U.S. patent application number 12/327245 was filed with the patent office on 2009-06-11 for novel pyrrolidine bicyclic compounds and its derivatives, compositions and methods of use.
This patent application is currently assigned to VICURON PHARMACEUTICALS INC.. Invention is credited to Jeffrey JACOBS, Rakesh K. JAIN, Jason G. LEWIS, Dinesh V. PATEL, Zhengyu YUAN.
Application Number | 20090149509 12/327245 |
Document ID | / |
Family ID | 23150431 |
Filed Date | 2009-06-11 |
United States Patent
Application |
20090149509 |
Kind Code |
A1 |
JACOBS; Jeffrey ; et
al. |
June 11, 2009 |
NOVEL PYRROLIDINE BICYCLIC COMPOUNDS AND ITS DERIVATIVES,
COMPOSITIONS AND METHODS OF USE
Abstract
N-[1-oxo-(optionally 2-aza)-2-alkyl-3-(carboxyl or thiol or
hydroxyaminocarbonyl or N-hydroxyformamido)-propyl]-(aryl or
heteroaryl)-azacyclo.sub.4-7alkanes or thiazacyclo.sub.4-7alkanes,
salts or prodrugs thereof have interesting properties, e.g., in the
treatment or prevention of disorders amenable to treatment by PDF
inhibitors, such as treatment of bacterial infections.
Inventors: |
JACOBS; Jeffrey; (San Mateo,
CA) ; JAIN; Rakesh K.; (Fremont, CA) ; LEWIS;
Jason G.; (Hayward, CA) ; PATEL; Dinesh V.;
(Fremont, CA) ; YUAN; Zhengyu; (Fremont,
CA) |
Correspondence
Address: |
LAHIVE & COCKFIELD, LLP;FLOOR 30, SUITE 3000
ONE POST OFFICE SQUARE
BOSTON
MA
02109
US
|
Assignee: |
VICURON PHARMACEUTICALS
INC.
Fremont
CA
|
Family ID: |
23150431 |
Appl. No.: |
12/327245 |
Filed: |
December 3, 2008 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11151131 |
Jun 13, 2005 |
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12327245 |
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10171705 |
Jun 14, 2002 |
6987104 |
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11151131 |
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60298418 |
Jun 15, 2001 |
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Current U.S.
Class: |
514/343 ;
514/365; 514/367; 514/374; 514/423; 546/279.1; 548/152; 548/204;
548/236; 548/540 |
Current CPC
Class: |
C07D 417/04 20130101;
C07D 207/08 20130101; A61P 43/00 20180101; C07D 403/04 20130101;
C07D 401/04 20130101; A61P 31/04 20180101; C07D 413/04
20130101 |
Class at
Publication: |
514/343 ;
548/236; 514/374; 548/152; 514/367; 548/204; 514/365; 546/279.1;
548/540; 514/423 |
International
Class: |
A61K 31/4439 20060101
A61K031/4439; C07D 413/04 20060101 C07D413/04; A61K 31/422 20060101
A61K031/422; C07D 415/00 20060101 C07D415/00; C07D 207/09 20060101
C07D207/09; A61K 31/40 20060101 A61K031/40; A61K 31/428 20060101
A61K031/428; A61K 31/427 20060101 A61K031/427; C07D 401/04 20060101
C07D401/04 |
Claims
1. (canceled)
2. A compound of formula (I): ##STR00044## wherein: R.sub.1 is an
aryl or heteroaryl which is linked to either the .alpha.- or
.beta.-position to the ring nitrogen; R.sub.2 is hydrogen, halogen
or hydroxy; R.sub.3 is hydrogen, halogen, C.sub.1-10 alkyl,
C.sub.1-10 heteroalkyl or (R.sub.2 and R.sub.3) collectively form a
C.sub.4-7 cycloalkyl, provided that when R.sub.3 is halogen,
R.sub.2 is not hydroxy; X is --CH.sub.2--; W is NR.sub.5 or
C.sub.4R.sub.5, wherein R.sub.4 is hydrogen, halogen, C.sub.1-10
alkyl, or C.sub.1-10 heteroalkyl and R.sub.5 is C.sub.1-10 alkyl or
(R.sub.4 and R.sub.5) collectively form a C.sub.4-7 cycloalkyl,
provided that when W is NR.sub.5, R.sub.2 and R.sub.3 are hydrogen,
C.sub.1-10 alkyl or heteroalkyl; Y is --COOH, --SH, --N(OH)--CHO or
--CO--NH(OH), provided that when Y is --N(OH)CHO or --SH, R.sub.2
is hydrogen and R.sub.3 is hydrogen, C.sub.1-10 alkyl or C.sub.1-10
heteroalkyl; n is 2; a pharmaceutically acceptable salt thereof or
a prodrug thereof.
3. The compound of formula (I) according to claim 2, wherein Y is
--CO--NH(OH).
4. (canceled)
5. The compound of formula (I) according to claim 2, wherein
R.sub.1 is hydroxy.
6. The compound of formula (I) according to claim 2, wherein
R.sub.5 is n-butyl.
7. The compound of formula (I) according to claim 2, wherein
R.sub.1 is heteroaryl.
8. The compound of formula (I) according to claim 2, wherein the
heteroaryl is oxazolyl, thiazolyl, pyridinyl and
benzimidazolyl.
9. The compound of formula (I) according to claim 2, wherein the
heteroaryl is linked to the .alpha.-position to the nitrogen of the
azacycloalkane represented in formula (I).
10. The compound of formula (I) according to claim 2, wherein the
heteroaryl is oxazolyl.
11. (canceled)
12. The compound of formula (I) according to claim 2, wherein the
heteroaryl is thiazolyl.
13. (canceled)
14. The compound of formula (I) according to claim 2, wherein the
heteroaryl is pyridinyl.
15. The compound of formula (I) according to claim 2, wherein the
heteroaryl is benzimidazolyl.
16. The compound of formula (I) according to claim 2, wherein
R.sub.2 is hydrogen and W is N.
17. The compound of formula (I) according to claim 16, wherein
R.sub.5 is a substituted lower alkyl.
18. The compound of formula (I) according to claim 16, wherein
R.sub.1 is a heteroaryl.
19. The compound of formula (I) according to claim 16, wherein the
heteroaryl is oxazolyl or thiazolyl.
20. The compound of formula (I) according to claim 16, wherein the
heteroaryl is linked to the .alpha.-position of the azacycloalkane
represented in formula (I).
21-23. (canceled)
24. The compound of formula (I) according to claim 2, wherein
R.sub.1 is an aryl.
25. (canceled)
26. The compound of formula (I) according to claim 2, wherein
R.sub.1 is halogen.
27-34. (canceled)
35. A pharmaceutical composition comprising a compound according to
claim 2 and a pharmaceutically acceptable carrier.
36-38. (canceled)
39. A method for inhibiting bacteria comprising contacting said
bacteria with a bacteria inhibiting amount of a compound according
to claim 2 or a pharmaceutically acceptable salt thereof.
40-41. (canceled)
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. divisional
application Ser. No. 11/151,131 (filed Jun. 13, 2005) which claims
the benefit of U.S. Ser. No. 10/171,705 (filed Jun. 14, 2002),
which in turn claims the benefit of U.S. Provisional Application
Ser. No. 60/298,418 (filed Jun. 15, 2001).
BACKGROUND OF THE INVENTION
[0002] Treatment of microbial infection in host organisms requires
an effective means to kill the microbe while doing as little harm
to the host as possible. Accordingly, agents which target
characteristics unique to a pathology-causing microorganism are
desirable for treatment. Penicillin is an extremely well-known
example of such an agent. Penicillin acts by inhibiting
biosynthesis of bacterial cell walls. Since mammalian cells do not
require cell walls for survival, administration of penicillin to a
human infected with bacteria may kill the bacteria without killing
human cells.
