without further purification. .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. ppm 10.22 (s, 1H), 7.04 (dd, J=8.6, 5.5 Hz,
1H), 6.61 (dd, J=10.4, 8.8 Hz, 1H); .sup.19F NMR (376 MHz,
DMSO-d.sub.6) .delta. ppm -131.69--131.65 (m, 1F).
3-Ethoxy-6-fluoro-2-hydroxy-benzaldehyde
##STR00041##
[0274] To a solution of 6-fluoro-2,3-dihydroxy-benzaldehyde (1.00
g, 6.40 mmol) in dry DMSO (10 mL) was added NaOBu.sup.t(1.23 g,
12.81 mmol) and ethyl bromide (0.77 g, 7.04 mmol) under N.sub.2 and
stirred at RT for 18 h. The resultant mixture was diluted with
water, acidified to pH .about.6 with 2N.HCl and extracted with
EtOAc (4.times.25 mL). Combined organic layers were washed with
water and brine and dried over MgSO.sub.4. Filtration and removal
of the solvent under reduced pressure provided 1.05 g (5.70 mmol,
89%) of the title compound as yellow solid. .sup.1H NMR (400 MHz,
CHLOROFORM-d) .delta. ppm 10.24 (s, 1H), 7.04 (dd, J=8.8, 5.3 Hz,
1H), 6.61-6.50 (m, 1H), 4.09 (q, J=7.0 Hz, 3H), 1.46 (t, J=7.0 Hz,
3H); .sup.19F NMR (376 MHz, CHLOROFORM-d) .delta. ppm
-132.19--132.15 (m, 1F).
Trifluoro-methanesulfonic acid 6-ethoxy-3-fluoro-2-formyl-phenyl
ester
##STR00042##
[0276] To a cold (0.degree. C.) solution of
3-ethoxy-6-fluoro-2-hydroxy-benzaldehyde (1.05 g, 5.70 mmol) in dry
DCM (90 ml) was added pyridine (675 mg, 8.53 mmol) under nitrogen
and stirred the reaction mixture at 0.degree. C. for 10 min. Then
added triflic anhydride (1.93 g, 6.84 mmol) slowly and continued
stirring for 3 h at RT. Diluted the reaction with 1N.HCl (25 mL)
and extracted with DCM (2.times.100 mL). The organic layer was
washed with water and brine and dried over MgSO.sub.4. Filtered and
concentrated the filtrate. Purification of the residue by flash
column chromatography with 5% ethyl acetate in hexanes gave 1.12 g,
(3.54 mmol, 62%) of the title compound as a white solid. .sup.1H
NMR (400 MHz, CHLOROFORM-d) .delta. ppm 10.34 (s, 1H), 7.31-7.13
(m, 2H), 4.13 (q, J=7.0 Hz, 2H), 1.48 (t, J=7.0 Hz, 3H); .sup.19F
NMR (376 MHz, CHLOROFORM-d) .delta. ppm -128.15--128.11 (m, 1F),
-73.56 (s, 3F).
3-Ethoxy-6-fluoro-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
##STR00043##
[0278] A solution of trifluoro-methanesulfonic acid
6-ethoxy-3-fluoro-2-formyl-phenyl ester (1.60 g, 5.05 mmol) in
anhydrous THF (50 mL) was degassed for 40 min. Added
bis(pinacolato)diborane (3.85 g, 15.17 mmol), KOAc (1.50 g, 15.17
mmol) and PdCl.sub.2(dppf) (296 mg, 8 mol %) and stirred the
reaction at 70.degree. C. (bath temp) for 3 h. Another addition of
bis(pinacolato)diborane (1.40 g, 5.51 mmol) and heating at
70.degree. C. for 2 h completed the reaction. The resultant mixture
was cooled to room temperature and filtered through a pad of
Celite.RTM.. Concentrated the filtrate. Purification of the residue
by flash column chromatography with hexanes and 5% EtOAc/hexanes
yielded 1.65 g of title compound as white solid. .sup.1H NMR
confirms presence of title compound but with some impurities. It
was used in next step without further purification. .sup.1H NMR
(400 MHz, CHLOROFORM-d) .delta. ppm 10.29 (s, 1H), 7.12-6.97 (m,
2H), 4.00 (q, J=7.0 Hz, 2H), 1.46 (s, 12H), 1.41 (t, J=6.8 Hz, 3H);
.sup.19F NMR (376 MHz, CHLOROFORM-d) .delta. ppm -133.71--133.67
(m, 1F).
7-Ethoxy-4-fluoro-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-ol
##STR00044##
[0280]
3-Ethoxy-6-fluoro-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)--
benzaldehyde (1.65 g, 5.61 mmol) was added to a solution of NaOH
(225 mg, 5.60 mmol) in H.sub.2O (10 mL) and stirred for 10 min at
RT. Added nitromethane (1.03 g, 16.83 mmol) dropwise and stirred
for 4 h at RT. The reaction mixture was acidified with 4N HCl and
extracted with ethyl acetate. Organic layer was washed with water,
brine and dried over MgSO.sub.4, filtered and concentrated in
vacuo. Purification of the residue by flash column chromatography
with 10 to 40% EtOAc/hexanes afforded 1.7 g of a mixture of
compounds by .sup.1H NMR spectrum. This mixture was used in next
step without further purification. .sup.1H NMR (400 MHz,
CHLOROFORM-d) .delta. ppm 7.13 (t, J=8.8 Hz, 1H), 6.79 (dd, J=8.8,
2.5 Hz, 1H), 5.95 (d, J=9.0 Hz, 1H), 5.14 (s, 1H), 5.01 (dd,
J=13.3, 2.3 Hz, 1 H), 4.44 (dd, J=13.3, 9.0 Hz, 1H), 4.10 (q, J=6.8
Hz, 2H), 1.45 (t, J=6.8 Hz, 3H); .sup.19F NMR (376 MHz,
CHLOROFORM-d) .delta. ppm -131.00--130.97 (m, 1F).
7-Ethoxy-4-fluoro-1-hydroxy-1, 3-dihydro-benzo[c][1,
2]oxaborol-3-ylmethyl)-carbamic acid tert-butyl ester
##STR00045##
[0282] To a cold (0.degree. C.) solution of
7-ethoxy-4-fluoro-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-ol (500
mg, 1.96 mmol) in dry MeOH (20 mL) was added (Boc).sub.2O (856 mg,
3.92 mmol) followed by NiCl.sub.2. 6H.sub.2O (466 mg, 1.96 mmol)
under nitrogen. Stirred the reaction mixture under nitrogen for 20
min and added NaBH.sub.4 (445 mg, 11.76 mmol) in portions and left
it for overnight at RT. Evaporated the solvent and diluted the
reaction with 30 ml of ethyl acetate and filtered it through
Celite. Filtrate was concentrated and residue was purified by flash
column chromatography using 5% MeOH/DCM, but a mixture (950 mg) of
products was obtained which was used in next step without further
purification.
3-Aminomethyl-7-ethoxy-4-fluoro-3H-benzo[c][1,2]oxaborol-1-ol;
hydrochloride
##STR00046##
[0284] A solution of
7-ethoxy-4-fluoro-1-hydroxy-1,3-dihydro-benzo[c][1,2]oxaborol-3-ylmethyl)-
-carbamic acid tert-butyl ester (450 mg, 1.38 mmol) in 4M HCl (in
1,4-dioxane, 15 mL) was stirred at RT for overnight. The solvent
was removed under reduced pressure. Recrystallization from
EtOAc/hexanes provided 245 mg (0.93 mmol, 62%) of the title
compound as white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. ppm 9.15 (br. s., 1H), 8.20 (br. s., 3H), 7.26 (t, J=9.0
Hz, 1H), 6.95-6.85 (m, 1H), 5.45 (d, J=6.3 Hz, 1H), 3.30 (hidden,
1H), 4.05 (q, J=6.9 Hz, 2H), 2.89 (br. s., 1H), 1.30 (t, J=6.8 Hz,
3H); .sup.19F NMR (376 MHz, DMSO-d.sub.6) .delta. ppm
-131.68--131.66 (m, 1F); MS (ESI) m/z=226 (M+1, positive); HPLC
purity: 91.87% (MaxPlot 200-400 nm), 90.33% (220 nm); Anal. Calcd
for C.sub.10H.sub.14BClFNO.sub.3 0.5 H.sub.2O: C 44.40%; H 5.59%; N
5.18%. Found: C 44.30%; H 5.42%; N 5.50%.
D.
3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
2-Bromo-3-hydroxybenzaldehyde
##STR00047##
[0286] The suspension of 3-hydroxybenzaldehyde (5 g, 0.04 mol),
iron powder (172 mg, 3 mmol) and sodium acetate (6.72 g, 0.08 mol)
in acetic acid (40 mL) was warmed until a clear solution was
obtained and then cooled to room temperature. To this mixture was
dropwise added a solution of bromine in glacial acetic acid (10 mL)
over 15 min. After the addition, the reaction mixture was stirred
for 2 h and then poured into ice-water. The resulting mixture was
extracted with dichloromethane (3.times.50 mL). The combined
extracts were dried over anhydrous Na.sub.2SO.sub.4 and
concentrated. The residue was recrystallized from dichloromethane
to afford the product (2.3 g, yield 28%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6.quadrature..quadrature. .delta..quadrature.10.30 (s,
1H), 7.54-7.51 (m, 1H), 7.39-7.35 (m, 1H), 7.31-7.27 (m, 1H), 5.90
(s, 1H).
2-Bromo-3-ethoxybenzaldehyde
##STR00048##
[0288] The suspension of 2-bromo-3-hydroxybenzaldehyde (120 g, 0.60
mol), K.sub.2CO.sub.3 (247 g, 1.79 mol) and bromoethane (135 mL,
1.79 mol) in DMF (700 mL) was stirred at 70.degree. C. for 3 h.
After the reaction was quenched with water (50 mL), the resulting
mixture was extracted with EtOAc (3.times.60 mL). The combined
organic layers were washed with water (50 mL) and aqueous LiCl
solution (50 mL), dried over anhydrous Na.sub.2SO.sub.4 and
concentrated to dryness in vacuo. The residue was purified by
column chromatography on silica gel to give the target compound
(128 g, yield 94%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6.quadrature..quadrature..delta. .quadrature.10.45 (s,
1H), 7.52-7.50 (d, 1H), 7.38-7.34 (t, 1H), 7.13-7.10 (d, 1H),
4.18-4.13 (m, 2H), 1.53-1.50 (m, 3H).
2-(2-Bromo-3-ethoxyphenyl)-1, 3-dioxolane
##STR00049##
[0290] To a solution of 2-bromo-3-ethoxybenzaldehyde (128 g, 0.56
mol) and glycol (253 mL, 4.49 mol) in toluene (600 mL) was added
p-toluenesulfonic acid (10 g, 0.06 mol). The reaction flask had a
Dean and Stark condenser attached and the reaction mixture was
refluxed to remove the water for 4 h. The reaction mixture was then
cooled to room temperature and concentrated in vacuo. The residue
was purified by column chromatography on silica gel to give the
target compound (132 g, yield 86%).
Diisopropyl 2-(1, 3-dioxolan-2-yl)-6-ethoxyphenylboronate
##STR00050##
[0292] To the solution of 2-(2-bromo-3-ethoxyphenyl)-1,3-dioxolane
(132 g, 0.48 mol) in anhydrous THF (500 mL) was dropwise added
n-BuLi (2.5 M in THF, 386 mL, 0.97 mol) at -78.degree. C. under
nitrogen protection. The mixture was stirred at -78.degree. C. for
2 h and then triisopropyl borate (227 mL, 0.97 mol) was dropwise
added. The resulting mixture was stirred at this temperature for 4
h. After the reaction was quenched by adding saturated aqueous
NH.sub.4Cl solution (200 mL), the resulting mixture was extracted
with EtOAc (3.times.300 mL). The combined organic layers were
washed with water (200 mL) and brine (200 mL), dried over anhydrous
Na.sub.2SO.sub.4 and concentrated to dryness. The residue was
purified by column chromatography on silica gel to give the target
compound (136 g, yield 87%).
2-Ethoxy-6-formylphenylboronic acid
##STR00051##
[0294] To the mixture of diisopropyl
2-(1,3-dioxolan-2-yl)-6-ethoxyphenylboronate (136 g, 0.42 mol) in
THF (500 mL) was added diluted HCl (2N, 200 mL) slowly at room
temperature with stirring. After stirred for 1.5 h at room
temperature, the reaction mixture was basified with 20% aqueous
solution of NaOH to pH=12 and then washed with EtOAc (2.times.100
mL). The aqueous layer was acidified by using the diluted HCl (2N)
to pH=2 and then extracted with EtOAc (3.times.100 mL). The
combined organic layers were dried over anhydrous Na.sub.2SO.sub.4
and concentrated to dryness. The residue was purified by column
chromatography on silica gel to give the target compound as white
solid (80 g, yield 83%). .sup.1H NMR (400 MHz, DMSO-d) .delta. 9.93
(s, 1H), 7.92 (s, 2H), 7.45-7.48 (m, 2H), 7.23-7.28 (d, 1H),
4.01-4.06 (m, 2H), 1.69-1.20 (m, 3H).
7-Ethoxy-3-(nitromethyl)benzo[c][1, 2]oxaborol-1 (3H)-ol
##STR00052##
[0296] The mixture of 2-ethoxy-6-formylphenylboronic acid (80 g,
0.41 mol), NaOH (16.5 g, 0.41 mol) and CTAB (7.7 g, 20 mmol) in
H.sub.2O (100 mL) and THF (500 mL) was stirred for 0.5 h at room
temperature After dropwise addition of nitromethane (14 mL, 2.4
mol), the reaction mixture was stirred at room temperature for 3 h.
Then the cyclization was afforded by adding the diluted HCl (2 N)
to pH=2 and then extracted with EtOAc (3.times.300 mL). The
combined organic layers were washed with brine (250 mL), dried over
anhydrous Na.sub.2SO.sub.4 and concentrated to dryness in vacuo.
The residue was purified by column chromatography on silica gel to
give the target compound as white solid (92 g, yield 94%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6
.quadrature..quadrature..quadrature..delta. 9.06 (s, 1H), 7.46-7.43
(t, 1H), 7.07-7.05 (d, 1H), 6.89-6.87 (d, 1H), 5.71-5.69 (m, 1H),
5.31-5.27 (m, 1H), 4.57-4.51 (m, 1H), 4.12-4.07 (m, 2H), 1.34-1.30
(t, 3H).
4-Chloro-7-ethoxy-3-(nitromethyl)benzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00053##
[0298] To a solution of
7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1(3H)-ol (42 g, 0.18
mol) in DMF (200 mL) at 80.degree. C. was added a solution of NCS
(11.8 g, 0.18 mol) in DMF (50 mL) in 30 min. The reaction was
quenched with an aqueous solution of LiCl solution (500 mL) and the
resulting mixture was extracted by EtOAc (3.times.250 mL). The
combined organic layers were dried over anhydrous Na.sub.2SO.sub.4
and concentrated in vacuo. The residue was purified by column
chromatography on silica gel to give the compound as white solid.