[0003] However, the use of antibiotics and antimicrobials has also
resulted in increased resistance to these agents. As bacteria
become resistant to older, more widely used antimicrobial agents,
new antimicrobials must be developed in order to provide effective
treatments for human and non-human animals suffering from microbial
infection.
[0004] Peptidyl deformylase (PDF) is a metallopeptidase found in
prokaryotic organisms, such as bacteria. Protein synthesis in
prokaryotic organisms begins with N-formyl methionine (fMet). After
initiation of protein synthesis, the formyl group is removed by the
enzyme PDF; this activity is essential for maturation of proteins.
It has been shown that PDF is required for bacterial growth (see
Chang et al., J. Bacteriol., Vol. 171, pp. 4071-4072 (1989);
Meinnel et al., J. Bacteriol., Vol. 176, No. 23, pp. 7387-7390
(1994); Mazel et al., EMBO J., Vol. 13, No. 4, pp. 914-923 (1994)).
Since protein synthesis in eukaryotic organisms does not depend on
fMet for initiation, agents that will inhibit PDF are attractive
candidates for development of new antimicrobial and antibacterial
drugs. Prokaryotic organisms, including disease-causing
prokaryotes, are described in Balows, Truper, Dworkin, Harder and
Schleifer, Eds., The Prokaryotes, 2.sup.nd Ed., Springer-Verlag, NY
(1992); and Holt, Editor-in-Chief, Bergey's Manual of Systematic
Bacteriology, Vols. 1-4, Williams & Wilkins, Baltimore (1982,
1986, 1989).
[0005] PDF is part of the metalloproteinase superfamily. While PDF
clearly shares many of the features which characterize
metalloproteinases, it differs from other members of the
superfamily in several important respects. First, the metal ion in
the active enzyme appears to be Fe (II), or possibly another
divalent cationic metal, instead of the zinc ion more commonly
encountered (see Rajagopalan et al., J. Am. Chem. Soc., Vol. 119,
pp. 12418-12419 (1997)). Second, the divalent ion appears to play
an important role, not only in catalysis, but also in the
structural integrity of the protein. Third, the third ligand of the
divalent ion is a cysteine, rather than a histidine or a glutamate,
as in other metalloproteinases and is not located at the C-terminal
side of the HEXXH motif but far away along the amino acid sequence
and N-terminal to the motif. Finally, the solution structure shows
significant differences in the secondary and tertiary structure of
PDF compared to other prototypical metalloproteinases (see Meinnel
et al., J. Mol. Biol., Vol. 262, pp. 375-386 (1996)). PDF from E.
coli, Bacillus stearothermophilus and Thermus thermophilus have
been characterized (see Meinnel et al., J. Mol. Biol., Vol. 267,
pp. 749-761 (1997)). The enzyme studied by Meinnel et al. contained
a zinc ion as the divalent ion and the structural features
summarized above were obtained from zinc-containing proteins. The
structure of the protein has also been determined by NMR (see
O'Connell et al., J. Biomol., NMR, Vol. 13, No. 4, pp. 311-324
(1999)).
[0006] Metalloproteinases are critical to many aspects of normal
metabolism. The class known as matrix metalloproteinases (MMPs) are
involved in tissue remodeling, such as degradation of the
extracellular matrix. These enzymes are believed to play a role in
normal or beneficial biological events, such as the formation of
the corpus luteum during pregnancy (see Liu et al., Endocrinology,
Vol. 140, No. 11, pp. 5330-5338 (1999)), wound healing (see
Yamagiwa et al., Bone, Vol. 25, No. 2, pp. 197-203 (1999)), and
bone growth in healthy children (see Bord et al., Bone, Vol. 23,
No. 1, pp. 7-12 (1998)). Disorders involving metalloproteinases
have been implicated in several diseases such as cancer, arthritis
and autoimmune diseases.
[0007] Because of the importance of MMPs in normal physiological
processes, it would be preferable to develop agents that inhibit
PDF, a metalloproteinase present only in prokaryotes, while
avoiding significant inhibition of MMPs. Alternatively, PDF
inhibitors which also inhibit MMPs may be of use where the
therapeutic benefits of inhibiting PDF outweigh the risk of side
effects from MMP inhibition.
[0008] A wide variety of compounds have been developed as candidate
inhibitors of MMPs and other metalloproteinases, and much effort
has also been directed at synthetic methods for these compounds and
related compounds (see Izquierdo-Martin et al., J. Am. Chem. Soc.,
Vol. 114, pp. 325-331 (1992); Cushman et al., Chapter 5, "Specific
Inhibitors of Zinc Metallopeptidases", Topics in Molecular
Pharmacology, Burgen & Roberts, Eds. (1981); Mohler et al.,
Nature, Vol. 370, pp. 218-220 (1994); Gearing et al., Nature, Vol.
370, pp. 555-557 (1994); McGeehan et al., Nature, Vol. 370, pp.
558-561 (1994); U.S. Pat. Nos. 4,052,511, 4,303,662, 4,311,705,
4,321,383, 4,599,361, 4,804,676, 5,128,346, 5,256,657, 5,268,384,
5,447,929, 5,453,423, 5,552,419, 5,614,625, 5,643,908, 5,712,300
and 5,869,518; European patent publications EP 236872, EP 274453,
EP 334244, EP 423943, EP 489577, EP 489579, EP 497192, EP 574758;
and International PCT Patent Applications Publication Nos. WO
90/05716, WO 90/05719, WO 91/02716, WO 92/13831, WO 92/22523, WO
93/09090, WO 93/09097, WO 93/20047, WO 93/24449, WO 93/24475, WO
94/02446, WO 94/02447, WO 94/21612, WO 94/25434, WO 94/25435, WO
95/33731, WO 96/25156, WO 96/26918 WO 97/30707, WO 97/49674, WO
98/55449 and WO 99/02510.
[0009] Research on inhibitors of PDF is much less extensive than
that for inhibitors of MMPs. N-formyl hydroxylamine derivatives are
described in International Patent Application WO 99/39704. Peptide
aldehyde inhibitors of PDFs are described in Durand et al., Arch.
Biochem. Biophys., Vol. 367, No. 2, pp. 297-302 (1999). The PDF
inhibitor
(S)-2-O--(H-phosphonoxy)-L-caproyl-L-leucyl-p-nitroanilide is
described in Hao et al., Biochem., Vol. 38, pp. 4712-4719 (1999),
and peptidyl H-phosphonate inhibitors of PDF are discussed in Hu et
al., Bioorg. Med. Chem. Lett., Vol. 8, pp. 2479-2482 (1998).
Formylated peptides and pseudopeptides are described in Meinnel et
al., Biochem., Vol. 38, No. 1, pp. 4288-4295 (1999) as inhibitors
of PDF.
[0010] In view of the importance of identifying new antibiotics to
treat bacteria resistant to existing antibiotics, and the
relatively small amount of work that has been carried out on PDF
inhibitors, it is desirable to develop novel inhibitors of PDF for
evaluation and use as antibacterial and antimicrobial agents. The
present invention fulfills this need.