(39.7 g, contaminated with 18% C6-Cl regioisomer). The mixture was
then recrystallized from Ether/PE (1/5) to give the pure compound
(28 g, yield 46.3%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
.quadrature. 9.32 (s, 1H), 7.50-7.48 (d, 1H), 6.98-6.96 (d, 1H),
5.77-5.74 (d, 1H), 5.35-5.31 (d, 1H), 4.73-4.67 (m, 1H), 4.12-4.07
(m, 2H), 1.34-1.28 (t, 3H).
3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1,2]oxaborol-1 (3H)-ol
hydrochloride
##STR00054##
[0300] A mixture of
4-chloro-7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1(3H)-ol (47
g, 0.17 mol), Raney Ni (2 g) and 2 M NH.sub.3 in EtOH (40 mL) in
EtOH (200 mL) was stirred under an atmosphere of H.sub.2 for 2 h
and then filtrated. The filtrate was acidified by using 4.5 N HCl
in EtOH (100 mL). After stirring for 30 min, the mixture was
concentrated and the residue was washed with CH.sub.3CN (2.times.50
mL) to give the product as white solid (43 g, yield 89%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 9.13 (s, 1H), 8.18 (s, 3H),
7.50-7.51 (d, 1H), 6.97-7.00 (d, 1H), 5.36-5.39 (m, 1H), 4.08-4.14
(m, 2H), 3.55-3.59 (m, 1H), 2.90-2.95 (m, 1H), 1.33-1.36 (m, 3H);
MS (ESI) m/z=242 [M+H].sup.+.
E. 3-(Aminomethyl)-4-bromo-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
2,2,2-trifluoroacetate salt
3-(Aminomethyl)-7-ethoxybenzo[c][1, 2]oxaborol-1(3H)-ol
hydrochloride salt
##STR00055##
[0302] To the solution of
7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1 (3H)-ol (2 g, 8.43
mmol), Raney Ni (200 mg) and 2 M NH.sub.3 in EtOH (10 mL) in
ethanol (35 mL) was shaken under an atmosphere of H.sub.2 for 2 h
at room temperature. The mixture was filtered through a bed of
Celite and the filtrate was concentrated in vacuo. The crude amine
was dissolved in EtOAc (10 mL) and HCl in Et.sub.2O (30 mL) was
added immediately. After 1 h, the suspension was filtered and the
resulting solid was washed with acetonitrile/hexanes (2:1,
2.times.20 mL) to give the compound as white solid (1 g, yield
57.2%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. .quadrature.
8.89 (s, 1H), 8.22 (s, 3H), 7.48-7.44 (t, 1H), 7.06-7.04 (d, 1H),
6.90-6.88 (d, 1H), 5.31-5.29 (m, 1H), 4.13-4.08 (m, 2H), 3.45-3.39
(m, 1H), 2.80-2.78 (m, 1H), 1.36-1.33 (m, 3H); MS (ESI) m/z=208
[M+H].sup.+.
tert-Butyl (7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1,2]oxaborol-3-yl)methyl-carbamate
##STR00056##
[0304] To the mixture of
3-(aminomethyl)-7-ethoxybenzo[c][1,2]oxaborol-(3H)-ol hydrochloride
salt (300 mg, 1.23 mmol) and triethylamine (622 mg, 6.16 mmol) in
dichloromethane (35 mL) at 0.degree. C. was added di-tert-butyl
dicarbonate (402.8 mg, 1.85 mmol) and the mixture was stirred for 2
h at room temperature. After the reaction was quenched with sat.
NaHCO.sub.3 (45 mL) and the resulting mixture was extracted with
EtOAc (3.times.30 mL), the combined organic layers were dried over
anhydrous Na.sub.2SO.sub.4 and concentrated in vacuo. The residue
was purified by flash-column chromatography to give the product
(320 mg, yield 84.6%).
tert-Butyl (4-bromo-7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,
2]oxaborol-3-yl)-methylcarbamate
##STR00057##
[0306] To the solution of tert-butyl
(7-ethoxy-1-hydroxy-1,3-dihydrobenzo
[c][1,2]oxaborol-3-yl)methyl-carbamate (250 mg, 0.81 mmol) and
1-bromopyrrolidine-2,5-dione (173.9 mg, 0.98 mmol) in CH.sub.3CN
(50 mL) was added 2,2'-Azobis(2-methylpropionitrile (10 mg) and the
mixture was stirred for 1 h at 90.degree. C. The reaction mixture
was then concentrated in high vacuo and the residue was purified by
prep-HPLC to give the product (200 mg, yield 63.6%). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 8.90 (s, 1H), 7.55-7.53 (d, 1H),
6.85-6.82 (d, 1H), 5.08-5.07 (d, 1H), 4.11-4.07 (m, 2H), 3.82-3.79
(d, 1H), 3.06-3.03 (m, 1H), 1.39 (s, 9H), 1.30 (t, 3H); MS (ESI)
m/z=387 [M+H].sup.+.
3-(Aminomethyl)-4-bromo-7-ethoxybenzo[c][1, 2]oxaborol-1 (3H)-ol 2,
2, 2-trifluoroacetate salt
##STR00058##
[0308] The mixture of tert-butyl
(4-bromo-7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-3-yl)-methyl-
carbamate (200 mg, 51.8 mmol) in 2,2,2-trifluoroacetic acid and
dichloromethane (1:1, 20 mL) was stirred at room temperature for 1
h and concentrated to dryness (water bath <30.degree. C.). The
residue was washed with acetonitrile (2.times.5 mL) and the white
solid was dried in high vacuo to give the product (190 mg, yield
91.6%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.12 (s, 1H),
8.04 (s, 3H), 7.65-7.62 (d, 1H), 6.94-6.92 (d, 1H), 5.27-5.25 (m,
1H), 4.13-4.08 (m, 2H), 3.64-3.61 (m, 1H), 2.99-2.92 (m, 1H),
1.36-1.33 (t, 3H); MS (ESI) m/z=287 [M+H].sup.+.
F. 3-(Aminomethyl)-7-ethoxy-4-methylbenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride salt
7-Ethoxy-4-methyl-3-(nitromethyl)benzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00059##
[0310] A mixture of
4-bromo-7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1 (3H)-ol
(200 mg, 0.63 mmol), tetramethylstannane (341.7 mg, 1.90 mmol) and
Pd(PPh.sub.3).sub.4 (Cat. 20 mg) in DMF (35 mL) was stirred
overnight at 90.degree. C. under N.sub.2 protection. The reaction
was quenched by adding ice-water and the resulting mixture was
extracted with ethyl acetate (3.times.30 mL). The combined extracts
were dried over anhydrous Na.sub.2SO.sub.4 and concentrated in
vacuo. The residue was purified by prep-TLC to give the product (72
mg, yield 45.3%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.00
(s, 1H), 7.23-7.21 (d, 1H), 6.83-6.81 (d, 1H), 5.77-5.75 (m, 1H),
5.27-5.24 (m, 1H), 4.50-4.44 (m, 1H), 4.08-4.03 (m, 2H), 2.25 (s,
3H), 1.33-1.29 (t, 3H).
3-(Aminomethyl)-7-ethoxy-4-methylbenzo[c][1, 2]oxaborol-1 (3H)-ol
hydrochloride salt
##STR00060##
[0312] A mixture of
7-ethoxy-4-methyl-3-(nitromethyl)benzo[c][1,2]oxaborol-1 (3H)-ol
(80 mg, 0.32 mmol), Raney Ni (50 mg) and NH.sub.3/EtOH (2 mL) in
EtOH (10 mL) was stirred under an atmosphere of H.sub.2 for 2 h and
then filtrated. The filtrate was acidified by using 4.5 N HCl in
EtOH (15 mL). After stirring for 30 min, the mixture was
concentrated in vacuo and the residue was washed with CH.sub.3CN
(2.times.3 mL) to give the product as white solid (39 mg, yield
47.5%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 8.80 (s, 1H),
8.15 (s, 3H), 7.24-7.22 (d, 1H), 6.83-6.81 (d, 1H), 5.37-5.35 (m,
1H), 4.08-4.03 (m, 2H), 3.36-3.28 (m, 1H), 2.73-2.70 (m, 1H), 2.23
(s, 3H), 1.34-1.30 (t, 3H); MS (ESI) m/z=222 [M+H].sup.+.
G.
3-(Aminomethyl)-7-ethoxy-4-ethylbenzo[c][1,2]oxaborol-1(3H)-ol
7-Ethoxy-3-(nitromethyl)-4-vinylbenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00061##
[0314] A mixture of
4-bromo-7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1(3H)-ol (900
mg, 2.85 mmol), vinyltributyltin (5.2 g, 53 mmol) and
Pd(Ph.sub.3P).sub.4 (230 mg, 0.2 mmol) in DMF (45 mL) was degassed
for 15 min with N.sub.2 and then stirred at 100.degree. C. for 30
min in microwave reactor (Biotage). After the reaction was quenched
with ice-water, the resulting mixture was extracted with EtOAc
(3.times.30 mL). The combined organic layers were washed with water
(20 mL) and brine (20 mL), dried over anhydrous Na.sub.2SO.sub.4
and then concentrated to dryness in vacuo. The residue was purified
by column chromatography on silica gel to give the compound (650
mg, yield 87%). .sup.1H NMR (400 MHz, DMSO-d.sub.6
.quadrature..quadrature..delta. 9.10 (s, 1H), 7.64-7.66 (d, 1H),
6.93-6.95 (d, 1H), 6.77-6.84 (m, 1H), 5.93-5.96 (d, 1H), 5.69-5.73
(d, 1H), 5.28-5.31 (d, 1H), 5.10-5.14 (d, 1H), 4.44-4.49 (m, 1H),
4.09-4.14 (m, 2H), 1.32-1.35 (m, 3H); MS (ESI) m/z=264
[M+H].sup.+.
3-(Aminomethyl)-7-ethoxy-4-vinylbenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00062##
[0316] A mixture of
7-ethoxy-3-(nitromethyl)-4-vinylbenzo[c][1,2]oxaborol-1(3H)-ol (205
mg, 0.78 mmol), Raney-Ni (50 mg) and 2 M NH.sub.3 in EtOH (5 mL) in
EtOH (10 mL) was shaken under an atmosphere of H.sub.2 for 2 h at
room temperature. The mixture was filtered through a bed of Celite
and the filtrate was concentrated in vacuo. The crude amine was
dissolved in EtOAc (2 mL) and HCl in Et.sub.2O (20 mL) was added
immediately. After 1 h, the suspension was filtered and the
resulting solid was used directly for the next step without further
purification.
3-(Aminomethyl)-7-ethoxy-4-ethylbenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00063##
[0318] To a suspension of
3-(aminomethyl)-7-ethoxy-4-vinylbenzo[c][1,2]oxaborol-1(3H)-ol (175
mg, 0.75 mmol) with Pd/C (40 mg) in EtOH (5 ml) was shaken under an
atmosphere of H.sub.2 for 2 h at room temperature. The mixture was
filtered through a bed of Celite and the filtrate was concentrated
in vacuo. The crude amine was dissolved in EtOAc (2 mL) and HCl in
Et.sub.2O (15 mL) was added immediately. After 1 h, the suspension
was filtered and the resulting solid was washed with hexanes to
give the target compound (23 mg, yield 13%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.81 (s, 1H), 8.18 (s, 3H), 7.31-7.29 (d,
1H), 6.68-6.88 (d, 1H), 5.38-5.40 (d, 1H), 4.04-4.09 (d, 2H),
3.30-3.35 (m, 1H), 2.66-2.71 (m, 1H), 1.31-1.34 (m, 3H), 1.15-1.17
(m, 3H); MS (ESI) m/z=236 [M+H].sup.+.
H.
3-(Aminomethyl)-7-ethoxy-4-phenylbenzo[c][1,2]oxaborol-1(3H)-ol
7-Ethoxy-3-(nitromethyl)-4-phenylbenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00064##
[0320] A mixture of
4-bromo-7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1 (3H)-ol
(315 mg, 1 mmol), tributyl-phenyl-stannane (750 mg, 2 mmol) and
Pd(Ph.sub.3P).sub.4 (Cat.) in DMF (15 mL) was degassed for 15 min
with N.sub.2 and then stirred at 100.degree. C. for 30 min in
microwave reactor (Biotage). After the reaction was quenched with
ice-water, the resulting mixture was extracted with EtOAc
(3.times.40 mL). The combined organic layers were washed with water
(20 mL) and brine (20 mL), dried over anhydrous Na.sub.2SO.sub.4
and then concentrated to dryness. The residue was purified by
column chromatography on silica gel to give the compound as white
solid (60 mg, yield 20%). .sup.1HNMR (400 MHz, DMSO-d.sub.6
.quadrature..quadrature.) .delta. 9.13 (s, 1H), 7.46-7.49 (m, 5H),
7.34-7.48 (m, 2H), 6.17-6.20 (m, 1H), 4.88-4.92 (m, 1H), 4.21-4.25
(m, 1H), 4.05-4.16 (m, 2H), 1.34-1.37 (m, 3H).
3-(Aminomethyl)-7-ethoxy-4-phenylbenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00065##
[0322] A mixture of
7-ethoxy-3-(nitromethyl)-4-phenylbenzo[c][1,2]oxaborol-1 (3H)-ol
(60 mg, 0.19 mmol), Raney-Ni (.about.25 mg) and 2 M NH.sub.3 in
EtOH (2 mL) in EtOH (10 mL) was shaken under an atmosphere of
H.sub.2 for 2 h at room temperature. The mixture was filtered
through a bed of Celite and the filtrate was concentrated in vacuo.
The crude amine was dissolved in EtOAc (1 mL) and HCl in Et.sub.2O
(5 mL) was added immediately. After 1 h, the suspension was
filtered and the resulting solid was washed with hexanes to give
the compound as white solid (30 mg, yield 51%). .sup.1HNMR (400
MHz, DMSO-d.sub.6.quadrature..quadrature.) .delta. 8.89 (s, 1H),
8.04 (s, 3H), 7.43-7.46 (d, 5H), 7.36-7.38 (d, 1H), 6.98-7.00 (d,
1H), 5.78-5.81 (d, 1H), 4.09-4.14 (m, 2H), 2.56-2.59 (m, 1H),
2.24-2.30 (m, 1H), 1.33-1.36 (m, 3H); MS (ESI) m/z=284
[M+H].sup.+.
I.
7-(4-Aminobutoxy)-3-(aminomethyl)-4-chlorobenzo[c][1,2]oxaborol-1(3H)-o-
l dihydrochloride
tert-Butyl 4-hydroxybutylcarbamate
##STR00066##
[0324] To a mixture of 4-aminobutan-1-ol (4.0 g, 45 mmol) and TEA
(7.5 mL, 54 mmol) in DCM (200 ml) was added (Boc).sub.2O (10.2 g,
47.2 mmol). The reaction mixture was stirred for 2 h at room
temperature and then washed with water (2.times.150 mL) and the
solution of citric acid (100 mL). The organic layer was dried over
anhydrous Na.sub.2SO.sub.4 and concentrated to give the product as
yellow oil 7.5 g. (yield 88%).