SUMMARY OF THE INVENTION
[0011] In particular, the present invention provides an
N-[1-oxo-(optionally 2-aza)-2-alkyl-3-(carboxyl or thiol or
hydroxyaminocarbonyl or N-hydroxyformamido)-propyl]-(aryl or
heteroaryl)-azacyclo.sub.4-7alkane or thiazacyclo.sub.4-7alkane
(referred to herein collectively as "compounds of the invention"),
a salt thereof or a prodrug thereof, e.g., a compound of formula
(I):
##STR00001##
wherein [0012] R.sub.1 is aryl or heteroaryl which is linked to
either the .alpha.- or .beta.-position to the ring nitrogen; [0013]
R.sub.2 is hydrogen, halogen or hydroxy; [0014] R.sub.3 is
hydrogen, halogen, C.sub.1-10alkyl, C.sub.1-10heteroalkyl or
(R.sub.2 and R.sub.3) collectively form a C.sub.4-7cycloalkyl,
provided that when R.sub.3 is halogen, R.sub.2 is not hydroxy;
[0015] X is --CH.sub.2-- or S; [0016] W is NR.sub.5 or
CR.sub.4R.sub.5, wherein R.sub.4 is hydrogen, halogen,
C.sub.1-10alkyl, or C.sub.1-10heteroalkyl and [0017] R.sub.5 is
C.sub.1-10alkyl or (R.sub.4 and R.sub.5) collectively form a
C.sub.4-7cycloalkyl, provided that when W is NR.sub.5, [0018]
R.sub.2 and R.sub.3 are hydrogen, C.sub.1-10alkyl or heteroalkyl;
[0019] Y is --COOH, --SH, --N(OH)--CHO or --CO--NH(OH), provided
that when Y is --N(OH)CHO or --SH, R.sub.2 is hydrogen and R.sub.3
is hydrogen, C.sub.1-10alkyl or C.sub.1-10heteroalkyl; [0020] n is
0 to 3, provided that when n is 0, X is --CH.sub.2--; a salt
thereof or a prodrug thereof.
DETAILED DESCRIPTION OF THE INVENTION
[0021] Unless otherwise stated, the following terms as used in the
specification have the following meaning.
[0022] The term "cycloalkane" or "cycloalkyl" is a cyclic saturated
alkyl group containing from 3- to 6-ring carbon atoms, and is,
e.g., cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.
[0023] The term "azacyclo.sub.4-7alkane" contains 1-ring heteroatom
which is a nitrogen. It contains from 4-7, and preferably 5-ring
atoms including the heteroatom.
[0024] The term "thiazacyclo.sub.4-7alkane" contains 2-ring
heteroatoms, nitrogen and sulfur. It contains from 4-7, and
especially 5-ring atoms including the heteroatom.
[0025] The azacyclo.sub.4-7alkane and thiazacyclo.sub.4-7alkane are
substituted at either the .alpha.- or .beta.-position to the
nitrogen of the ring by a heteroaryl or aryl as defined below.
[0026] The term "alkyl" refers to saturated and unsaturated
aliphatic groups, cycloalkyl or substituted alkyl including
straight-chain, branched-chain and cyclic groups having from 1-10
carbons atoms, and is preferably a saturated lower alkyl having
from 1-7 carbon atoms, and especially 1-4 carbon atoms. Examples of
alkyl include, but are not limited to, methyl, ethyl, n-propyl,
isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, n-pentyl,
neopentyl, n-hexyl or n-heptyl, cyclopropyl and especially
n-butyl.
[0027] The term "substituted alkyl" refers to an alkyl group that
is substituted with one or more substitutents preferably 1-3
substitutents including, but not limited to substituents, such as
halogen, lower alkoxy, hydroxy, mercapto, carboxy, cycloalkyl,
aryl, heteroaryl and the like. Examples of substituted alkyl groups
include, but are not limited to, --CF.sub.3, --CF.sub.2--CF.sub.3,
hydroxymethyl, 1- or 2-hydroxyethyl, methoxymethyl, 1- or
2-ethoxyethyl, carboxymethyl, 1- or 2-carboxyethyl,
cyclopentylethyl and the like.
[0028] The term "aryl" or "Ar" refers to an aromatic carbocyclic
group of 6-14 carbon atoms having a single ring including, but not
limited to, groups, such as phenyl; or multiple condensed rings
including, but not limited to, groups, such as naphthyl or anthryl;
and is especially phenyl. An aryl group can be unsubstituted or
substituted with one or more substituents, preferably 1-3
substituents including, but not limited to, groups, such as lower
alkyl, halogen and lower alkoxy.
[0029] The term "heteroaryl" or "HetAr" refers to a 4- to
7-membered, monocyclic aromatic heterocycle or a bicycle that is
composed of a 4- to 7-membered, monocyclic aromatic heterocycle and
a fused-on benzene ring. The heteroaryl has at least one hetero
atom, preferably one or two heteroatoms including, but not limited
to, heteroatoms, such as N, O and S, within the ring.
Representative examples include, but are not limited to, oxazolyl,
thiazolyl, pyridinyl and imidazolyl, benzimidazolyl, isoxazolyl,
benzthiazolyl and the like. The heteroaryl may be unsubstituted or
substituted by one or more substituents including, but not limited
to lower alkyl, halo-lower alkyl, halogen, hydroxy, lower alkoxy,
aryl (preferably aryl) and the like.
[0030] The term "heteroalkyl" refers to a saturated or unsaturated
alkyl as defined above, having from 1-10 carbon atoms, and
especially a saturated lower heteroalkyl of 1-4 carbon atoms which
contain one or more heteroatoms, as part of the main, branched or
cyclic chains in the group. Heteroatoms are independently selected
from the group consisting of --NR-- where R is hydrogen or alkyl,
--S--, --O-- and --P--; preferably --NR-- where R is hydrogen or
alkyl and/or --O--. Heteroalkyl groups may be attached to the
remainder of the molecule either at a heteroatom (if a valence is
available) or at a carbon atom. Examples of heteroalkyl groups
include, but are not limited to, groups, such as --O--CH.sub.3,
--CH.sub.2--O--CH.sub.3, --CH.sub.2--CH.sub.2--O--CH.sub.3,
--S--CH.sub.2--CH.sub.2--CH.sub.3,
--CH.sub.2--CH(CH.sub.3)--S--CH.sub.3 and
--CH.sub.2--CH.sub.2--NH--CH.sub.2--CH.sub.2--.
[0031] The heteroalkyl group may be unsubstituted or substituted
with one or more substituents, preferably 1-3 substituents
including, but not limited to, alkyl, halogen, alkoxy, hydroxyl,
mercapto, carboxy and phenyl. The heteroatom(s) as well as the
carbon atoms of the group may be substituted. The heteroatom(s) may
also be in oxidized form.
[0032] The term "alkoxy" as used herein refers to a C.sub.1-10alkyl
linked to an oxygen atom, or preferably a saturated lower alkoxy
having from 1-7 carbon atoms. Examples of alkoxy groups include,
but are not limited to, groups, such as methoxy, ethoxy,
tert-butoxy and allyloxy.
[0033] The term "halogen" or "halo" as used herein refer to
chlorine, bromine, fluorine, iodine, and is especially
fluorine.
[0034] "Protecting group" refers to a chemical group that exhibits
the following characteristics: 1) reacts selectively with the
desired functionality in good yield to give a protected substrate
that is stable to the projected reactions for which protection is
desired; 2) is selectively removable from the protected substrate
to yield the desired functionality; and 3) is removable in good
yield by reagents compatible with the other functional group(s)
present or generated in such projected reactions. Examples of
suitable protecting groups may be found in Greene et al.,
Protective Groups in Organic Synthesis, 2.sup.nd Ed., John Wiley
& Sons, Inc., NY (1991). Preferred amino protecting groups
include, but are not limited to, benzyloxycarbonyl (CBz),
t-butyl-oxycarbonyl (Boc), t-butyldimethylsilyl (TBDMS),
9-fluorenylmethyl-oxycarbonyl (Fmoc), or suitable photolabile
protecting groups, such as 6-nitroveratryloxy carbonyl (Nvoc),
nitropiperonyl, pyrenylmethoxycarbonyl, nitrobenzyl, dimethyl
dimethoxybenzyl, 5-bromo-7-nitroindolinyl and the like. Preferred
hydroxyl protecting groups include Fmoc, TBDMS, photolabile
protecting groups, such as nitroveratryl oxymethyl ether (Nvom)),
Mom (methoxy methyl ether) and Mem (methoxy ethoxy methyl ether).
Particularly preferred protecting groups include NPEOC
(4-nitrophenethyloxycarbonyl) and NPEOM
(4-nitrophenethyloxy-methyloxycarbonyl).
[0035] It will be appreciated that the compounds of the invention,
e.g., the compounds of formula (I), may exist in the form of
optical isomers, racemates or diastereoisomers. For example, a
compound of formula (I) wherein R.sub.2 and R.sub.3 are different
residues, is asymmetric and may have the R- or S-configuration. It
is to be understood that the present invention embraces all
enantiomers and their mixtures. Similar considerations apply in
relation to starting materials exhibiting asymetric carbon atoms as
mentioned.
[0036] The compounds of the invention, e.g., the compounds of
formula (I), may exist in free form or in salt form, e.g., in form
of a pharmaceutically acceptable salt. A "pharmaceutically
acceptable salt" of a compound means a physiologically and
pharmaceutically acceptable salt that possesses the desired
pharmacological activity of the parent compound and does not impart
undesired toxicological effects. Such salts include:
1. Acid addition salts, formed with inorganic acids, such as
hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,
phosphoric acid and the like; or formed with organic acids, such as
acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic
acid, glycolic acid, pyruvic acid, lactic acid, malonic acid,
succinic acid, malic acid, maleic acid, fumaric acid, tartaric
acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid,
cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic
acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid,
benzenesulfonic acid, 4-chlorobenzenesulfonic acid,
2-napthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic
acid, 3-phenylpropionic acid, trimethylacetic acid, tertiary
butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic
acid, hydroxynapthoic acid, salicylic acid, stearic acid, muconic
acid and the like; or 2. Salts formed when an acidic proton present
in the parent compound either is replaced by a metal ion, e.g., an
alkali metal ion, an alkaline earth ion or an aluminum ion; or
coordinates with an organic base, such as ethanolamine,
diethanolamine, triethanolamine, tromethamine, N-methylglucamine
and the like.
[0037] A compound of the invention, e.g., a compound of formula
(I), may act as a prodrug. Prodrug means any compound which
releases an active parent drug according to formula (I) in vivo
when such prodrug is administered to a mammalian subject. Prodrugs
of a compound of formula (I) are prepared by modifying functional
groups present in the compound of formula (I) in such a way that
the modifications may be cleaved in vivo to release the parent
compound. Prodrugs include compounds of formula (I) wherein a
hydroxy, amino or sulfhydryl group in compound (I) is bonded to any
group that may be cleaved in vivo to regenerate the free hydroxyl,
amino or sulfhydryl group, respectively. Examples of prodrugs
include, but are not limited to, esters, e.g., acetate, formate and
benzoate derivatives; carbamates, e.g., N,N-dimethylamino-carbonyl;
of hydroxy functional groups in compounds of formula (I) and the
like.
[0038] In the compounds of the invention, e.g., the compounds of
formula (I), the following significances are preferred individually
or in any subcombination: [0039] 1. Y is COOH, SH, --CO--NH(OH) or
--N(OH)CHO, preferably --CO--NH(OH). [0040] 2. X is --CH.sub.2--.
[0041] 3. R.sub.2 is hydroxy, fluorine or hydrogen. [0042] 4.
R.sub.3 is hydrogen. [0043] 5. W is CR.sub.4R.sub.5 wherein R.sub.4
is hydrogen and R.sub.5 is hydrogen or C.sub.1-10alkyl, preferably
n-butyl; [0044] 6. n is 1. [0045] 7. R.sub.1 is phenyl or
heteroaryl. [0046] 8. Heteroaryl as R.sub.1 is oxazolyl, thiazolyl,
pyridinyl and benzimidazolyl. When heteroaryl is oxazolyl, the
oxazolyl may be substituted with a lower alkyl, especially methyl.
When the heteroaryl is thiazolyl, the thiazolyl may be substituted
by one or two substituents selected from the group consisting of
lower alkyl and phenyl. Preferably R.sub.1 is oxazolyl or
methyloxazolyl. [0047] 9. R.sub.1 is preferably linked to the
.alpha.-position of the azacycloalkane represented in formula
(I).
[0048] Compounds of the invention, e.g., the compounds of formula
(I), may be prepared in accordance with methods well-known in the
art of organic chemistry. Compounds of formula I may be prepared by
reacting a compound of formula II
##STR00002##
wherein X, R.sub.1, R.sub.2, R.sub.3, W and n are as defined above
and Y.sub.a is COOH or a functional derivative thereof, with a
hydroxylaminating agent, e.g., NH.sub.2OH and, where required,
converting the resulting compounds obtained in free form into salt
forms or vice versa.
[0049] Functional derivatives of COOH as Y.sub.a are, e.g.,
halogenides, e.g., acid chloride, esters or acid anhydride.
[0050] Above reaction may be carried out according to methods known
in the art or as disclosed in Schemes A to K and in the examples
below.
[0051] Insofar as the production of starting materials is not
particularly described, the compounds are known or may be prepared
analogously to methods known in the art or as disclosed in the
examples hereinafter.
[0052] The following abbreviations are used: [0053] AcOH=acetic
acid [0054] BuLi=n-butyl lithium [0055] DAST=diethylaminosulfur
trifluoride [0056] DCC=dicyclohexylcarbodiimide [0057]
DCE=dichloroethane [0058] DCM=dichloromethane [0059]
DIC=diisopropylcarbodiimide [0060] DIAD=diisopropylazodicarboxylate
[0061] DIEA=diisopropylethylamine [0062] DME=1,2-dimethoxyethane
[0063] DMF=dimethylformamide [0064] DMSO=dimethylsulfoxide [0065]
EDC=N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide [0066]
EtOAc=ethyl acetate [0067]
HATU=O-(7-aza-benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate [0068] LDA=lithium diisopropylamine [0069]
MeOH=methanol [0070] NaHMDS=sodium hexamethyldisilazide [0071]
PyBOP=benzotriazole-1-yl-oxy-tris-pyrrolidinophosphonium
hexafluorophosphate [0072] rt=room temperature [0073]
TEA=triethylamine [0074] TFA=trifluoroacetic acid [0075]
THF=tetrahydrofuran [0076] p-TSA=p-toluenesulfonic acid [0077]
TMSCl=trimethylsilyl chloride
General Procedure A
Synthesis of N-hydroxy-3-aminocarbonylpropionamide
##STR00003##
[0079] Step 1: To a solution of 4-(S)-benzyloxazolidin-2-one (56
mmol) (Aldrich, Milwaukee, Wis.) in THF at -78.degree. C. is added
2.5 M n-BuLi in hexane (22.4 mL, 56 mmol) and the reaction is
stirred at -78.degree. C. for 2 hours. To this is added via cannula
a -78.degree. C. solution of acid chloride A-1 (R=hexanoyl, 65
mmol) in THF and the mixture is stirred at -78.degree. C. for 2
hours, then allowed to warm to rt and stirred overnight. The
reaction is then quenched with aqueous saturated NH.sub.4Cl,
extracted with EtOAc, dried and purified by silica gel
chromatography (hexanes/EtOAc) to afford
N-hexanoyl-4-(S)-benzyloxazolidin-2-one (A-2).
[0080] Step 2: To a solution of
N-hexanoyl-4-(S)-benzyloxazolidin-2-one A-2 (7.3 mmol) in THF at
-78.degree. C. is added 1.0 M NaHMDS (8.8 mmol) and the reaction
stirred at -78.degree. C. for 1 hour. A solution of methyl
bromoacetate (8.8 mmol) in THF is then added dropwise, and the
resulting mixture is stirred at -78.degree. C. for 1 hour and then
at rt overnight. The reaction is quenched with NH.sub.4Cl,
concentrated, then suspended in EtOAc and washed with 0.5 N HCl and
brine, dried and purified by silica gel chromatography
(EtOAc/hexanes) to afford the methyl
3-(R)-(n-butyl)-3-[4-(S)-benzyloxazolidin-2-one-3-ylcarbonyl)propi-
onate (A-3).