4-(tert-Butoxycarbonyl)butyl methanesulfonate
##STR00067##
[0326] To a mixture of tert-butyl 4-hydroxybutylcarbamate (7.5 g,
40 mmol) and TEA (3.6 mL, 48 mmol) in DCM (100 mL) at 0.degree. C.
was dropwise added MsCl (6.6 mL, 48 mmol). The mixture was stirred
at room temperature for 1 h and then washed with water (2.times.100
mL) and the solution of citric acid (100 mL). The organic layer was
dried over anhydrous Na.sub.2SO.sub.4 and concentrated to give the
product as yellow oil (10.0 g, yield 94%).
tert-Butyl 4-(2-bromo-3-formylphenoxy)butylcarbamate
##STR00068##
[0328] To a mixture of 2-bromo-3-hydroxybenzaldehyde (3.0 g, 15
mmol) and tert-butoxycarbonyl)butyl methanesulfonate (4.8 g, 18
mmol) in DMF (40 mL) was added K.sub.2CO.sub.3 (6.2 g, 45 mmol).
The mixture was stirred at 80.degree. C. for 45 min and quenched by
addition of aqueous LiCl solution (80 mL). The mixture was
extracted with EtOAc (2.times.80 mL) and the combined organic
layers were dried over anhydrous Na.sub.2SO.sub.4 and then
concentrated to give the crude product as brown oil (6.0 g).
tert-Butyl 4-(3-formyl-2-(4, 4, 5, 5-tetramethyl-1, 3,
2-dioxaborolan-2-yl)phenoxy)-butylcarbamate
##STR00069##
[0330] A mixture of tert-butyl 4-(2-bromo-3-formy
lphenoxybutylcarbamate (6.0 g, 16 mmol), KOAc (5.0 g, 48 mmol),
(Pin).sub.2B.sub.2(7.7 g, 86 mmol) and Pb(dppf)Cl.sub.2 (1.25 g,
1.6 mmol) in dioxane (100 mL) was degassed fro 15 min with N.sub.2
and refluxed for 2 h under N.sub.2 protection. The mixture was
filtered and the filtrate was concentrated in vacuo. The residue
was purified by chromatography on silica gel to give the product as
yellow oil (3.5 g, yield 55%).
tert-Butyl-4-(1-hydroxy-3-(nitromethyl)-1, 3-dihydrobenzo[c][1,
2]oxa-borol-7-yloxy)butylcarbamate
##STR00070##
[0332] To a solution of tert-butyl
4-(3-formyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-phenoxy)-buty-
lcarbamate (3.5 g, 8.3 mmol) and CTAB (cat.) in THF (50 mL) was
added MeNO.sub.2 (2.8 mL, 49 mmol), followed by an aqueous solution
of NaOH (0.36 g, 9.1 mmol) in H.sub.2O (5 mL). The mixture was
stirred at room temperature for 45 min. The cyclization was
afforded by adding 2N HCl solution until pH=2 at 0.degree. C. The
reaction mixture was extracted with EtOAc (3.times.50 mL) and the
organic layers were dried over anhydrous Na.sub.2SO.sub.4, and then
concentrated in vacuo. The residue was purified by chromatography
on silica gel (PE:EtOAc=3:1) to give the product as yellow oil (1.7
g, yield 53.6%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.05
(s, 1H), 7.44-7.47 (t, 1H), 7.06-7.08 (d, 1H), 6.88-6.90 (d, 1H),
9.86 (t, 1H), 5.70-5.72 (m, 1H), 5.29-5.33 (m, 1H), 4.53-4.59 (m,
1H), 4.02-4.06 (t, 2H), 2.95-2.30 (m, 2H), 1.67-1.72 (m, 2H),
1.52-1.57 (m, 2H), 1.38 (s, 9H).
tert-Butyl-4-(4-chloro-1-hydroxy-3-(nitromethyl)-1,
3-dihydrobenzo[c][1, 2]oxaborol-7-yloxy)butylcarbamate
##STR00071##
[0334] A mixture of tert-butyl
4-(1-hydroxy-3-(nitromethyl)-1,3-dihydrobenzo[c][1,2]oxaborol-7-yloxy)-bu-
tylcarbamate (640 mg, 1.7 mmol) in DMF (20 mL) was added NCS (226
mg, 1.7 mmol) in DMF (5 mL). The mixture was heated to 80.degree.
C. for 2 h. After the reaction was quenched with an aqueous LiCl
solution (100 mL), the resulting mixture was extracted with EtOAc
(3.times.50 mL). The combined organic layers were dried over
anhydrous Na.sub.2SO.sub.4 and concentrated in vacuo. The residue
was purified by prep-HPLC to give the product (280 mg, yield
67.5%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.29 (s, 1H),
7.47-7.49 (d, 1H), 6.96-6.98 (d, 1H), 6.80 (s, 1H), 5.74-5.77 (m,
1H), 5.31-5.35 (m, 1H), 4.67-4.72 (m, 1H), 4.02-4.05 (m, 2H),
2.94-2.99 (m, 2H), 1.68-1.72 (m, 2H), 1.50-1.56 (m, 2H), 1.36 (s,
9H).
tert-Butyl 4-(3-(aminomethyl)-4-chloro-1-hydroxy-1,
3-dihydro-benzo[c][1, 2]oxa-borol-7-yloxy)butylcarbamate
##STR00072##
[0336] A mixture of tert-butyl
4-(1-hydroxy-3-(nitromethyl)-1,3-dihydrobenzo[c][1,2]oxaborol-7-yl-oxy)-b-
utylcarbamate (410 mg, 1 mmol), Raney-Ni (100 mg) and 2 N NH.sub.3
in EtOH (3 mL) in EtOH (15 mL) was shaken under an atmosphere of
H.sub.2 for 2 h at room temperature. The mixture was filtered
through a bed of Celite and the filtrate was concentrated in vacuo.
The resulting solid was used directly for the next step.
7-(4-Aminobutoxy)-3-(aminomethyl)-4-chlorobenzo[c][1, 2]oxaborol-1
(3H)-ol dihydrochloride
##STR00073##
[0338] To a mixture of the crude
tert-butyl-4-(3-(aminomethyl)-4-chloro-1-hydroxy-1,3-dihydrobenzo[c]-[1,2-
]-oxa-borol-7-yloxy)butylcarbamate in DCM (5 mL) was added
CF.sub.3COOH (2 mL) at room temperature. The reaction mixture was
stirred for 1 h and concentrated in vacuo. The crude amine was
dissolved in EtOAc (1 mL) and HCl in Et.sub.2O (10 mL) was added
immediately. After 1 h, the suspension was filtered and the
resulting solid was washed with hexanes to give the target compound
as white solid (180 mg, yield: 42%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 9.16 (s, 1H), 8.28 (s, 3H), 8.03 (d, 3H),
7.49-7.51 (d, 1H), 6.99-7.01 (d, 1H), 5.38-5.40 (m, 1H), 4.05-4.08
(m, 2H), 3.56-3.59 (d, 1H), 2.84-2.91 (m, 3H), 1.71-1.83 (m, 4H);
MS (ESI) m/z=285 [M+H].sup.+.
J.
3-(Aminomethyl)-7-(3-aminopropoxy)-4-chlorobenzo[c][1,2]oxaborol-1(3H)--
ol tert-Butyl 3-bromopropylcarbamate
##STR00074##
[0340] To a mixture of 3-bromopropan-1-amine (10.95 g, 50 mmol) and
TEA (15.4 mL, 110 mmol) in DCM (100 ml) at 0.degree. C. was added
(Boc).sub.2O (11.4 g, 52.5 mmol). The reaction mixture was stirred
at room temperature overnight and then washed with water
(3.times.100 mL) and the solution of citric acid (100 mL). The
organic layer was dried over anhydrous Na.sub.2SO.sub.4 and
concentrated to give the product as yellow oil (9.0 g, yield
76%).
tert-Butyl 3-(2-bromo-3-formylphenoxy)propylcarbamate
##STR00075##
[0342] A mixture of 2-bromo-3-hydroxybenzaldehyde (5 g, 24.9 mmol),
3-(tert-butoxy carbonylamino)-propyl methanesulfonate (7.55 g, 30
mmol) and Cs.sub.2CO.sub.3 (24 g, 75 mmol) in DMF (60 mL) was
stirred at 50.degree. C. for 3 h and quenched with water (600 mL).
The resulting mixture was extracted with EtOAc (3.times.60 mL) and
the combined organic layers were washed with brine (60 mL), dried
over anhydrous Na.sub.2SO.sub.4 and then concentrated in vacuo. The
residue was purified by column chromatography on silica gel
(PE/EtOAc=10/1) to give the product (7.2 g, yield 81%). .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 10.43 (s, 1H), 7.53 (dd, J=7.8 Hz,
1.6 Hz, 1H), 7.37 (t, J=7.8 Hz, 1H), 7.12 (dd, J=8.2 Hz, 1.6 Hz,
1H), 5.16 (s, 1H), 4.15 (t, J=5.9 Hz, 2H), 3.42 (m, 2H), 2.10 (m,
2H), 1.44 (s, 9H); MS (ESI) m/z=358 [M+H].sup.+.
tert-Butyl 3-(3-formyl-2-(4, 4, 5, 5-tetramethyl-1, 3,
2-dioxaborolan-2-yl)-phenoxy) propylcarbamate
##STR00076##
[0344] A solution of tert-butyl
3-(2-bromo-3-formylphenoxy)propylcarbamate (7.1 g, 20 mmol),
B.sub.2pin.sub.2 (10 g, 40 mmol), Pd(dppf)Cl.sub.2 (800 mg, 2 mmol)
and KOAc (5.9 g, 60 mmol) in 1, 4-dioxane (30 mL) was degassed with
N.sub.2 and stirred at 80.degree. C. for 5 h. The mixture was
cooled to room temperature and diluted with EtOAc (100 mL). The
organic layer was washed with water (50 mL) and brine (50 mL),
dried over anhydrous Na.sub.2SO.sub.4 and then concentrated. The
residue was purified by chromatography on silica gel (PE/EtOAc=5/1)
to give the product (3.1 g, yield 38.3%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 9.95 (s, 1H), 7.48 (t, J=7.8 Hz, 1H), 7.40 (m,
1H), 7.09 (d, J=8.2 Hz, 1H), 4.76 (s, 1H), 4.05 (t, J=6.3 Hz, 2H),
3.32 (m, 2H), 2.00 (m, 2H), 1.45 (s, 12H), 1.43 (s, 9H); MS (ESI)
m/z=406 [M+H].sup.+.
tert-Butyl
3-(1-hydroxy-3-(nitromethyl)-1,3-dihydrobenzo[c][1,2]oxaborol-7-
-yloxy)-propylcarbamate
##STR00077##
[0346] A mixture of tert-butyl
3-(3-formyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)
phenoxy)propylcarbamate (3.1 g, 8.47 mmol), MeNO.sub.2 (775 mg,
12.7 mmol), CTAB (310 mg, 0.85 mmol) and NaOH (407 mg, 10 mmol) in
THF (35 mL) and H.sub.2O (8 mE) was stirred at room temperature for
3 h. The mixture was adjusted to pH 2-3 using 2N HCl and then
stirred for 30 min. The mixture was extracted with EtOAc
(2.times.80 mL). The organic layer was washed with water (30 mL)
and brine (30 mL), dried over anhydrous Na.sub.2SO.sub.4 and then
concentrated. The residue was purified by column chromatography on
silica gel (PE/EtOAc=5/1) to give the product (2 g, yield 64.5%).
.sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.05 (s, 1H), 7.47 (t,
J=7.8 Hz, 1H), 7.07 (d, J=7.4 Hz, 1H), 6.91 (m, 2H), 5.72 (dd,
J=9.0, 2.7 Hz, 1H), 5.31 (dd, J=13.3, 2.7 Hz, 1H), 4.54 (dd,
J=13.3, 9.4 Hz, 1H), 4.05 (t, J=6.3 Hz, 2H), 3.09 (m, 2H), 1.83 (m,
2H), 1.37 (s, 9H); MS (ESI) m/z=367 [M+H].sup.+.
tert-Butyl-4-(4-chloro-1-hydroxy-3-(nitromethyl)-1,
3-dihydrobenzo[c][1, 2]oxaborol-7-yloxy) propylcarbamate
##STR00078##
[0348] To a mixture of tert-butyl
4-(1-hydroxy-3-(nitromethyl)-1,3-dihydrobenzo[c][1,2]oxaborol-7-yl-oxy)pr-
opylcarbamate (2.9 g, 8 mmol) in DMF (35 mL) was added NCS (1.0 g,
8 mmol) in DMF (15 mL). The reaction mixture was heated to
80.degree. C. for 2 h and then quenched with an aqueous LiCl
solution (300 mL). The resulting mixture was extracted with EtOAc
(3.times.100 mL). The combined organic layers were dried over
anhydrous Na.sub.2SO.sub.4 and concentrated in vacuo. The residue
was purified by prep-HPLC to give the product as white solid (480
mg, yield 15.2%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.28
(s, 1H), 7.49-7.51 (d, 1H), 6.96-6.98 (d, 1H), 6.85 (s, 1H),
5.74-5.77 (m, 1H), 5.31-5.35 (m, 1H), 4.67-4.72 (m, 1H), 4.03-4.06
(m, 2H), 3.07-3.11 (m, 2H), 1.82-1.86 (m, 2H), 1.37 (s, 9H); MS
(ESI) m/z=401 [M+H].sup.+.
tert-Butyl 3-(3-(aminomethyl)-4-chloro-1-hydroxy-1,
3-dihydro-benzo[c][1, 2]oxa-borol-7-yloxy)propylcarbamate
##STR00079##
[0350] A mixture of tert-butyl
3-(4-chloro-1-hydroxy-3-(nitromethyl)-1,3-dihydrobenzo[c][1,2]oxaborol-7--
yloxy)propylcarbamate (480 mg, 1.2 mmol), Raney-Ni (500 mg) and 2 M
NH.sub.3 in EtOH (3 mL) in EtOH (15 mL) was shaken under an
atmosphere of H.sub.2 for 2 h at room temperature. The mixture was
filtered through a bed of Celite and the filtrate was concentrated
in vacuo. The resulting solid was used directly for the next
step.
3-(Aminomethyl)-7-(3-aminopropoxy)-4-chlorobenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00080##
[0352] To a mixture of the crude tert-butyl
3-(3-(aminomethyl)-4-chloro-1-hydroxy-1,3-dihydrobenzo[c]-[1,2]oxaborol-7-
-yloxy)propylcarbamate in DCM (10 mL) was added CF.sub.3COOH (2.0
mL) at 0.degree. C. The reaction mixture was stirred for 1 h and
concentrated in vacuo. The crude amine was dissolved in EtOH (2 mL)
and HCl in Et.sub.2O (2 mL) was added immediately. After 1 h, the
mixture was concentrated in vacuo. The residue was recrystallized
by EtOH/Et.sub.2O to give the target compound (173.4 mg, yield
42%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 9.28 (s, 1H),
8.36 (s, 3H), 8.17 (s, 3H), 7.50-7.52 (d, 1H), 6.98-7.00 (d, 1H),
5.39-5.42 (m, 1H), 4.13-4.16 (m, 2H), 3.56-3.59 (d, 1H), 2.98-2.99
(m, 2H), 2.87 (s, 1H), 2.04-2.10 (m, 2H); MS (ESI) m/z=271
[M+H].sup.+.
K.
(R)-3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride and
L.