[0081] Step 3: To methyl
3-(R)-(n-butyl)-3-[4-(S)-benzyloxazolidin-2-one-3-ylcarbonyl)-propionate
A-3 (1.44 mmol) in THF/water at 0.degree. C. is added 30%
H.sub.2O.sub.2 (5.76 mmol) and solid lithium hydroxide (1.44 mmol)
and the reaction is stirred at 0.degree. C. for 3 hours. The
reaction is then quenched with 2.0 M Na.sub.2SO.sub.3,
concentrated, suspended in EtOAc and subjected to standard aqueous
workup. The crude product is purified by silica gel chromatography
(MeOH/DCM) to afford methyl 3-(R)-(n-butyl)-propionate (A-4).
[0082] Step 4: To a solution of mono-protected succinate, e.g.,
mono-4-methyl 2-(R)-butylsuccinic acid A-4 (1 mmol) in DMF is added
amine A-5 (1 mmol), DIEA (0.4 mL, 2.3 mmol), and an activating
reagent (e.g., EDC, PyBOP, DIC, DCC, etc.; 1 mmol). The mixture is
stirred overnight, then diluted with EtOAc and washed with aqueous
HCl (1 N), water, saturated NaHCO.sub.3, brine and then dried
(Na.sub.2SO.sub.4). The filtrate is concentrated and then purified
on silica gel (Merck 60; EtOAc/hexane) to afford
3-aminocarbonylpropionate A-6.
[0083] Step 5: 3-aminocarbonylpropionate A-6 (0.1 mmol) is treated
with dioxane (1 mL) and hydroxylamine (50% in water, 2 mL) for 1-3
days, and then purified by preparative reverse-phase (C18) HPLC to
afford the desired N-hydroxy-3-aminocarbonylpropionamide (A-7).
General Procedure B
Synthesis of
2(S)-hydroxy-3(R)-[2(S)-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
[0084] Step 1: To a solution of diisopropylamine (14 mL, 100 mmol)
in THF at 0.degree. C. is added BuLi (2.5 M in hexane, 40 mL, 100
mmol) over 10 minutes. The mixture is stirred at rt for 30 minutes,
and then added via cannula to a -78.degree. C. solution of dimethyl
malate B-1 (7.71 g, 47.6 mmol) in THF (130 mL). The mixture is
warmed to -20.degree. C. over 2 hours, and then cooled to
-78.degree. C. Crotyl bromide (8.1 g, 60 mmol) is added, then the
mixture is allowed to warm to rt and then stirred overnight. The
solution is then cooled to -10.degree. C. and quenched with
NH.sub.4Cl (10%, 100 mL). The THF is removed and the residue
extracted with EtOAc (2.times.200 mL). The combined organic layers
are washed with HCl (1 N, 3.times.50 mL), saturated aqueous
NaHCO.sub.3 (3.times.50 mL), and brine, then dried over
Na.sub.2SO.sub.4. The solution is filtered and concentrated to give
a residue, which is purified on silica gel (EtOAc/hexane 1:4) to
afford (2S,3R)-3-(2-butenyl)-2-hydroxysuccinic dimethyl ester
B-2.
##STR00004##
[0085] Step 2: To (2S,3R)-3-(2-butenyl)-2-hydroxysuccinic dimethyl
ester B-2 (2.5 g) in EtOAc (50 mL) is added 10% Pd/C (0.25 g) and
the reaction stirred under a hydrogen atmosphere for 20 hours. The
suspension is filtered through Celite, washed with EtOAc (3.times.)
and then concentrated in vacuo to afford
(2S,3R)-3-(n-butyl)-2-hydroxysuccinic dimethyl ester B-3.
[0086] Step 3: To (2S,3R)-3-(n-butyl)-2-hydroxysuccinic dimethyl
ester B-3 in MeOH (28 mL) is added a solution of NaOH (2.2 g, 55
mmol) in water (28 mL). After 24 hours, the MeOH is removed, the
crude reaction is acidified with HCl (6 N, 12 mL) to pH=1, and then
extracted with EtOAc (3.times.50 mL). The combined organic layers
are dried (Na.sub.2SO.sub.4) and concentrated to give
(2S,3R)-3-(n-butyl)-2-hydroxysuccinic acid B-4.
[0087] Step 4: To a solution of
(2S,3R)-3-(n-butyl)-2-hydroxysuccinic acid B-4 (300 mg, 1.58 mmol)
in 2,2-dimethoxypropane (10 mL) is added p-TSA (20 mg) and the
reaction is stirred at rt for 16 hours. The solution is diluted
with DCM and washed with brine, dried (Na.sub.2SO.sub.4) and then
purified by silica gel chromatography to afford 1.2 mmol
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5.
[0088] Step 5: To a solution of
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 (1.2
mmol) in DMF (10 mL) is added bicyclic pyrrolidine A-5 (1.2 mmol),
HATU (1.2 mmol), and DIEA (2.5 mmol). The mixture is stirred
overnight, then concentrated and purified on silica gel
(EtOAc/hexane 1:4) to afford 275 mg of the desired amide B-6.
[0089] Step 6: To a cold solution of B-6 in dioxane (5 mL) 50%
aqueous hydroxylamine is added (400 .mu.L), and the solution
stirred at 4.degree. C. for 8 hours. The crude reaction mixture is
then purified by preparative reverse-phase (C18) HPLC to afford
2(S)-hydroxy-3(R)-[2(S)-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
acid hydroxamide B-7.
General Procedure C
Synthesis of
2-fluoro-3-(R)-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00005##
[0091] Step 1: To
2,2-dimethyl-5-[2(S)-oxazol-2-yl-pyrrolidin-1-carbonyl)-pentyl]-[1,3]diox-
olan-4-one B-6 (5 mmol, 50%) (5 mmol) in MeOH (20 mL) is added
sodium methoxide (catalytic; pH adjusted to 10) and the solution
stirred for 1 hour. Amberlite IR-120 resin (H.sup.+ form) is added,
then the solution is filtered and concentrated to afford
2(S)-hydroxy-3-(2(S)-oxazol-2-yl-pyrrolidin-1-carbonyl)-heptanoic
acid methyl ester C-1.
[0092] Step 2: To
2(S)-hydroxy-3-(2(S)-oxazol-2-yl-pyrrolidin-1-carbonyl)-heptanoic
acid methyl ester C-1 (5 mmol) in DCM (5 mL) is added DAST (15
mmol) at -20.degree. C. The solution is stirred for 16 hours at rt.
The reaction mixture is washed with aqueous NaHCO.sub.3 and brine,
dried (Na.sub.2SO.sub.4) and concentrated then purified on silica
gel (EtOAc/hexanes) to afford
2-(R/S)-fluoro-3-(2(S)-oxazol-2-yl-pyrrolidin-1-carbonyl)-heptanoic
acid methyl ester C-2.
[0093] 1H NMR analysis of this product suggested approximately 1:2
ratio of SIR diastereomers. The two isomers are separated by silica
gel column chromatography.
[0094] Step 3: To intermediate C-2 (0.15 mmol, each isomer is
treated separately) in dioxane (1 mL), aqueous 50% hydroxylamine is
added (0.5 mL) and the reaction stirred for 16 hours at 5.degree.
C. The crude reaction mixture is then purified by preparative
reverse-phase (C18) HPLC to afford
2-fluoro-3-(R)-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptano- ic
acid hydroxamide C-3.
General Procedure D
N-hydroxy-N-{2-R-[2-S-(oxazol-2-yl)-pyrrolidine-1-carbonyl]-hexyl}-formami-
de
##STR00006## ##STR00007##
[0095] Step 1: 2-n-butyl Acrylic Acid (D-2) (R=n-butyl) is Prepared
as Indicated Above
Step 2: 4-benzyl-3-(2-butyl-acryloyl)-oxazolidin-2-one (D-3)
[0096] 2-n-butyl acrylic acid (9.90 g, 77.2 mmol, 1 equiv.) is
dissolved in dry THF (260 ml) and cooled to -78.degree. C. under a
blanket of nitrogen. Hunig's base (17.5 mL, 100.4 mmol, 1.3 equiv.)
and pivaloyl chloride (9.5 mL, 77.2 mmol, 1 equiv.) are added at
such a rate that the temperature remained below -60.degree. C. The
mixture is stirred at -78.degree. C. for 30 minutes, warmed to rt
for 2 hours, and finally cooled back to -78.degree. C.
[0097] In a separate flask, (S)-(-)-4-benzyl-2-oxazolidinone (13.49
g, 77.24 mmol) is dissolved in dry THF (150 mL) and cooled to
-78.degree. C. under nitrogen. BuLi (2.5 M solution in hexanes,
30.9 mL, 77.2 mmol, 1 equiv.) is added slowly at -78.degree. C.,
and the mixture is stirred for 30 minutes at rt. The resulting
anion is slowly transferred via a cannula into the original
reaction vessel. The mixture is allowed to warm to rt and is
stirred overnight at rt. The reaction is quenched with 1 M
KHCO.sub.3, and the solvents are removed under reduced pressure.
The residue is partitioned between EtOAc and water. The organic
layer is washed with brine, dried over anhydrous Na.sub.2SO.sub.4,
filtered and concentrated to give a yellow oil which is purified by
flash chromatography (hexane:EtOAc=4:1) to yield the title compound
D-3 as a white solid.
[0098] 1H NMR (CDCl.sub.3): .delta. 7.39-7.20 (m, 5H), 5.42-5.40
(d, J=7.14 Hz, 2H), 4.76-4.68 (m, 1H), 4.29-4.156 (m, 2H),
3.40-3.35 (dd, J=3.57, 13.46 Hz, 1H), 2.86-2.79 (dd, J=9.34, 13.46
Hz, 1H), 2.42-2.37 (t, J=7.69 Hz, 2H), 1.55-1.30 (m, 4H),
0.951-0.904 (t, J=7.14 Hz, 3H). ES-MS: calcd. For
C.sub.17H.sub.21NO.sub.3 (287.35). found: 288.5 [M+H].
Step 3:
4-benzyl-3-[2-(benzyloxyamino-methyl)-hexanoyl]-oxazolidin-2-one
(p-toluenesulfonic Acid Salt)
[0099] Compound D-3 (8.25 g, 28.7 mmol) is mixed with
O-benzylhydroxylamine (7.07 g, 57.4 mmol, 2 equiv.) and stirred for
40 hours at rt under nitrogen. The mixture is dissolved in EtOAc
and p-TSA (21.84 g, 114.8 mmol, 4 equiv.) is added to precipitate
excess O-benzylhydroxylamine as a white solid. The white solid is
filtered off, and the filtrate is concentrated to give a crude
yellow oil. Charging the crude yellow oil with excess diethyl ether
and cooling to 0.degree. C. for 30 minutes gave a solid which is
collected by filtration and dried in vacuo to afford the title
compound as a white crystalline solid (single diastereomer).
[0100] 1H NMR (CDCl.sub.3): .delta. 8.07-8.04 (d, J=8.24 Hz, 2H),
7.59-7.39 (m, 10H), 7.18-7.15 (d, J=7.69 Hz, 2H), 5.49-5.40 (q,
J=8.61 Hz, 2H), 4.65-4.56 (m, 1H), 4.25-4.08 (m, 3H), 3.83-3.79 (d,
J=13.46 Hz, 1H), 3.15-3.11 (d, J=13.46 Hz, 1H), 2.56 (s, 3H),
1.83-1.67 (m, 4H), 1.40 (bs, 4H), 1.00-0.951 (t, J=6.87, 3H).
ES-MS: calcd. For
C.sub.24H.sub.30N.sub.2O.sub.4*C.sub.7H.sub.8O.sub.3S (582.71).
found: 411.7 [M+H] free base.
Step 4:
4-benzyl-3-[2-(benzyloxyamino-methyl)-hexanoyl]-oxazolidin-2-one
(D-5)
[0101] To a solution of p-TSA salt (22.9 g, 39.3 mmol) dissolved in
EtOAc (400 mL), is added 1 M Na.sub.2CO.sub.3 (200 mL, 5 equiv.)
and stirred at rt for 30 minutes. The layers are separated, and the
aqueous layer extracted with EtOAc. The combined organic layers are
dried over anhydrous Na.sub.2SO.sub.4, filtered and concentrated to
give the title compound as a pale opaque oil.
[0102] 1H NMR (CDCl.sub.3): .delta. 7.57-7.38 (m, 10H), 4.98-4.90
(m, 2H), 4.87-4.79 (m, 1H), 4.38-4.28 (m, 3H), 3.64-3.57 (dd,
J=9.21, 12.64 Hz, 1H), 3.46-3.36 (td, J=3.76, 13.05 Hz, 2H),
2.68-2.60 (dd, J=10.03, 13.46 Hz, 1H), 1.90-1.88 (m, 1H), 1.78-1.71
(m, 1H), 1.51-1.44 (m, 4H), 1.10-1.06 (t, J=6.73 Hz, 3H). ES-MS:
calcd. For C.sub.24H.sub.30N.sub.2O.sub.4 (410.51). found: 411.7
[M+H].
Step 5:
N-[2-(4-benzyl-2-oxo-oxazolidine-3-carbonyl)-hexyl]-N-benzyloxy-fo-
rmamide (D-6)
[0103] A solution of compound D-5 (5.38 g, 13.1 mmol, 1 equiv.) in
formic acid (7.4 mL, 196.6 mmol, 15 equiv.) is cooled to 0.degree.
C. under nitrogen. In a separate flask, formic acid (7.4 ml, 196.6
mmol, 15 equiv.) is cooled to 0.degree. C. under nitrogen, and
acetic anhydride (2.47 mL, 26.2 mmol, 2 equiv.) is added dropwise.
The solution is stirred at 0.degree. C. for 15 minutes. The
resulting mixed anhydride is slowly transferred via syringe into
the original reaction vessel. The mixture is stirred at 0.degree.
C. for 1 hour, then at rt for 3 hours. The mixture is concentrated,
taken up in DCM, and washed successively with saturated NaHCO.sub.3
and brine. The organic layer is dried over anhydrous
Na.sub.2SO.sub.4, filtered and concentrated to give an opaque oil
which is purified by flash chromatography (hexane:EtOAc=2:1 then
DCM:acetone=9:1) to yield the title compound as a colorless
oil.
[0104] 1H NMR (CDCl.sub.3, rotamers): .delta. 8.38 (s, 0.7H), 8.21
(s, 0.3H), 7.54-7.35 (m, 10H), 5.0-5.00 (m, 2H), 4.88-4.81 (m, 1H),
4.39-4.29 (m, 4H), 4.07-4.03 (m, 1H), 3.43-3.39 (m, 1H), 2.66-2.58
(m, 1H), 1.89 (bs, 1H), 1.73 (bs, 1H), 1.49-1.44 (m, 3H), 1.10-1.06
(t, J=6.73 Hz, 3H). ES-MS: calcd. For
C.sub.25H.sub.30N.sub.2O.sub.5 (438.52). found: 439.7 [M+H].
Step 6: 2-[(benzyloxy-formyl-amino)-methyl]-hexanoic acid (D-7)
[0105] Compound D-6 (0.163 g, 0.372 mmol, 1 equiv.) is dissolved in
THF (4.5 mL) and water (1.5 mL) and cooled to 0.degree. C. Hydrogen
peroxide (30% in water, 228 .mu.L, 2.23 mmol, 6 equiv.) is added
dropwise followed by the slow addition of a solution of lithium
hydroxide (0.019 g, 0.446 mmol, 1.2 equiv.) in water (350 .mu.L).