(S)-3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride
##STR00081##
[0353] tert-Butyl ((4-chloro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1,2]oxaborol-3-yl)methyl)carbamate
##STR00082##
[0355] A solution of
3-(aminomethyl)-4-chloro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride (38.4 g, 0.16 mol) and Et.sub.3N (47.8 g, 0.47 mol)
in CH.sub.2Cl.sub.2 (350 mL) at 0.degree. C. was added
di-tert-butyl dicarbonate (172 g, 0.79 mol) and the reaction was
stirred for 2 h at room temperature. After the reaction was
quenched by addition of sat. NaHCO.sub.3 (100 mL) and the resulting
mixture was extracted with EtOAc (3.times.120 mL). The combined
organic layers were dried over Na.sub.2SO.sub.4 and concentrated in
vacuo. The residue was purified by column chromatography on silica
gel to give the compound as white solid (27 g, yield 50%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 8.92 (s, 1H), 7.40-7.42 (d,
1H), 6.88-6.90 (d, 1H), 6.77-6.79 (m, 1H), 5.15-5.16 (d, 1H),
4.06-4.13 (m, 2H), 3.75-3.78 (d, 1H), 3.03-3.08 (m, 1H), 1.31-1.34
(m, 12H); MS (ESI) m/z=286 [M+H].sup.+.
(S)-tert-Butyl ((4-chloro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)methyl)carbamate and
(R)-tert-butyl ((4-chloro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)methyl) carbamate
##STR00083##
[0357] 25.7 g of tert-butyl
((4-chloro-7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-3-yl)methy-
l)carbamate dissolved in acetonitrile (10 mg/ml) was resolved via
chiral HPLC using ChiralPak AD-H (250.times.30 mm I.D.) and SF
CO.sub.2/methanol as eluent. Flow rate is 70 mL/min. UV detection
was monitored at 220 nm. Two peaks were collected and evaporated to
give 10.65 g of enantiomer A (faster eluting isomer) and 10.15 g of
enantiomer B (slower eluting isomer). Analysis of the pooled
fractions using a ChiralPak AD-3 (150.times.4.6 mm I.D.) and the
same mobile phase showed enantiomer A with a retention time of 3.12
min and 98.7% e.e, and enantiomer B with a retention time of 3.44
min and 98.5% e.e.
(R)-3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1, 2]oxaborol-1
(3H)-ol hydrochloride
##STR00084##
[0359] Enantiomer A (7.0 g, 20.5 mmol) was dissolved in 30 mL of
dioxane and treated with 4M HCl (26.7 mL, 106.6 mmol) in dioxane.
The reaction mixture was stirred at room temperature for overnight
until the reaction was completed indicated by LC/MS. After dioxane
was removed in vacuo and diethyl ether was added, an off-white
solid was collected and dried under high-vacuum. This material was
re-dissolved in acetonitrile and water (1:1, v/v) and lyophilized
to give 5.17 g of the title compound as an off-white solid. .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 9.11 (s, 1H), 8.22 (s, 3H),
7.47 (d, 1H), 6.95 (d, 1H), 5.34-5.37 (m, 1H), 4.06-4.11 (m, 2H),
3.53-3.56 (m, 1H), 2.89 (m, 1H), 1.30-1.34 (m, 3H); MS (ESI)
m/z=242.0 [M+H].sup.+.
(S)-3-(Aminomethyl)-4-chloro-7-ethoxybenzo[c][1, 2]oxaborol-1
(3H)-ol hydrochloride
##STR00085##
[0361] Enantiomer B (7.0 g, 20.5 mmol) was dissolved in 30 mL of
dioxane and treated with 4M HCl (26.7 mL, 106.6 mmol) in dioxane.
The reaction mixture was stirred at room temperature for overnight
until the reaction was completed indicated by LC/MS. After dioxane
was removed in vacuo and diethyl ether was added, an off-white
solid was collected and dried under high-vacuum. This material was
re-dissolved in acetonitrile and water (1:1, v/v) and lyophilized
to give 5.23 g of the title compound as an off-white solid. .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 9.11 (s, 1H), 8.25 (s, 3H),
7.47 (d, 1H), 6.95 (d, 1H), 5.35-5.38 (m, 1H), 4.06-4.11 (m, 2H),
3.53-3.56 (m, 1H), 2.88 (m, 1H), 1.30-1.33 (m, 3H); MS (ESI)
m/z=242.0 [M+H].sup.+.
M.
(R)-3-(Aminomethyl)-4-fluoro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride and
N.
(S)-3-(Aminomethyl)-4-fluoro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride
##STR00086##
[0362] tert-Butyl ((4-fluoro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)methyl)carbamate
##STR00087##
[0364] This compound was prepared from
3-aminomethyl-7-ethoxy-4-fluoro-3H-benzo[c][1,2]-oxaborol-1-ol,
hydrochloride, using the similar procedure as described above.
(S)-tert-Butyl ((4-fluoro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)methyl)carbamate and
(R)-tert-butyl ((4-fluoro-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)methyl)carbamate
##STR00088##
[0366] 4.5 g of tert-buty
((4-fluoro-7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-3-yl)methy-
l)carbamate dissolved in ethanol (100 mg/ml) was resolved via
chiral HPLC using ChiralCel OZ--H column (250.times.30 mm I.D.) and
SF CO.sub.2/hexane:ethanol (1:1) as eluent. Flow rate is 70 mL/min.
UV detection was monitored at 220 nm. Two peaks were collected and
evaporated to give 2.1 g of enantiomer A (faster eluting isomer)
and 2.2 g of enantiomer B (slower eluting isomer). Analysis of the
pooled fractions using a ChiralCel OZ--H (150.times.4.6 mm I.D.)
and SF CO.sub.2/ethanol (0.05% DEA) as mobile phase showed
enantiomer A with a retention time of 2.66 min and 99.5% e.e, and
enantiomer B with a retention time of 3.31 min and 98.1% e.e.
(R)-3-(Aminomethyl)-4-fluoro-7-ethoxybenzo[c][1, 2]oxaborol-1
(3H)-ol hydrochloride
##STR00089##
[0368] Enantiomer A (2.1 g) was treated with 200 mL of 1.6 N HCl in
MeOH and stirred at room temperature for 5 hours until the reaction
was completed indicated by LC/MS. After water (100 mL) was added,
the residue was lyophilized overnight to give 1.40 g of the title
compound as an off-white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. ppm 9.16 (br. s., 1H), 8.34 (br. s., 3H), 7.26 (t, 1H),
6.70-6.92 (m, 1H), 5.48 (d, 1H), 4.06 (q, 2H), 3.35 (m, 1H), 2.88
(m, 1H), 1.30 (t, 3H); MS (ESI) m/z=226.1 (M+1, positive).
(S)-3-(Aminomethyl)-4-fluoro-7-ethoxybenzo[c][1, 2]oxaborol-1
(3H)-ol hydrochloride
##STR00090##
[0370] Enantiomer B (2.1 g) was treated with 200 mL of 1.6 N HCl in
MeOH and stirred at room temperature for 5 hours until the reaction
was completed indicated by LC/MS. After water (100 mL) was added,
the residue was lyophilized overnight to give 1.43 g of the title
compound as an off-white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. ppm 9.16 (br. s., 1H), 8.31 (br. s., 3H), 7.26 (t, 1H),
6.70-6.92 (m, 1H), 5.48 (d, 1H), 4.06 (q, 2H), 3.35 (m, 1H), 2.88
(m, 1H), 1.31 (t, 3H); MS (ESI) m/z=226.1 (M+1, positive).
O.
3-Aminomethyl-5-chloro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,2]oxaborol-1-
-ol hydrochloride
##STR00091## ##STR00092##
[0371] 5-Chloro-2, 3-dihydroxy-benzaldehyde
[0372] To a solution of 5-chloro-2-hydroxy-3-methoxy-benzaldehyde
(7 g, 37.5 mmol) in anhydrous CH.sub.2Cl.sub.2 (200 mL) at
0.degree. C. was added a solution of BBr.sub.3 in CH.sub.2Cl.sub.2
(1 M, 93.7 mL, 93.7 mmol) and the reaction mixture was stirred
overnight at room temperature. The solution was diluted with
CH.sub.2Cl.sub.2 (200 mL), washed with water, brine, dried over
Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure
generating the title compound (6.2 g, 36.0 mmol, 96%) as a light
yellow solid. .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta. ppm 11.02
(s, 1H), 9.83 (s, 1H), 7.18 (s, 1H), 7.14 (d, J=2.3 Hz, 1H), 5.71
(s, 1H).
3-(3-Benzyloxy-propoxy)-5-chloro-2-hydroxy-benzaldehyde
[0373] To a solution of 5-chloro-2,3-dihydroxy-benzaldehyde (3.1 g,
17.7 mmol) in anhydrous DMSO (20 mL) was added NaH (60% in mineral
oil, 1.50 g, 35.4 mmol) portion-wise and the mixture was stirred
for 30 minutes. The solution was cooled to 0.degree. C. and a
solution of 3-benzyloxy-1-bromopropane (3.1 mL, 17.7 mmol) in DMSO
(3 mL) was added dropwise over 10 minutes period. The ice bath was
removed. After overnight, the solution was diluted with EtOAc (100
mL), washed with water, brine, dried over Na.sub.2SO.sub.4,
filtered and concentrated under reduced pressure. The product was
purified by silica gel column chromatography (2:1 hexanes-EtOAc
mobile phase) generating the title compound (5.3 g, 16.6 mmol, 94%)
as a light yellow oil. .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta.
ppm 10.81 (s, 1H), 9.87 (s, 1H), 7.42-7.28 (m, 5H), 7.17 (s, 1H),
7.08 (s, 1H), 4.53 (s, 2H), 4.17 (t, J=6.2 Hz, 2H), 3.69 (t, J=5.8
Hz, 2H), 2.15 (t, J=6.2 Hz, 2H).
Trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-4-chloro-6-formyl-phenyl ester
[0374] To a solution of
3-(3-benzyloxy-propoxy)-5-chloro-2-hydroxy-benzaldehyde (5.3 g,
16.6 mmol) and pyridine (3.4 mL, 41.5 mmol) in CH.sub.2Cl.sub.2 (70
mL) at 0.degree. C. was added Tf.sub.2O (3.1 mL, 18.3 mmol)
drop-wise over 5 minutes period and the reaction mixture was
stirred for 3 h at room temperature. The solution was diluted with
CH.sub.2Cl.sub.2 (100 mL), washed with water, brine, dried over
Na.sub.2SO.sub.4, then concentrated under reduced pressure. The
product was purified by silica gel column chromatography (2:1
hexanes-EtOAc mobile phase) generating the title compound (3.8 g,
8.5 mmol, 51%) as a light yellow oil. .sup.1H NMR (400 MHz,
CHLOROFORM-d) .delta. ppm 10.18 (s, 1H), 7.48 (d, J=2.3 Hz, 1H),
7.37-7.29 (m, 6H), 4.52 (s, 2H), 4.23 (t, J=6.2 Hz, 2H), 3.69 (t,
J=5.8 Hz, 2H), 2.16 (t, J=6.0 Hz, 2H); .sup.19F NMR (376 MHz,
CHLOROFORM-d) .delta. ppm -73.23 (s).
3-(3-Benzyloxy-propoxy)-5-chloro-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0375] To a solution of trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-4-chloro-6-formyl-phenyl ester (3.8 g, 8.4
mmol) in anhydrous 1,4-dioxane (50 mL) was added
bis(pinacolato)diborane (4.3 g, 16.9 mmol) and KOAc (2.5 g, 25.4
mmol) successively and the resulting solution was degassed with
N.sub.2 for 20 minutes. PdCl.sub.2(dppf) (0.5 g, 0.67 mmol) was
added and the resulting mixture was stirred overnight at 90.degree.
C. The solution was diluted with EtOAc (100 mL), washed with water,
brine, dried over Na.sub.2SO.sub.4, filtered and concentrated under
reduced pressure. The product was purified by silica gel column
chromatography (4:1 hexanes-EtOAc mobile phase) generating the
title compound (3.4 g, 7.8 mmol, 92%) as a colorless oil. .sup.1H
NMR (400 MHz, CHLOROFORM-d) .delta. ppm 9.87 (s, 1H), 7.40-7.28 (m,
6H), 7.03 (br s, 1H), 4.50 (s, 2H), 4.09 (t, J=6.4 Hz, 2H),
3.70-3.60 (m, 2H), 2.10 (t, J=6.2 Hz, 2H), 1.42 (s, 12H).
7-(3-Benzyloxy-propoxy)-5-chloro-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
[0376] To a solution of
3-(3-benzyloxy-propoxy)-5-chloro-2-(4,4,5-trimethyl-[1,3,2]dioxaborolan-2-
-yl)-benzaldehyde (3.4 g, 7.8 mmol) and nitromethane (1.7 mL, 31.3
mmol) in THF (20 mL) was added a solution of NaOH (0.025 M, 40 mL).
After 12 h, 2 N HCl was added until pH was 1. The solution was
diluted with EtOAc (150 mL), washed with water, dried over
Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure.
The product was purified by silica gel column chromatography (4:1
hexanes-EtOAc mobile phase) to give the title compound product (1.7
g, 56%) as a colorless gel. .sup.1H NMR (400 MHz, CHLOROFORM-d)
.delta. ppm 7.39-7.28 (m, 5H), 6.92 (s, 1H), 6.83 (s, 1H), 5.85 (br
s, 1H), 5.81 (dd, J=8.5, 3.9 Hz, 1H), 4.70 (dd, J=13.2, 3.9 Hz,
1H), 4.59 (s, 2H), 4.47 (dd, J=13.0, 8.7 Hz, 1H), 4.21-4.07 (m,
2H), 3.71-3.60 (m, 2H), 2.10 (quin, J=5.7 Hz, 2H).
5-Chloro-7-(3-hydroxy-propoxy)-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
##STR00093##
[0378]
7-(3-Benzyloxy-propoxy)-5-chloro-3-nitromethyl-3H-benzo[c][1,2]oxab-
orol-1-ol (0.3 g, 0.91 mmol) in MeOH (30 mL) was added conc HCl (1
mL) and Pd(OH).sub.2 (10% w/w on carbon, 0.2 g) and the reaction
vessel was pressurized to 40 psi with hydrogen for 30 minutes at
room temperature. The mixture was filtered through a pad of
Celite.RTM. and washed with EtOAc. The filtrate was concentrated in
vacuo and the product was purified by prep. HPLC (C18 column, using
acetonitrile and 0.1% AcOH/water solution gradient) provided the
title compound (80 mg, 33%). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. ppm 9.19 (s, 1H), 7.22 (s, 1H), 6.97 (s, 1H), 5.71 (dd,
J=8.9, 2.3 Hz, 1H), 5.32 (dd, J=13.2, 2.7 Hz, 1H), 4.64 (dd,
J=13.6, 8.9 Hz, 1H), 4.53 (t, J=4.8 Hz, 1H), 4.12 (t, J=6.0 Hz,
2H), 3.57 (q, J=5.5 Hz, 2H), 1.86 (t, J=6.2 Hz, 2H).
3-Aminomethyl-5-chloro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,
2]oxaborol-1-ol hydrochloride
##STR00094##
[0380] To a
5-chloro-7-(3-hydroxy-propoxy)-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-o-
l 7 (80 mg, 0.27 mmol) in methanolic ammonia solution (2 M, 20 mL)
was added Ra/Ni (.about.0.1 g, 2800 Nickel slurry in water) and the
reaction vessel was pressurized to 40 psi with hydrogen overnight
at room temperature. The mixture was filtered through a pad of
Celite.RTM. and washed with EtOAc. The filtrate was concentrated in
vacuo and to the resulting residue was added water (1 mL), followed
by conc HCl to pH 1. The heterogeneous mixture was lyophilized
providing the title compound as a hygroscopic ivory solid (79 mg,
quantitative). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. ppm 7.20
(s, 1H), 6.99 (s, 1H), 5.28 (dd, J=8.0, 2.5 Hz, 1H), 4.13 (t, J=6.0
Hz, 2H), 3.59 (t, J=6.0 Hz, 2H), 3.47 (dd, J=13.0, 2.5 Hz, 1H),
2.89 (dd, J=13.2, 8.6 Hz, 1H), 1.89 (t, J=6.0 Hz, 2H); MS (ESI)
m/z=272 (M+1, positive); HPLC purity: 96.83% (MaxPlot 200-400 nm),
95.40% (220 nm).
P.
3-Aminomethyl-7-(3-hydroxy-propoxy)-6-methoxy-3H-benzo[c][1,2]oxaborol--
1-ol hydrochloride
##STR00095##
[0381] 3-(3-Benzyloxy-propoxy)-2-bromo-4-methoxy-benzaldehyde
[0382] Synthesized according to the methods of general procedure 4
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
2-bromo-3-hydroxy-4-methoxy-benzaldehyde (1.0 g, 4.32 mmol),
(3-bromo-propoxymethyl)-benzene (0.76 mL, 4.32 mmol), cesium
carbonate (2.11 g, 6.5 mmol), DMF (30 mL). Purification: flash
chromatography (10% EtOAc/hexanes): yield 1.54 g (95%). .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. (ppm): 10.26 (s, 1H), 7.73 (d, J=8.6
Hz, 1H), 7.46-7.18 (m, 5H), 6.95 (d, J=8.6 Hz, 1H), 4.56 (s, 2H),
4.14 (t, J=6.1 Hz, 2H), 3.92 (s, 3H), 3.77 (t, J=6.2 Hz, 2H), 2.15
(quin, J=6.5 Hz, 2H); MS (ESI): m/z=381 (M+1, positive).
3-(3-Benzyloxy-propoxy)-4-methoxy-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0383] Synthesized according to the methods of general procedure 5
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-benzyloxy-propoxy)-2-bromo-4-methoxy-benzaldehyde (14.82 g, 39
mmol), bis(pinacolato)diboran (14.86 g, 58.5 mmol), KOAc (11.46 g,
117 mmol), PdCl.sub.2(dppf) (8.5 g, 11.7 mmol), dioxane (200 mL).
Purification: flash column chromatography (15% EtOAc/hexanes):
yield 3.42 g (22%). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta.
(ppm): 9.79 (s, 1H), 7.53 (d, J=8.2 Hz, 1H), 7.40-7.24 (m, 5H),
6.98 (d, J=8.2 Hz, 1H), 4.53 (s, 2H), 4.12 (t, J=6.4 Hz, 2H), 3.88
(s, 3H), 3.69 (t, J=6.4 Hz, 2H), 2.10 (quin, J=6.5 Hz, 2H), 1.44
(s, 12H).
7-(3-Benzyloxy-propoxy)-6-methoxy-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
##STR00096##
[0385] Synthesized according to the methods of general procedure 8
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-benzyloxy-propoxy)-4-methoxy-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaboro-
lan-2-yl)-benzaldehyde (3.36 g, 7.88 mmol), nitromethane (1.28 mL,
23.66 mmol), NaOH (0.22 g, 5.52 mmol), THF (6 mL), water (18 mL).
Purification: flash column chromatography (30% EtOAc/hexanes):
yield 1.2 g (41%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
(ppm): 9.36 (s, 1H), 7.40-7.22 (m, 5H), 7.22-7.04 (m, 2H), 5.68
(dd, J=9.4, 2.7 Hz, 1H), 5.29 (dd, J=13.4, 2.5 Hz, 1H), 4.52 (dd,
J=13.3, 9.4 Hz, 1H), 4.45 (s, 2H), 4.25 (t, J=6.2 Hz, 2H), 3.75 (s,
3H), 3.61 (t, J=6.2 Hz, 2H), 1.92 (quin, J=6.5 Hz, 2H).
3-Aminomethyl-7-(3-hydroxy-propoxy)-6-methoxy-3H-benzo[c][1,
2]oxaborol-1-ol; hydrochloride
##STR00097##
[0387] To a solution of
7-(3-benzyloxy-propoxy)-6-methoxy-3-nitromethyl-3H-benzo[c][1,2]oxaborol--
1-ol (0.5 g, 1.29 mmol) in methanolic ammonia (10 mL) was added
palladium hydroxide (0.25 g, 45 wt %) in a hydrogenation bottle and
the flask was charged with hydrogen at 45 psi for 18 h. The
catalyst was filtered off and the solvent was evaporated under
reduced pressure. In order to assure all the ammonia has been
stripped off, the compound was subjected to the high vacuum for 1
h. The crude (0.4 g) obtained was further dissolved in methanol (15
mL) and transferred to a hydrogenation bottle and concentrated HCl
(5-6 drops) was added to make it to pH 2. To this solution
palladium hydroxide (0.11 g, 25 wt %) was added and the flask was
charged with hydrogen to 45 psi for 1.5 h. The catalyst was
filtered off through a pad of Celite and the solvent evaporated.
Purification was accomplished by preparative HPLC generating 0.16 g
(41%) of the title compound as a white solid. .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. (ppm): 8.06 (br. s, 1H), 7.17 (d, J=8.2 Hz,
1H), 7.12-7.02 (m, 1H), 5.32 (dd, J=7.8, 2.3 Hz, 1H), 4.46-4.27 (m,
2H), 4.14 (t, J=4.5 Hz, 2H), 3.78 (s, 3H), 3.44 (dd, J=13.3, 2.7
Hz, 1H), 2.88 (dd, J=13.3, 8.2 Hz, 1H), 2.08-1.94 (m, 2H); MS
(ESI): m/z=268 (M+1, positive); HPLC purity: 95.35% (MaxPlot
200-400 nm), 97.48% (220 nm).
Q.
3-Aminomethyl-7-(3-hydroxy-propoxy)-6-methyl-3H-benzo[c][1,2]oxaborol-1-
-ol; hydrochloride
##STR00098## ##STR00099##
[0388] 1, 2-Dimethoxy-3-methyl-benzene
[0389] To a cooled (0.degree. C.) solution of
1,2-dimethoxy-3-methylbenzene (2.05 g, 13.45 mmol) and TMEDA (2.8
mL, 18.83 mmol) in diethyl ether (100 mL) was added t-butyllithium
(1.7 M in pentane, 9.5 mL, 16.14 mmol). The color of the solution
changed to light yellow and after a few minutes a white precipitate
was observed. The suspension was stirred at room temperature for 18
h, cooled to 0.degree. C. and dimethylformamide (2.08 mL, 26.90
mmol) was added dropwise. The precipitate disappeared and the color
of the solution changed to light pink. After stirring for 0.5 h,
ice was added followed by 1N HCl (30 mL), the compound was
extracted into ethyl acetate, dried (Na.sub.2SO.sub.4) and the
solvent was evaporated to obtain light brown oil. Purification by
flash column chromatography (5% EtOAc/hexane) generated the title
compound: yield 1.4 g (58%). .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. (ppm): 10.34 (s, 1H), 7.49 (d, J=8.2 Hz, 1H), 7.01 (d,
J=7.8 Hz, 1H), 4.00 (s, 3H), 3.86 (s, 3H), 2.33 (s, 3H). MS (ESI):
m/z=181 (M+1, positive).
2,3-Dihydroxy-4-methyl-benzaldehyde
[0390] To a solution of 1,2-dimethoxy-3-methyl-benzene (13.8 g,
76.66 mmol) cooled to -30.degree. C. (dry ice/acetone) in
dichloromethane (200 mL) was added boron trichloride (230 mL, 230
mmol) dropwise and the mixture was left to stir overnight at room
temperature. The solution was cooled to 0.degree. C. and ice/water
was added carefully, and then extracted with excess of
dichloromethane. The organic layer was washed with water, dried
(Na.sub.2SO.sub.4) and the solvent was evaporated. Purification by
silica gel column chromatography (10-20% EtOAc/hexane) gave the
title compound as a crystalline solid: yield 9.2 g (80%). .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. (ppm): 11.11 (s, 1H), 9.82 (s,
1H), 7.05 (d, J=8.2 Hz, 1H), 6.81 (d, J=8.6 Hz, 1H), 5.67 (s, 1H),
2.33 (s, 3H).
3-(3-Benzyloxy-propoxy)-2-hydroxy-4-methyl-benzaldehyde
[0391] Synthesized according to the methods of general procedure 4
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
2,3-dihydroxy-4-methyl-benzaldehyde (9 g, 59.21 mmol),
(3-bromo-propoxymethyl)-benzene (11.5 mL, 65.13 mmol), sodium
tert-butoxide (12.52 g, 130.26 mmol) and DMSO (100 mL).
Purification: flash column chromatography (5-10% EtOAc/hexane):
yield 15.1 g (85%). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta.
(ppm): 11.05 (s, 1H), 9.84 (s, 1H), 7.38-7.26 (m, 5H), 7.20 (d,
J=7.8 Hz, 1H), 6.80 (d, J=8.2 Hz, 1H), 4.55 (s, 2H), 4.15 (t, J=6.2
Hz, 2H), 3.73 (t, J=6.2 Hz, 2H), 2.31 (s, 3H), 2.19-2.00 (m,
2H).
Trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-6-formyl-3-methyl-phenyl ester
[0392] Synthesized according to the methods of general procedure 6
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-Benzyloxy-propoxy)-2-hydroxy-4-methyl-benzaldehyde (0.3 g, 1.0
mmol), trifluoromethanesulfonic acid (0.34 mL, 2.0 mmol), pyridine
(0.25 mL, 3.1 mmol), dichloromethane (15 mL). Purification: flash
column chromatography (10-15% EtOAc/hexane): yield 0.25 g (59%).
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. (ppm): 10.14 (s, 1H),
7.61 (d, J=7.8 Hz, 1H), 7.42-7.29 (m, 6H), 4.53 (s, 2H), 4.07 (t,
J=6.4 Hz, 2H), 3.71 (t, J=6.1 Hz, 2H), 2.40 (s, 3H), 2.07-2.22 (m,
2H). .sup.19F NMR (400 MHz, CDCl.sub.3) .delta. (ppm): -73.63
(s).
3-(3-Benzyloxy-propoxy)-4-methyl-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0393] Synthesized according to the methods of general procedure 5
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-6-formyl-3-methyl-phenyl ester (0.26 g, 0.6
mmol), bis(pinacolato)diboran (0.31 g, 1.2 mmol), KOAc (0.18 g, 1.8
mmol), PdCl.sub.2(dppf) (0.13 g, 0.18 mmol), THF (10 mL).
Purification: flash column chromatography (15% EtOAc/hexane): yield
0.091 g (36%). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. (ppm):
9.89 (s, 1H), 7.47 (d, J=7.4 Hz, 1H), 7.40-7.25 (m, 6H), 4.52 (s,
2H), 4.02 (t, J=6.4 Hz, 2H), 3.70 (t, J=6.2 Hz, 2H), 2.32 (s, 3H),
2.20-2.08 (m, 2H), 1.45 (s, 12H).
7-(3-Benzyloxy-propoxy)-6-methyl-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
##STR00100##
[0395] Synthesized according to the methods of general procedure 8
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-benzyloxy-propoxy)-4-methyl-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborol-
an-2-yl)-benzaldehyde (1.14 g, 2.78 mmol), nitromethane (0.45 mL,
8.34 mmol), NaOH (0.78 g, 1.95 mmol), THF (3 mL) and water (9 mL).
Purification: flash column chromatography (25% EtOAc/hexanes):
yield 0.42 g (41%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
(ppm): 9.41 (s, 1H), 7.37-7.18 (m, 6H), 7.03 (d, J=7.8 Hz, 1H),
5.69 (dd, J=9.2, 2.5 Hz, 1H), 5.28 (dd, J=13.7, 2.7 Hz, 1H), 4.52
(dd, J=13.3, 9.4 Hz, 1H), 4.46 (s, 2H), 4.33 (t, J=6.1 Hz, 2H),
3.58 (t, J=6.1 Hz, 2H), 2.12 (s, 3H), 2.01-1.87 (m, 2H). MS (ESI):
m/z=370 (M-1, negative).
3-Aminomethyl-7-(3-hydroxy-propoxy)-6-methyl-3H
benzo[c][1,2]oxaborol-1-ol hydrochloride
##STR00101##
[0397] To a solution of
7-(3-benzyloxy-propoxy)-6-methyl-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-
-ol (0.42 g, 1.13 mmol) in methanolic ammonia (15 mL) was added
palladium hydroxide (0.2 g, 45 wt %) in a hydrogenation bottle and
the flask was charged with hydrogen at 45 psi for 18 h. The
catalyst was filtered off and the solvent was evaporated under
reduced pressure. In order to assure all the ammonia has been
stripped off, the compound was subjected to the high vacuum for 1
h. The crude (0.38 g) obtained was further dissolved in methanol
(15 mL) and transferred to a hydrogenation bottle and concentrated
HCl (5-6 drops) was added to make it to pH 2. To this solution
palladium hydroxide (0.1 g, 25 wt %) was added and the flask was
charged with hydrogen at 45 psi for 1.5 h. The catalyst was
filtered off through a pad of Celite and the solvent evaporated.
Purification by preparative HPLC provided 0.12 g (39%) of the title
compound as a white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. (ppm): 8.40 (s, 1H), 7.12 (d, J=7.4 Hz, 1H), 6.80 (d, J=7.4
Hz, 1H), 5.08-4.91 (m, 1H), 4.49-4.21 (m, 2H), 3.64 (t, J=5.7 Hz,
2H), 3.17 (dd, J=12.9, 3.1 Hz, 1H), 2.66 (dd, J=12.7, 8.0 Hz, 1H),
2.14 (s, 3H), 1.81 (quin, J=5.7 Hz, 2H). MS (ESI): m/z=252 (M+1,
positive); HPLC purity: 98.25% (MaxPlot 200-400 nm), 98.39% (220
nm).
R.
3-Aminomethyl-6-fluoro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,2]oxaborol-1-
-ol; hydrochloride salt
##STR00102##
[0398] 4-Fluoro-2, 3-dihydroxy-benzaldehyde
[0399] To a solution of 3-fluoro-benzene-1,2-diol (20 g, 156 mmol)
in anhydrous acetonitrile (400 mL) was added magnesium chloride
(37.1 g, 312 mmol), paraformaldehyde (31.6 g) and triethylamine
(134 mL, 975 mmol). The reaction mixture was heated at 80.degree.
C. for 8 h. The reaction mixture was cooled to room temperature and
the solid was collected by filtration. The solid was treated with
cold 2 N HCl and the aqueous layer was extracted with EtOAc. The
organic layer was concentrated in vacuo yielding 20.4 g of crude.
After a second run 40.8 g of crude was dissolved in DMF (1 L),
cooled to 0.degree. C., added to Cs.sub.2CO.sub.3 (340 g, 1.04 mol)
portion-wise. Then methyl iodide (330 mL, 5.28 mol) was added.