The resulting mixture is stirred at 0.degree. C. for 1.5 hours. The
basic reaction mixture is quenched with Amberlite IR-120 resin
(H.sup.+) to pH 4-5 at 0.degree. C. The resin is filtered off and
rinsed with EtOAc. The mixture is concentrated to remove THF, and
then taken up in EtOAc. The aqueous layer is separated, and the
organic layer dried over anhydrous Na.sub.2SO.sub.4, filtered and
concentrated to give an opaque oil which is purified by flash
chromatography (DCM:acetone=4:1 then acetone:MeOH=99:1) to yield
the title compound D-7 as a colorless oil.
[0106] 1H NMR (DMSO-d.sub.6, rotamers): .delta. 11.2 (s, 1H), 8.20
(s, 0.2H), 7.95 (s, 0.8H), 7.33-7.41 (m, 5H), 4.87 (s, 2H), 3.71
(bs, 2H), 2.50 (bs, 1H), 1.35-1.45 (m, 2H), 1.14-1.28 (m, 4H),
0.857-0.813 (t, J=13.1 Hz, 3H). ES-MS: calcd. For
C.sub.15H.sub.21NO.sub.4 (279.33). found: 278.5 [M-H], 302.5
[M+Na].
Step 7:
1-{2-[(benzyloxy-formyl-amino)-methyl]-hexanoyl}-pyrrolidine-2-car-
boxylic Acid Amide
[0107] To a solution of compound D-7 (0.190 g, 0.680 mmol, 1
equiv.) in dry dioxane (4 mL) at rt under nitrogen is added
successively Hunig's base (391 .mu.L, 2.24 mmol, 3.3 equiv.),
compound A-5 (0.748 mmol, 1.1 equiv.) and HATU (0.284 g, 0.748
mmol, 1.1 equiv.). The resulting mixture is stirred at rt for 22
hours. The mixture is partitioned between EtOAc and 10% citric
acid. The organic layer is washed with brine and saturated
NaHCO.sub.3, dried over anhydride Na.sub.2SO.sub.4, filtered and
concentrated. The residue is purified by flash chromatography
(DCM:acetone=3:1) to give the title compound as a colorless
oil.
Step 8:
{2-[(formyl-hydroxy-amino)-methyl]-hexanoyl}-pyrrolidine-2-carboxy-
lic Acid Amide (D-8)
[0108] Pd--C (0.059 g, 0.1 equiv.) is added to a solution of above
compound (0.550 mmol, 1 equiv.) in a 1:1 EtOAc/ethanol solution (12
mL) under nitrogen. The mixture is stirred under hydrogen
atmosphere for 36 hours. The catalyst is removed by filtration
through Celite. The filtrate is concentrated, and the residue is
purified by preparative TLC (DCM:acetone=2:1) to give the title
compound as an amorphous solid.
General Procedure E
Synthesis of 2-pyrrolidine-2-yl-oxazole (Hydrobromic Acid Salt)
##STR00008##
[0110] 2-(S)-pyrrolidin-2-yl-oxazole E-5 (X.dbd.CH.sub.2, n=1) is
prepared as described below.
Step 1: 2-chlorocarbonyl-pyrrolidine-1-carboxylic Acid Benzyl Ester
(E-2)
[0111] A mixture of Z-L-Pro-OH E-1 (10.0 g, 40.1 mmol, 1 equiv.)
and thionyl chloride (30 mL, 401.2 mmol, 10 equiv.) in DCM (200 mL)
is heated to reflux under nitrogen for 20 minutes and concentrated
in vacuo. The residual oil is coevaporated with toluene 3.times.,
and concentrated to give Z-L-Pro-Cl as an opaque oil.
Step 2: 2-(trimethyl-silanyl)-2H-[1,2,3]triazole (E-3)
[0112] To a solution of 1H-1,2,3-triazole (4.98 g, 72.10 mmol, 1
equiv.) in dry benzene (145 mL) at rt under nitrogen, is added TEA
(11.05 mL, 79.31 mmol, 1.1 equiv.) followed by the dropwise
addition of TMSCl (9.15 mL, 72.10 mmol, 1 equiv.). A white
precipitate develops. Reaction mixture is stirred at rt under
nitrogen for 1 hour. The resultant precipitate is removed by
filtration and washed thoroughly with dry benzene. The filtrate is
gently concentrated to avoid evaporating the highly volatile
product to give a quantitative yield of TMS-triazole with a trace
of benzene.
Step 3: 2-oxazol-2-yl-pyrrolidine-1-carboxylic Acid Benzyl Ester
(E-4)
[0113] To a solution of TMS-triazole (14.17 g, 100.3 mmol, 1
equiv.) in sulfolane (290 mL) at rt under nitrogen, is added
dropwise Z-L-Pro-Cl (26.85 g, 100.3 mmol, 1 equiv.) in sulfolane
(70 mL). The resulting mixture is stirred at rt for 30 minutes,
then the temperature is elevated to 140.degree. C. for 3 hours.
After the reaction mixture is cooled to rt, poured into excess
brine and extracted with diethyl ether. The diethyl ether extracts
are washed with brine, dried over anhydrous Na.sub.2SO.sub.4,
filtered and evaporated. The residue is purified by flash
chromatography (DCM:acetone=9:1) to give the title compound as a
light yellow oil.
[0114] 1H NMR (CDCl.sub.3): .delta. 7.79-7.70 (s, 1H), 7.67-7.47
(m, 5H), 7.37-7.23 (m, 1H), 5.42-5.19 (m, 2H), 3.92-3.69 (m, 2H),
2.49-2.14 (m, 5H). ES-MS: calcd. for C.sub.15H.sub.16N.sub.2O.sub.3
(272.30). found: 273.5 [M+H].
Step 4: 2-pyrrolidin-2-yl-oxazole (Hydrobromic Acid Salt) (E-5)
[0115] A solution of compound E-4 (6.33 g, 23.25 mmol, 1 equiv.) in
AcOH (116 ml) at rt is treated with HBr (5.7 M, 33% in AcOH, 204
ml, 1162 mmol, 50 equiv.), and the mixture is stirred at rt for 1
hour. Charging the reaction mixture with excess diethyl ether and
cooling to 0.degree. C. for 30 minutes gives a solid which is
collected by filtration and dried in vacuo to afford the title
compound as a brownish powder.
[0116] 1H NMR (DMSO-d.sub.6): .delta. 9.98 (bs, 1H), 8.47 (s, 1H),
7.53 (s, 1H), 5.14-5.05 (m, 1H), 3.54-3.46 (m, 2H), 2.62-2.53 (m,
1H), 2.43-2.33 (m, 1H), 2.28-2.15 (m, 2H). ES-MS: calcd. for
C.sub.7H.sub.10N.sub.2O*HBr (219.08). found: 139.4 [M+H] free
base.
General Procedure F
Synthesis of 5-alkyl-2(S)-pyrrolidine-2-yl-oxazole
##STR00009##
[0117] 5-methyl-2-(S)-pyrrolidin-2-yl-oxazole (F-5)
[0118] A solution of (S)--N-(trifluoroacetyl)prolyl chloride
(Aldrich) 1 equiv. in DCM is treated with 1-amino-propan-2-one (1.5
equiv.) in pyridine at rt for 5 hours. After usual work up, it
provides 1-trifluoroacetyl-pyrrolidine-2-carboxylic acid
(2-oxo-propyl)-amide.
[0119] 1H NMR (CDCl.sub.3): .delta.4.55-4.50 (m, 1H), 4.15 (s, 2H),
3.78-3.69 (m, 2H), 2.18 (s, 3H), 2.15-1.85 (m, 4H).