After warming to room temperature and stirring overnight the
solution was filtered, ethyl acetate was added and the organic
layer was washed with water (3.times.). After concentration in
vacuo the product was purified by Biotage silica gel chromatography
(2% to 3% to 10% to 20% EtOAc/hexanes) resulting in 14.8 g of
dimethoxy compound. This material was dissolved in DCM and cooled
to -30.degree. C. and BCl.sub.3 (1 M in DCM, 134 mL, 0.1343 mol)
was added to the solution at -30.degree. C. After overnight at room
temperature, the solution was cooled to -70.degree. C. and
BBr.sub.3 (1 M in DCM, 67.25 mL, 0.067 mol) was added. After
overnight warming to room temperature, the solution was cooled in
an ice bath and slowly ice water was added. The DCM layer was
separated and the aqueous layer was extracted with DCM (2.times.).
The combined organic layer was extracted with brine (2.times.),
dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo.
After triturating the residue obtained with hexanes/DCM (6:4) the
5.60 g (11% yield) of the title compound obtained was a brownish
pink solid. This material was used in the next step without further
purification. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. (ppm):
11.36 (s, 1H), 9.83 (s, 1H), 7.16-7.13 (m, 1H), 6.82-6.78 (m, 1H),
5.48 (brs, 1H). .sup.19F NMR (376 MHz, DMSO-d.sub.6 with D.sub.2O)
.delta. (ppm): -119.03--119.08 (m, 1F).
3-(3-Benzyloxy-propoxy)-4-fluoro-2-hydroxy-benzaldehyde
[0400] Synthesized according to the methods of general procedure 4
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
4-fluoro-2,3-dihydroxy-benzaldehyde (5.15 g, 32.9 mmol), NaOtBu
(6.95 g, 72.3 mmol), DMSO (200 mL), (3-bromo-propoxymethyl)-benzene
(8.31 g, 36.3 mmol). Purification: Biotage silica gel
chromatography (hexanes/ethyl acetate gradient) generated 4.00 g of
a mixture of the title compound and the dialkylated product. This
material was used in the next step without further purification.
.sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. (ppm): 10.13 (s, 1H),
7.66-7.54 (m, 5H), 7.53-7.33 (m, 1H), 6.92-6.90 (m, 1H), 4.53 (s,
2H), 4.52-4.44 (m, 2H), 3.71-3.62 (m, 2H), 2.18-2.13 (m, 2H).
.sup.19F NMR (376 MHz, DMSO-d.sub.6 with D.sub.2O) .delta. (ppm):
-121.03--121.08 (m, 1F).
3-(3-Benzyloxy-propoxy)-4-fluoro-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0401] Synthesized according to the methods of general procedure 6
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-benzyloxy-propoxy)-4-fluoro-2-hydroxy-benzaldehyde (4.00 g,
13.1 mmol), pyridine (2.34 mL, 28.9 mmol), DCM (100 mL), triflate
anhydride (2.21 mL, 13.5 mmol). Purification: Biotage silica gel
chromatography (hexanes/ethyl acetate gradient) generated 2.00 g of
triflate that was used immediately according to general procedure 5
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692).
[0402] Synthesized according to the methods of general procedure 5
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
trifluoro-methanesulfonic acid
2-(3-benzyloxy-propoxy)-3-fluoro-6-formyl-phenyl ester (2.0 g, 4.57
mmol), THF (15 mL), B.sub.2pin.sub.2 (2.20 g, 8.66 mmol), KOAc
(1.60 g, 16.3 mmol), PdCl.sub.2(dppf)-DCM (0.40 g, 0.55 mmol).
Purification: Biotage silica gel chromatography (hexanes/ethyl
acetate gradient) generated 0.50 g (26% yield) of the title
compound. .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. (ppm): 9.86
(s, 1H), 7.51 (dd, J=8.2, 4.0 Hz, 1H), 7.34-7.31 (m, 5H), 7.21 (dd,
J=11.0, 8.2 Hz, 1H), 4.51 (s, 2H), 4.24 (td, J=6.5, 2.0 Hz, 2H),
3.67 (t, J=6.2 Hz, 2H), 2.11-2.07 (m, 2H); 1.33 (s, 12H); .sup.19F
NMR (376 MHz, DMSO-d.sub.6 with D.sub.2O) .delta. (ppm):
-120.3--121.1 (m, 1F).
7-(3-Benzyloxy-propoxy)-6-fluoro-3-nitromethyl-3H-benzo[c][1,
2]oxaborol-1-ol
##STR00103##
[0404] Synthesized according to the methods of general procedure 8
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
3-(3-benzyloxy-propoxy)-4-fluoro-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborol-
an-2-yl)-benzaldehyde (0.40 g, 0.966 mmol), nitromethane (0.15 mL,
2.89 mmol), NaOH (0.038 g, 0.96 mmol), THF (10 mL), water (10 mL).
This generated 0.34 g (94% yield) of the title compound. .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. (ppm): 9.83 (s, 1H), 7.32-7.02
(m, 7H), 5.78-5.75 (m, 1H), 5.28-5.23 (m, 1H), 4.60-4.56 (m, 1H),
4.42 (s, 2H), 4.37 (brs, 2H), 3.57 (brs, 2H), 1.92 (brs, 2H).
.sup.19F NMR (376 MHz, DMSO-d.sub.6 with D.sub.2O) .delta. (ppm):
-132.3 (1F).
3-Aminomethyl-6-fluoro-7-(3-hydroxy-propoxy)-3H-benzo[c][1,
2]oxaborol-1-ol; hydrochloride salt
##STR00104##
[0406] To a mixture of
7-(3-benzyloxy-propoxy)-6-fluoro-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-
-ol (0.34 g, 0.906 mmol) and methanolic ammonia (2M, 20 mL) in a
Parr apparatus was added Pd(OH).sub.2 on carbon (0.30 g). The
apparatus was charged with hydrogen (.about.40 psi) and was shaken
overnight at rt. The suspension was filtered through Celite.RTM.
with methanol washing and was concentrated in vacuo. The 310 mg of
cream colored solid was dissolved in methanol (20 mL), transferred
to Parr apparatus and the pH was adjusted to .about.3 with a few
drops of concentrated HCl. Then Pd(OH).sub.2 on carbon (0.20 g) was
added and the apparatus was charged with hydrogen (.about.40 psi).
After 35 minutes, the suspension was filtered through Celite.RTM.
with methanol washing and was concentrated in vacuo. Purification
was accomplished by reverse phase preparative HPLC
(acetonitrile/water (0.1% AcOH) gradient) generating 100 mg (43%
yield) of the title compound as a white solid. mp 265-267.degree.
C.; .sup.1H NMR (400 MHz, DMSO-d.sub.6 with D.sub.2O) .delta.
(ppm): 7.41 (dd, J=11.1, 8.2 Hz, 1H), 7.07 (dd, J=7.9, 2.8 Hz, 1H),
5.29 (d, J=7.0 Hz, 1H), 4.36 (t, J=6.0 Hz, 2H), 3.62 (br.s, 2H),
3.45 (d, J=12.9 Hz, 1H), 2.92-2.86 (m, 1H), 1.93-1.83 (m, 2H);
.sup.19F NMR (376 MHz, DMSO-d.sub.6 with D.sub.2O) .delta. (ppm):
-135.0 (1F); MS (ESI) m/z=256 (M+1, positive); HPLC purity: 98.57%
(MaxPlot 200-400 nm), 97.28% (220 nm).
S. 3-Aminomethyl-7-ethoxy-6-methoxy-3H-benzo[c][1,2]oxaborol-1-ol;
hydrochloric acid salt
##STR00105##
[0407] 2-Bromo-3-ethoxy-4-methoxy-benzaldehyde
[0408] Ethyl bromide (2.88 g, 26.4 mmol) was added to a mixture of
2-bromo-3-hydroxy-4-methoxybenzaldehyde (5.08 g, 22 mmol) and
potassium carbonate (4.56 g, 33 mmol) in anhydrous DMF (50 mL) at
room temperature under nitrogen. The reaction mixture was stirred
at 35.degree. C. for 18 h, diluted with EtOAc (150 mL), washed with
water (2.times.50 mL), brine, dried over Na.sub.2SO.sub.4 and
concentrated to give crude product as a white solid. Purification
by silica column chromatography (eluant: 30% EtOAc in Hexanes) to
generate 5.65 g (99% yield) of the title compound as a white solid.
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. (ppm) 10.26 (s, 1H), 7.73
(d, J=8.4 Hz, 1H), 6.95 (d, J=8.8 Hz, 1H), 4.08 (q, J=7.1 Hz, 2H),
3.95 (s, 3H), 1.46 (t, J=7.0 Hz, 3H); MS (ESI) m/z=261 (M+1,
positive).
[3-Ethoxy-4-methoxy-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0409] Synthesized according to the methods of general procedure 5
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
2-bromo-3-ethoxy-4-methoxy-benzaldehyde (4 g, 15.43 mmol), KOAc
(4.55 g, 46.29 mmol), bis(pinacolato)diboron (7.84 g, 30.86 mmol).
PdCl.sub.2(dppf) (0.91 g, 1.24 mmol) in dry dioxane (90 mL). The
crude product was purified by silica gel column chromatography
(eluant: EtOAc/hexanes 1:9 then 1:3) to afford the title compound
as a white solid (1.50 g, 32% yield). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. (ppm) 9.71 (s, 1H), 7.44 (d, J=8.4 Hz, 1H),
6.89 (d, J=8.4 Hz, 1H), 4.01 (q, J=7.1 Hz, 2H), 3.82 (s, 3H), 1.38
(s, 12H), 1.35 (t, J=7.0 Hz, 3H).
7-Ethoxy-6-methoxy-3-nitromethyl-3H-benzo[c][1, 2]oxaborol-1-ol
##STR00106##
[0411] Synthesized according to the methods of general procedure 9
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
[3-ethoxy-4-methoxy-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-benz-
aldehyde (1.47 g, 4.80 mmol), nitromethane (0.92 g, 14.4 mmol),
CATBr (88 mg, 0.24 mmol) in dry THF (20 mL) and NaOH (0.025 M
aqueous solution). Purification by silica gel column chromatography
(eluant: 10% EtOAc/hexane to 30% EtOAc/hexane) to obtain the title
compound as a yellow solid (0.75 g, 59%). .sup.1H NMR {400 MHz,
DMSO-d.sub.6+D.sub.2O (0.01 ml)} .delta. (ppm) 9.34 (s, 1H), 7.18
(d, J=8.1 Hz, 1H), 7.11 (d, J=8.0 Hz, 1H), 5.68 (dd, J=9.2, 2.0 Hz,
1H), 5.29 (dd, J=13.2, 2.8 Hz, 1H), 4.55-4.50 (m, 1H), 4.20 (q,
J=7.0 Hz, 2H), 3.77 (s, 3H), 1.27 (t, J=7.0 Hz, 3H); MS (ESI)
m/z=261 (M-1, negative).
3-Aminomethyl-7-ethoxy-6-methoxy-3H-benzo[c][1, 2]oxaborol-1-ol;
hydrochloride salt
##STR00107##
[0413] Synthesized according to the methods of general procedure 13
in U.S. Pat. Pub. No. 20090227541 (U.S. patent application Ser. No.
12/142,692) using the following reactants and amounts:
7-Ethoxy-6-methoxy-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-ol
(0.97 g, 3.63 mmol), glacial acetic acid (20 mL), Pd(OH).sub.2 on
carbon (20% metal content, 50% weight-wet) (300 mg). Purification:
preparative HPLC (C18 column, using acetonitrile and 0.1%
AcOH/water solution) provided the title compound (0.28 g; 28%
yield). m.p. 202-204.degree. C. .sup.1H NMR {400 MHz, CD.sub.3OD)
.delta. (ppm) 7.20 (d, J=8.0 Hz, 1H), 7.08 (d, J=8.1 Hz, 1H), 5.40
(dd, J=8.4, 2.8 Hz, 1H), 4.23 (q, J=7.1 Hz, 2H), 3.31 (s, 3H), 3.56
(dd, J=13.6, 7.2 Hz, 1H), 2.92 (dd, J=13.2, 7.2 Hz 1H), 1.33 (t,
J=7 Hz, 3H); MS (ESI) m/z=238 (M+1, positive); HPLC purity: 98.79%
(MaxPlot 200-400 nm) and 99.13% (220 nm).
T. 3-Aminomethyl-7-ethoxy-6-fluoro-3H-benzo[c][1,2]oxaborol-1-ol;
hydrochloride salt
##STR00108## ##STR00109##
[0414] 4-Fluoro-2, 3-dihydroxybenzaldehyde
[0415] To a solution at -78.degree. C. of
2,3-dimethoxy-4-fluorobenzaldehyde (7.0 g, 38.0 mmol) in dry
dichloromethane (150 mL) was added dropwise BBr.sub.3 (23.8 g, 95.0
mmol) in dichloromethane (30 mL). Reaction mixture was allowed to
attain room temperature and stirred for 18 h. Then reaction mixture
was cooled to -78.degree. C., and quenched with a mixture of
methanol (10 mL) and water (50 mL) and stirred at room temperature
for 30 min. Precipitated solid was separated by filtration and
washed with cold dichloromethane. Dichloromethane layer was
concentrated to yield the title compound as a solid (5.2 g, 88%).
.sup.1H NMR (400 MHz, CHLOROFORM-d) .delta. ppm: 11.38 (s, 1H),
9.84 (s, 1H), 7.15 (dd, J=8.6, 5.5 Hz, 1H), 6.81 (t, J=9.4 Hz, 1H),
5.47 (s, 1H); MS (ESI) m/z=155 (M-1, negative).
3-Ethoxy-4-fluoro-2-hydroxybezaldehyde
[0416] To a solution of 2,3-dihydroxy-4-fluorobenzaldehyde (3.0 g,
19.23 mmol) in DMSO (100 mL), NaOBu-t (3.692 g, 38.46 mmol) was
added in portions at room temperature and stirred for 15 min. Then
iodoethane was added dropwise at room temperature and stirred for
18 h. The reaction mixture was poured onto crushed ice (200 mL) and
acidified with 2.5 M HCl to pH 3.0. The product was extracted with
ethyl acetate (2.times.100 mL), concentrated and the product was
chromatographed on a column of silica gel (Hex:EtOAc=95:5) to give
the title compound as a crystalline solid (2.3 g, 65%). .sup.1H NMR
(400 MHz, CHLOROFORM-d) .delta. ppm: 11.36 (s, 1H), 9.83 (s, 1H),
7.39-7.19 (m, 1H), 6.77 (t, J=9.2 Hz, 1H), 4.22 (q, J=7.0 Hz, 2H),
1.40 (t, J=7.0 Hz, 3H); MS (ESI) m/z=183 (M+1, positive).
Trifluoromethanesulfonic acid 2-ethoxy-3-fluoro-6-formyl-phenyl
ester
[0417] To a mixture of 3-ethoxy-4-fluoro-2-hydroxybezaldehyde
(2.208 g, 12.0 mmol) and pyridine (1.986 g, 24.0 mmol) in
dichloromethane (30.0 mL) at 0.degree. C. was added dropwise
trifluoromethanesulfonic anhydride (4.060, 14.4 mmol) in
dichloromethane (5.0 mL). The reaction mixture was stirred at
0.degree. C. for 2 h and room temperature for 3 h. Then diluted
with dichloromethane (40 mL), washed with 2M HCl, brine and dried
over anhydrous sodium sulfate. The solvent was removed under
reduced pressure to give the title compound as a light yellow
liquid (3.3 g, 87%). .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta.
ppm: 10.15 (s, 1H), 7.66 (dd, J=8.6, 5.5 Hz, 1H), 7.28-7.22 (m,
1H), 4.36 (q, J=6.9 Hz, 2H), 1.47 (t, J=7.0 Hz, 3H).
3-Ethoxy-4-fluoro-2-(4, 4, 5, 5-tetramethyl-[1, 3,
2]dioxaborolan-2-yl)-benzaldehyde
[0418] To a solution of trifluoromethanesulfonic acid
2-ethoxy-3-fluoro-6-formyl-phenyl ester (2.2 g, 6.96 mmol) in dry
THF (35.0 mL) bis(pinacolato)diboron (2.134 g, 8.4 mmol),
PdCl.sub.2(dppf) (367 mg, 0.5 mmol) and potassium acetate (1.372 g,
14.0 mmol) were added and purged with nitrogen for 15 min. The
reaction mixture was heated under reflux for 24 h. Cooled to room
temperature and diluted with ethyl acetate (40 mL) and filtered
through Celite. The solvent was removed under reduced pressure and
the residue was chromatographed on a column of silica gel
(Hex:EtOAc=9:1) to give the title compound as an off-white solid
(850.0 mg, 42%). .sup.1H NMR (400 MHz, CHLOROFORM-d) .delta. ppm:
9.87 (s, 1H), 7.51 (dd, J=8.2, 4.3 Hz, 1H), 7.22 (dd, J=10.9, 8.6
Hz, 1H), 4.20 (q, J=7.0 Hz, 2H), 1.46 (s, 12H), 1.40 (t, J=7.0 Hz,
3H); MS (ESI) m/z=295 (M+1, positive).
7-Ethoxy-6-fluoro-3-nitromethyl-3H-benzo[c][1, 2]oxaborol-1-ol
##STR00110##
[0420] To a cooled solution of sodium hydroxide (80 mg, 2.0 mmol)
in water (3.0 mL), nitromethane (244.0 mg, 4.0 mmol) was added at
0.degree. C. and stirred for 10 min. Then
3-ethoxy-4-fluoro-2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-benzal-
dehyde (588.0 mg, 2.0 mmol) in THF (5.0 mL) was added. The reaction
mixture was stirred at for 1 h at 0.degree. C. and for 2 h at room
temperature. The reaction mixture was acidified with 2.5 M HCl (1.0
mL) and the product was extracted with ethyl acetate (2.times.20
mL). The organic extracts were combined and washed with brine and
dried over anhydrous sodium sulfate. The solvent was removed under
reduced pressure and product was chromatographed on a column of
silica gel (CH.sub.2Cl.sub.2: MeOH=95:5) to give the title compound
as a solid (350 mg, 69%). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. ppm: 7.40 (dd, J=11.3, 8.2 Hz, 1H), 7.16 (dd, J=8.0, 3.3
Hz, 1H), 5.74 (d, J=9.0 Hz, 1H), 5.30 (dd, J=13.5, 2.2 Hz, 1H),
4.62 (dd, J=13.5, 9.2 Hz, 1H), 4.35 (q, J=6.8 Hz, 2H), 1.28 (t,
J=6.8 Hz, 3H); MS (ESI) m/z=254 (M-1, negative).
3-Aminomethyl-7-ethoxy-6-fluoro-3H-benzo[c][1, 2]oxaborol-1-ol;
hydrochloride salt
##STR00111##
[0422] To a solution of
7-ethoxy-6-fluoro-3-nitromethyl-3H-benzo[c][1,2]oxaborol-1-ol
(320.0 mg, 1.25 mmol) in methanol (5.0 mL), 5.0 mL of 2M ammonia in
methanol and 160 mg of Pd(OH).sub.2 on C were added and
hydrogenated at 45 PSI for 18 h. Catalyst was removed by filtration
and the filtrate was concentrated to generating an off-white solid
(250 mg). This solid was dissolved in methanol (3 mL) and 3.0 mL of
1.2 M HCl in methanol was added and stirred at room temperature for
3 h. Excess HCl and solvent were removed under reduced pressure and
the product was triturated with ether to give the title compound as
an off-white solid (140 mg, 43%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. ppm: 9.43 (s, 1H), 8.13 (br. s., 3H), 7.40
(dd, J=11.5, 8.0 Hz, 1H), 7.16 (dd, J=7.8, 3.1 Hz, 1H), 5.32 (d,
J=6.3 Hz, 1H), 4.35 (q, J=7.0 Hz, 2H), 3.43 (br. s., 1H), 2.92 (br.
s., 1H), 1.29 (t, J=7.0 Hz, 3H); .sup.19F NMR (376 MHz,
DMSO-d.sub.6) .delta. ppm -135 (s, 1F); MS (ESI) m/z=226 (M+1,
positive); HPLC purity: 95.81% (MaxPlot 200-400 nm), 94.73% (220
nm).
U.
3-(Aminomethyl)-5-chloro-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
##STR00112##
[0423] 4-Chloro-2-ethoxy-6-formylphenyl
trifluoromethanesulfonate
[0424] To a solution of 3-ethoxy-2-hydroxybenzaldehyde (20 g, 120.4
mmol) in AcOH (200 mL) was added N-chlorosuccinimide (16.1 g, 120.4
mmol). The reaction mixture was heated up to 105.degree. C. for 30
min. After cooled down to room temperature, the mixture was stirred
for additional 2.5 h. Subsequently, 200 mL of water was added
slowly over 10 min. The mixture was filtered and dried to give a
yellow solid, which was recrystallized in ethanol to give 4 g of
the target compound (4 g, 17% yield).
4-Chloro-2-ethoxy-6-formylphenyl trifluoromethanesulfonate
[0425] To a solution of 5-chloro-3-ethoxy-2-hydroxybenzaldehyde
(2.0 g, 10.0 mmol) in pyridine (2 mL) and DCM (20 mL) at 0.degree.
C. was dropwise added trifluoromethanesulfonic anhydride (1 mL).
The reaction was stirred for 1 h at 0.degree. C. before quenched
with ice-water. The organic layer was washed with sat. aqueous
NaHCO.sub.3 (20 mL) and brine (20 mL), dried over anhydrous
Na.sub.2SO.sub.4 and then concentrated in vacuo. The residue was
purified by column chromatography on silica gel to give target
compound (2.0 g, yield: 60%).
5-Chloro-3-ethoxy-2-(4, 4, 5,
5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzaldehyde
[0426] A mixture of 4-chloro-2-ethoxy-6-formylphenyl
trifluoromethanesulfonate (330 mg, 1 mmol), KOAc (350 mg, 2.0
mmol), bis(pinacolato)diborane (600 mg, 2.0 mmol) and
PdCl.sub.2(dppf)CH.sub.2Cl.sub.2 (65 mg, 0.08 mmol, 8 mol %) in
dioxane (30 mL) was degassed for 15 min with N.sub.2 and stirred at
100.degree. C. for 3 h. After quenched with ice-water, the reaction
mixture was extracted with EtOAc (3.times.30 mL). The combined
organic layers were washed with sat. aqueous NaHCO.sub.3 (20 mL)
and brine (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and then
concentrated. The residue was purified by column chromatography on
silica gel to give the compound (150 mg, yield: 43%).
5-Chloro-7-ethoxy-3-(nitromethyl)benzo[c][1, 2]oxaborol-1
(3H)-ol
[0427] The mixture of
5-chloro-3-ethoxy-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzalde-
hyde (310 mg, 1 mmol), NaOH (40 mg, 1 mmol) and CTAB (5 mg, 0.05
mmol) in H.sub.2O (2 mL) and THF (10 mL) was stirred for 0.5 h at
room temperature. After dropwise addition of nitromethane (0.2 mL,
2 mmol), the reaction mixture was stirred at room temperature for 3
h. Then the cyclization was afforded by adding the diluted aqueous
HCl solution (2 N) to pH=2 and then extracted with EtOAc
(3.times.30 mL). The combined organic layers were washed with brine
(25 mL), dried over anhydrous Na.sub.2SO.sub.4 and concentrated to
dryness. The residue was purified by prep-HPLC to give the compound
(100 mg, yield: 56%). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
9.20 (s, 1H), 7.22 (s, 1H), 6.96 (s, 1H), 5.69-5.72 (m, 1H),
5.30-5.34 (m, 1H), 4.61-4.67 (m, 1H), 4.10-4.15 (m, 2H), 1.31-1.34
(m, 3H);
3-(Aminomethyl)-5-chloro-7-ethoxybenzo[c][1, 2]oxaborol-1
(3H)-ol
##STR00113##
[0429] A mixture of
5-chloro-7-ethoxy-3-(nitromethyl)benzo[c][1,2]oxaborol-1 (3H)-ol
(270 mg, 1.0 mmol), Raney-Ni (.about.125 mg) and 2 M NH.sub.3 in
EtOH (2 mL) in EtOH (10 mE) was shaken under an atmosphere of
H.sub.2 for 2 h at room temperature. The mixture was filtered
through a bed of Celite and the filtrate was concentrated in vacuo.
The crude amine was dissolved in EtOAc (2 mL) and HCl solution in
Et.sub.2O (20 mL) was added immediately. After 1 h, the suspension
was filtered and the resulting solid was washed with hexanes to
give compound the target compound (100 mg, yield: 43%). .sup.1HNMR
(400 MHz, DMSO-d.sub.6.quadrature..quadrature.) 9.06 (s, 1H), 8.18
(s, 3H), 7.22 (s, 1H), 6.96 (s, 1H), 5.27-5.29 (m, 1H), 4.10-4.13
(m, 2H), 3.40-3.47 (m, 1H), 2.87-2.92 (m, 1H), 1.30-1.36 (m, 3H);
MS (ESI) m/z=242 [M+H].sup.+.
V.
(S)-3-(aminomethyl)-4-bromo-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride
Step 1: tert-butyl (7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1,2]oxaborol-3-yl)methyl-carbamate
##STR00114##
[0431] To the mixture of
3-(aminomethyl)-7-ethoxybenzo[c][1,2]oxaborol-1(3H)-ol
hydrochloride salt (5.0 g, 20.5 mmol) and triethylamine (10.4 g,
103.0 mmol) in dichloromethane (250 mL) at 0.degree. C. was added
di-tert-butyl dicarbonate (6.7 g, 30.8 mmol). The mixture was
stirred for 4 h at room temperature. After the reaction was
quenched with sat. NaHCO.sub.3 (500 mL), the resulting mixture was
extracted with EtOAc (3.times.300 mL), and the combined organic
layers were dried over anhydrous Na.sub.2SO.sub.4 and concentrated
in vacum. The residue was purified by flash-column chromatography
(2.5% to 5.0% MeOH in DCM) to give the product (5.51 g, yield
87%).
Step 2: tert-butyl (4-bromo-7-ethoxy-1-hydroxy-1,
3-dihydrobenzo[c][1, 2]oxaborol-3-yl)-methylcarbamate
##STR00115##
[0433] To the solution of tert-butyl
(7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-3-yl)methyl-carbamat-
e (5.5 g, 17.9 mmol) and 1-bromopyrrolidine-2,5-dione (3.8 g, 21.5
mmol) in CH.sub.3CN (1100 mL) was added
2,2'-Azobis(2-methylpropionitrile (220 mg). The mixture was stirred
for 1 h at 90.degree. C. The reaction mixture was then concentrated
in high vacuum and the residue was purified by column
chromatography (2.5% to 5.0% MeOH in DCM) to give the product (3.7
g, yield 54%). .sup.1H NMR (300 MHz, DMSO-d.sub.6) 8.90 (s, 1H),
7.55-7.53 (d, 1H), 6.85-6.82 (d, 1H), 5.08-5.07 (d, 1H), 4.11-4.07
(m, 2H), 3.82-3.79 (bd, 1H), 3.06-3.03 (m, 1H), 1.39 (s, 9H), 1.30
(t, 3H); MS (ESI) m/z=387 [M+H].sup.+.
Step 3: (S)-3-(aminomethyl)-4-bromo-7-ethoxybenzo[c][1,
2]oxaborol-1 (3H)-ol hydrochloride
##STR00116##
[0435] The mixture of tert-butyl
(4-bromo-7-ethoxy-1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-3-yl)-methyl-
carbamate (3.7 g, 9.6 mmol) in 4N HCl in dioxane (12 ml, 48.0 mmol)
was stirred at room temperature for 2 h and then concentrated to
dryness (water bath <30.degree. C.). The residue was triturated
with DCM/ether (1/10, 2.times.10 mL) and the white solid was dried
in high vacuum to give the product (2.96 g, yield: 92%). .sup.1H
NMR (300 MHz, DMSO-d.sub.6) 9.11 (s, 1H), 8.1 (bs, 3H), 7.63-7.60
(d, 1H), 6.92-6.89 (d, 1H), 5.27-5.24 (m, 1H), 4.12-4.05 (m, 2H),
3.62-3.57 (m, 1H), 2.99-2.92 (m, 1H), 1.34-1.30 (t, 3H); MS (ESI)
m/z=287 [M+H].sup.+.
Example 2
[0436] LeuRS IC50 Testing
[0437] Experiments were performed in 96-well microtiter plates,
using 80 .mu.L reaction mixtures containing 50 mM HEPES-KOH (pH
8.0), 30 mM MgCl.sub.2 and 30 mM KCl, 13 .mu.M [.sup.14C]leucine
(306 mCi/mmol, Perkin-Elmer), 15 uM total E. coli tRNA (Roche,
Switzerland), 0.02% (w/v) BSA, 1 mM DTT, 0.2 pM LeuRS and 4 mM ATP
at 30.degree. C. Reactions were started by the addition of 4 mM
ATP. After 7 minutes, reactions were quenched and tRNA was
precipitated by the addition of 50 .mu.L of 10% (w/v) TCA and
transferred to 96-well nitrocellulose membrane filter plates
(Millipore Multiscreen HTS, MSHAN4B50). Each well was then washed
three times with 100 .mu.L of 5% TCA. Filter plates were then dried
under a heat lamp and the precipitated [.sup.14C]leucine
tRNA.sup.Leu was quantified by liquid scintillation counting using
a Wallac MicroBeta Trilux model 1450 liquid scintillation counter
(PerkinElmer, Waltham Mass.).
[0438] To determine the inhibitor concentration which reduces
enzyme activity by 50% (IC.sub.50), increasing concentrations of
inhibitor were incubated with LeuRS enzyme, tRNA and leucine for 20
minutes. Reactions were initiated by the addition of 4 mM ATP and
stopped after 7 minutes then precipitated and counted to quantify
radioactivity.
[0439] Biochemical testing results for exemplary compounds of the
invention are provided in FIG. 1.
Example 3
Antibacterial MIC Testing
[0440] All MIC testing of bacteria followed the Clinical and
Laboratory Standards Institute (CLSI) guidelines for antimicrobial
testing of aerobic bacteria (Methods for Dilution Antimicrobial
Susceptibility Tests for Bacteria That Grow Aerobically; Approved
Standard--Seventh Edition)(M07-A7) and anaerobic bacteria (Methods
for Antimicrobial Susceptibility Testing of Anaerobic Bacteria;
Approved Standard--Seventh Edition) (M11-A7).
[0441] Antibacterial MIC testing results for exemplary compounds of
the invention are provided in FIG. 1.
Example 4
Microplate Alamar Blue Assay (MABA)
[0442] The microplate alamar blue assay (MABA) was essentially
performed as described by Collins, L., et al., Antimicrob Agents
Chemother 41: 1004-1009 (1997). For example, black, clear-bottomed
96-well microplates (black view plates; Packard Instrument Company,
Meriden, Conn.) with the outer perimeter wells filled with sterile
water to prevent dehydration in experimental wells. Initial drug
dilutions were prepared in dimethyl sulfoxide and subsequent two
fold dilutions were performed in 0.1 ml of 7H9GC (no Tween 80) in
the microplates. Frozen inocula were initially diluted 1:20 in
BACTEC 12B medium followed by a 1:50 dilution in 7H9GC. Addition of
100 .mu.L to wells resulted in final bacterial titers of between
2.0.times.10.sup.5 and 5.times.10.sup.4 CFU/mL for H37Rv and H37Ra,
respectively. Wells containing drug only were used to detect
autofluorescence of compounds plus additional control wells
consisted of bacteria only (B) and medium only (M). Plates were
incubated at 37.degree. C. Starting at day 4 of incubation, 20
.mu.L of 10.times. alamar Blue solution (Alamar
Biosciences/Accumed, Westlake, Ohio) and 12.5 .mu.L of 20% Tween 80
were added to one B well and one M well, and plates were
reincubated at 37.degree. C. Wells were observed at 12 and 24 h for
a color change from blue to pink and for a reading of greater than
or equal to 50,000 fluorescence units (FU). Fluorescence was
measured in a Cytofluor II microplate fluorometer (PerSeptive
Biosystems, Framingham, Mass.) in bottom-reading mode with
excitation at 530 nm and emission at 590 nm. If the B wells became
pink by 24 h, reagent was added to the entire plate. If the well
remained blue or 50,000 FU was measured, additional M and B wells
were tested daily until a color change occurred, at which time
reagents were added to all remaining wells. Plates were then
incubated at 37.degree. C., and results were recorded at 24 h
post-reagent addition. Visual MICs were defined as the lowest
concentration of drug that prevented a color change. For
fluorometric MICs, a background subtraction was performed on all
wells with a mean of triplicate M wells. Percent inhibition was
defined as 1-(test well FU/mean FU of triplicate B
wells).times.100. The lowest drug concentration effecting an
inhibition of 90% was considered the MIC.
[0443] Biochemical testing results for exemplary compounds of the
invention are provided in FIG. 1.
Example 5
Low Oxygen Recovery Assay (LORA)
[0444] The low-oxygen recovery assay (LORA) was essentially
performed as described by Cho et al. Antimicrob Agents Chemother
51: 1380-1385 (2007). A recombinant M. tuberculosis H.sub.37R.sub.v
bearing luxAB on a plasmid, pFCA-luxAB, was used in all the LORA
experiments. Frozen aliquots from a low oxygen adapted culture were
thawed, diluted in Middlebrook 7H12 broth (Middlebrook 7H9 broth
containing 1 mg/mL Casitone, 5.6 g/mL palmitic acid, 5 mg/mL bovine
serum albumin, and 4 .mu.g/ml filter-sterilized catalase), and
sonicated for 15 s. The cultures were diluted to obtain an
A.sub.570 of 0.03 to 0.05 and 3,000 to 7,000 RLUs per 100 .mu.L.
This corresponds to 5.times.10.sup.5 to 2.times.10.sup.6 CFU/mL.
Twofold serial dilutions were prepared in a volume 100 .mu.L in
black 96-well microtiter plates, and 100 .mu.L of the cell
suspension was added. For LORA, the microplate cultures were placed
under anaerobic conditions (oxygen concentration, less than 0.16%)
by using an Anoxomat model WS-8080 (MART Microbiology) and three
cycles of evacuation and filling with a mixture of 10% H.sub.2, 5%
CO.sub.2, and 85% N.sub.2. An anaerobic indicator strip was placed
inside the chamber to visually confirm the removal of oxygen. The
plates were incubated at 37.degree. C. for 10 days and then
transferred to an ambient gaseous condition (5% CO.sub.2-enriched
air) incubator for a 28-h "recovery." On day 11 (after the 28-h
aerobic recovery), 100 .mu.L culture was transferred to white
96-well microtiter plates for determination of luminescence. A 10%
solution of n-decanal aldehyde (Sigma) in ethanol was freshly
diluted 10-fold in PBS, and 100 .mu.l was added to each well with
an autoinjector. Luminescence was measured in a Victor2 multilabel
reader (Perkin-Elmer Life Sciences) by using a reading time of 1 s.
The MIC was defined as the lowest drug concentration effecting
growth inhibition of 90% relative to the growth for the drug-free
controls.
[0445] Biochemical testing results for exemplary compounds of the
invention are provided in FIG. 1.
Example 6
Tuberculosis In Vivo Efficacy Experiments
[0446] The TB in vivo efficacy experiments were essentially
performed as described in Lenaerts et al. Antimicrob Agents
Chemother 47: 783-785 (2003) with a few modifications. A highly
susceptible gamma interferon specific pathogen-free
C57BL/6-Ifngtmlts (GKO) mice (Jackson Laboratories, Bar Harbor,
Me.) were exposed to a low-dose aerosol infection with M.
tuberculosis strain Erdman in a Glas-Col inhalation exposure system
as previously described in Kelly et al. Antimicrob Agents Chemother
40: 2809-2812 (1996). Every treatment group consisted of five mice
for every following time point. Treatment was started 10 days after
infection. One control group of infected mice was sacrificed at the
start of treatment. A second group of infected but untreated mice
was sacrificed after the cessation of treatment at 24 days. C and L
were formulated in saline and E was formulated in 50% water/35%
PEG400/5% PG, while rifampicin was formulated in 20% cyclodextrin.
All compounds were administered by oral gavage. Rifampicin was
dosed at 10 mg/kg QD PO. C was dosed at 100 mg/kg BID PO. E was
dosed at 100 mg/kg BID PO. L was dosed at 100 mg/kg QD PO. After
completion of therapy, the mice were sacrificed by carbon dioxide
inhalation. Lungs were aseptically removed and disrupted in a
tissue homogenizer. The number of viable organisms was determined
by serial dilution of the homogenates on nutrient Middlebrook 7H11
agar plates (GIBCO BRL, Gaithersburg, Md.). The plates were
incubated at 37.degree. C. in ambient air for 4 weeks prior to the
counting of viable M. tuberculosis colonies (CFU).
[0447] On day 3, the control group had a mean log 10 CFU/lung of
2.83 (0.40). On day 10, the control group had a log 10 CFU/lung of
4.81 (0.08). On day 24, the control group had a log 10 CFU/lung of
8.96 (0.14). On day 24, the group treated with rifampicin had a log
10 CFU/lung of 6.16 (0.10). On day 24, the group treated with C had
a log 10 CFU/lung of 5.06 (0.26). On day 24, the group treated with
E had a log 10 CFU/lung of 2.73 (0.05). On day 24, the group
treated with L had a log 10 CFU/lung of 3.08 (0.06).
Example 7
Tuberculosis In Vivo Efficacy Experiments
[0448] The TB in vivo efficacy experiments were essentially
performed as described in Lenaerts et al. Antimicrob Agents
Chemother 47: 783-785 (2003) with a few modifications. A highly
susceptible gamma interferon specific pathogen-free
C57BL/6-Ifngtmlts (GKO) mice (Jackson Laboratories, Bar Harbor,
Me.) were exposed to a low-dose aerosol infection with M.
tuberculosis strain Erdman in a Glas-Col inhalation exposure system
as previously described in Kelly et al. Antimicrob Agents Chemother
40: 2809-2812 (1996). Every treatment group consisted of five mice
for every following time point. Treatment was started 13 days after
infection. One control group of infected mice was sacrificed at the
start of treatment. A second group of infected but untreated mice
was sacrificed after the cessation of treatment at 22 days. N was
formulated in saline and E was formulated in 50% water/35%
PEG400/5% PG, while Isoniazid (INH) was formulated in distilled
water. All compounds were administered by oral gavage. INH was
dosed at 25 mg/kg QD PO. E was dosed at 100 mg/kg QD PO. N was
dosed at 100 mg/kg BID PO. After completion of therapy, the mice
were sacrificed by carbon dioxide inhalation. Spleens and lungs
were aseptically removed and disrupted in a tissue homogenizer. The
number of viable organisms was determined by serial dilution of the
homogenates on nutrient Middlebrook 7H11 agar plates (GIBCO BRL,
Gaithersburg, Md.). The plates were incubated at 37.degree. C. in
ambient air for 4 weeks prior to the counting of viable M.
tuberculosis colonies (CFU).
[0449] On day 13, the control group had a mean log 10 0 CFU of 7.02
(0.08) for lungs and mean log 10 CFU for spleens of 3.99 (0.21). On
day 22, the control group had a log 10 CFU for lungs of 7.82 (0.11)
and spleens of 6.69 (0.08). On day 22, the group treated with INH
had a log 10 CFU for lungs of 5.29 (0.13) and for spleens of 4.27
(0.25). On day 22, the group treated with E had a log 10 CFU for
lungs of 5.27 (0.12) and for spleens of 4.27 (0.25). On day 22, the
group treated with N had a log 10 CFU for lungs of 5.51 (0.09) and
spleens of 2.42 (0.48).
[0450] It is understood that the examples and embodiments described
herein are for illustrative purposes only and that various
modifications or changes in light thereof will be suggested to
persons skilled in the art and are to be included within the spirit
and purview of this application and scope of the appended claims.
All publications, patents, and patent applications cited herein are
hereby incorporated by reference in their entirety for all
purposes.
Sequence CWU 1
1
112919DNAMycobacterium tuberculosis 1catatgaccg aaagcccgac
cgcaggtccg ggtggtgtgc cgcgtgcgga tgatgcagat 60agcgatgtgc cgcgttatcg
ttataccgcg gaactggcgg cgcgtctgga acgtacctgg 120caggaaaact
gggcgcgtct gggcaccttt aacgtgccga acccggtggg tagcctggca
180ccgccggatg gtgcagcagt gccggatgat aaactgtttg tgcaggatat
gtttccgtat 240ccgagcggcg aaggcctgca tgtgggccat ccgctgggct
atattgcgac cgatgtgtat 300gcgcgttatt ttcgtatggt gggccgtaac
gtgctgcatg cgctgggctt tgatgcgttt 360ggtctgccgg cggaacagta
tgcggtgcag accggcaccc atccgcgtac ccgtaccgaa 420gcgaacgtgg
tgaactttcg tcgtcagctg ggccgtctgg gctttggcca tgatagccgt
480cgtagcttta gcaccaccga tgtggatttt tatcgttgga cccagtggat
ttttctgcag 540atttataacg cgtggtttga taccaccgcg aacaaagcgc
gtccgattag cgaactggtg 600gcggaatttg aaagcggtgc acgttgcctg
gatggtggtc gtgattgggc aaaactgacc 660gcaggtgaac gtgcggatgt
gattgatgaa tatcgtctgg tgtatcgtgc ggatagcctg 720gtgaactggt
gcccgggtct gggtaccgtg ctggcaaacg aagaagtgac cgcagatggc
780cgtagcgatc gtggcaactt tccggtgttt cgtaaacgtc tgcgtcagtg
gatgatgcgt 840attaccgcgt atgcggatcg tctgctggat gatctggatg
tgctggattg gccggaacag 900gtgaaaacca tgcagcgtaa ctggattggc
cgtagcaccg gcgcggtggc gctgtttagc 960gcgcgtgcgg cgagcgatga
tggctttgaa gtggatattg aagtgtttac cacccgtccg 1020gataccctgt
ttggcgcgac ctatctggtg ctggcgccgg aacatgatct ggtggatgaa
1080ctggtggcgg caagctggcc ggcaggtgtg aacccgctgt ggacctatgg
cggtggtacc 1140ccgggtgaag caattgcagc atatcgtcgt gcgattgcgg
cgaaaagcga tctggaacgt 1200caggaaagcc gtgaaaaaac cggcgtgttt
ctgggcagct atgcgattaa cccggcgaac 1260ggcgaaccgg tgccgatttt
tattgcggat tatgtgctgg cgggctatgg caccggcgcg 1320attatggcgg
tgccgggcca tgatcagcgt gattgggatt ttgcgcgtgc gtttggcctg
1380ccgattgtgg aagtgattgc aggtggaaac attagcgaaa gcgcgtatac
cggcgatggc 1440attctggtga acagcgatta tctgaacggc atgagcgtgc
cggcagcaaa acgtgcaatt 1500gtggatcgtc tggaaagcgc aggtcgtggt
cgtgcacgta ttgaatttaa actgcgtgat 1560tggctgtttg cgcgtcagcg
ttattggggc gaaccgtttc cgattgtgta tgatagcgat 1620ggccgtccgc
atgcgctgga tgaagcggcg ctgccggtgg aactgccgga tgtgccggat
1680tatagcccgg tgctgtttga tccggatgat gcggatagcg aaccgagccc
gccgctggcg 1740aaagcgaccg aatgggtgca tgtggatctg gatctgggcg
atggcctgaa accgtatagc 1800cgtgatacca acgtgatgcc gcagtgggcg
ggcagcagct ggtatgaact gcgttatacc 1860gatccgcata acagcgaacg
tttttgcgcg aaagaaaacg aagcgtattg gatgggtccg 1920cgtccggcag
aacatggtcc ggatgatccg ggtggtgtgg atctgtatgt gggcggcgcg
1980gaacatgcgg tgctgcatct gctgtatagc cgtttttggc ataaagtgct
gtatgatctg 2040ggccatgtga gcagccgtga accgtatcgt cgtctggtga
accagggcta tattcaggcg 2100tatgcgtata ccgatgcgcg tggcagctat
gtgccggcgg aacaagtgat tgaacgtggc 2160gatcgttttg tgtatccggg
cccggatggc gaagtggaag tgtttcagga atttggcaaa 2220attggcaaaa
gcctgaaaaa cagcgtgagc ccggatgaaa tttgcgatgc gtatggcgcg
2280gataccctgc gtgtgtatga aatgagcatg ggcccgctgg aagcgagccg
tccgtgggcg 2340accaaagatg tggtgggcgc gtatcgtttt ctgcagcgtg
tgtggcgtct ggtggtggat 2400gaacataccg gcgaaacccg tgtggcggat
ggcgtggaac tggatattga taccctgcgt 2460gcgctgcatc gtaccattgt
gggcgtgagc gaagattttg cggcgctgcg taacaacacc 2520gcgaccgcga
aactgattga atataccaac catctgacca aaaaacatcg tgatgcagtg
2580ccgcgtgcgg cagtggaacc gctggtgcag atgctggcac cgctggcacc
gcatattgcg 2640gaagaactgt ggctgcgtct gggcaacacc accagcctgg
cgcatggccc gtttccgaaa 2700gcggatgcgg cgtatctggt ggatgaaacc
gtggaatatc cggtgcaggt gaacggcaaa 2760gtgcgtggtc gtgtggtggt
ggcggcggat accgatgaag aaaccctgaa agcggcggtg 2820ctgaccgatg
aaaaagtgca ggcgtttctg gcgggcgcga ccccgcgtaa agtgattgtg
2880gtggcgggcc gtctggtgaa cctggtgatt taactcgag 2919
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