[0120] Treatment of this intermediate with POCl.sub.3 at 70.degree.
C. for 2 hours provides
2,2,2-trifluoro-1-{2(S)-(5-methyl-oxazol-2-yl)-pyrrolidin-1-yl}-ethanone.
[0121] Treatment of this material with 2 M methanolic ammonia for 5
hours affords title compound.
[0122] MS: m/z=153.4 (M+1).
General Procedure G
Synthesis of 2-(alkylated-thiaazole)-2-pyrrolidine
##STR00010##
[0124] 2-(S)-pyrrolidin-2-yl-(4,5-dimethyl-thiaazole) G-4
(X.dbd.CH.sub.2, n=1, R.sub.2.dbd.R.sub.3.dbd.CH.sub.3) is prepared
as follows:
[0125] To a solution of thioamide G-2 (0.11 g, 1 equiv.) in DME (5
mL) is added 3-bromo-2-butanone (0.16 mL, 3 equiv.) and KHCO.sub.3
(0.40 g, 8 equiv.) and stirred for 2 hours. The reaction mixture is
cooled to 0.degree. C., then pyridine (0.4 mL, 8.5 equiv.) and
trifluoroacetic anhydride (0.32 mL, 4 equiv.) is added and the
mixture is stirred at rt 16 hours. It is diluted with EtOAc and
washed with water, brine, dried (Na.sub.2SO.sub.4) and concentrated
under reduced pressure. The residue is purified by silica gel
chromatography using hexane-EtOAc (1:1) as solvent gradient to give
the desired intermediate.
[0126] 1H NMR (CDCl.sub.3): .delta. 5.18-4.86 (m, 1H), 3.68-3.43
(m, 2H), 2.35 & 2.30 (each s, 2.times.3H), 2.15-1.81 (m, 2H),
2.54-1.45 (m, 2H), 1.35, (s, 9H). ES-MS: calcd. for
C.sub.14H.sub.22N.sub.2O.sub.2S (282.14). found: 283.3 [M+1].
[0127] Treatment of above compound with 4 M HCl in dioxane provides
the title compound.
[0128] 2-(S)-pyrrolidin-2-yl-(5-phenyl-thiaazole) G-4
(X.dbd.CH.sub.2, n=1, R.sub.2.dbd.C.sub.6H.sub.5, R.sub.3.dbd.H) is
prepared reacting a solution of thioamide G-2 (0.13 g, 1 equiv.) in
DME (5 mL) with 2-bromo-acetophenone (0.34 g, 3 equiv.) and
KHCO.sub.3 (0.45 g, 8 equiv.) and further working up as disclosed
above.
[0129] 1H NMR (CDCl.sub.3): .delta.8.25-8.15 (m, 3H), 7.71-7.45 (m,
3H), 5.39-5.22 (dd, J=2.8 Hz, 1H), 3.79-3.65 (m, 2H), 2.59-2.41 (m,
1H), 2.38-2.22 (m, 1H), 2.19-2.12 (m, 2H), 1.551 (bs, 9H). ES-MS:
calcd. for C.sub.18H.sub.22N.sub.2O.sub.2S (330.45). found: 331.5
[M+1].
[0130] Treatment of above compound with 4 M HCl in dioxane provides
the title compound.
General Procedure H
Synthesis of 2-pyrrolidin-2-yl-benzimidazole
##STR00011##
[0132] N-Cbz-L-Proline amide [X.dbd.CH.sub.2, n=1] (4 g, 16.1 mmol)
and o-phenylenediamine (1.7 g, 15.5 mmol) are heated to 165.degree.
C. in a nitrogen atmosphere for 2.5 hours then the temperature is
increased to 220.degree. C. for an additional 40 minutes. The
reaction mixture is cooled to rt and dissolved in DCM and washed
with saturated NaHCO.sub.3, water and brine then dried over
MgSO.sub.4 and evaporated to dryness. The residue is recrystallized
from 2:1/PrOH/water then diethyl ether/hexanes to provide the
Cbz-protected benzimidazole 630 mg. The protected material is then
taken up in AcOH (3 mL) and 33% HBr in AcOH (6 mL) is added. After
40 minutes, diethyl ether is added and the solution cooled to
0.degree. C. the precipitate is collected on a glass filter.
Recrystallization from MeOH/diethyl ether provides the title
compound as HBr salt.
[0133] 1H NMR (CDCl.sub.3): .delta. 7.63 (dd, J=3.3, 6.3, 2H), 7.43
(dd, J=3.0, 6.3, 2H), 5.13 (dd, J=8.0, 8.0, 1H), 4.57-3.37 (m, 2H),
2.64-2.58 (m, 1H), 2.43-2.04 (m, 3H). ES-MS: calcd. for
C.sub.11H.sub.13N.sub.3 (187.1). found: 188.1 [M+H].
General Procedure I
##STR00012##
[0134] Step 1
[0135] To an aminoacid methyl ester hydrochloride I-1 (P=methyl,
R.sub.1.dbd.H, 88 mmol) in saturated aqueous NaHCO.sub.3 (25 mL) is
added with vigorous stirring a solution of 2-nitrobenzene-sulfonyl
chloride (77 mmol) in THF (50 mL). Additional NaHCO.sub.3 is added
over 2 hours to maintain a basic pH. The reaction mixture is then
extracted with DCM (500 mL) and the organic layer washed with
water, dried (Na.sub.2SO.sub.4), concentrated and the crude product
recrystallized from 2:1 water/isopropanol then dried over
P.sub.2O.sub.5 in vacuo to provide N-2-nitrophenylsulfonyl
aminoacid methyl ester I-2 as colorless crystals. To a 0.degree. C.
solution of N-2-nitrophenylsulfonyl aminoacid methyl ester I-2 (28
mmol), an alcohol R.sub.10H (25 mmol) and triphenylphosphine (28
mmol) in THF (10 mL) is slowly added DIAD (28 mmol) over 5 minutes.
The reaction is allowed to warm to rt, then stirred additional 24
hours. The solvent is removed and the crude product purified on
silica gel (Merck 60; hexanes/DCM/THF 12:6:1) to afford
N-2-nitrophenylsulfonyl-N-alkyl-aminoacid methyl ester I-3.
Step 3
[0136] To a stirred suspension of
N-2-nitrophenylsulfonyl-N-alkyl-aminoacid methyl ester I-3
(R.sub.2=2-cyclopentylethyl, 11 mmol) and polymer bound
1,3,4,6,7,8-hexahydro-2H-pyrimidino[1,2-a]pyrimidine (12 mmol) in
DMF (40 mL) is added thiophenol (22 mmol). After 1 hour, the
reaction mixture is diluted with ether (300 mL), filtered and the
filtrate washed with brine (5.times.50 mL) and saturated
NaHCO.sub.3 (50 mL). The combined basic aqueous washes are then
back extracted with DCM (2.times.50 mL) and the DCM layers
combined. The ether phase is then extracted with 0.5 M HCl
(5.times.25 mL), the aqueous extract is made basic with solid
NaHCO.sub.3, then saturated with NaCl and extracted with
(5.times.50 mL) DCM. All DCM layers are combined, dried
(Na.sub.2SO.sub.4), acidified with 4 N HCl in dioxane and
evaporated to provide the secondary amine hydrochloride salt I-4.
In a separate flask, phosgene (20% in toluene; 0.11 mol) is added
to a vigorously stirred 0.degree. C. suspension of NaHCO.sub.3 (1
mole) in water (125 mL) and ether (200 mL). To this is added
dropwise over 30 min. the secondary amine HCl salt in water (125
mL) and an additional aliquot of phosgene (0.11 mol). The reaction
mixture is allowed to warm to rt then stirred additional 15
minutes. The phases are separated and the organic phase washed with
1 M HCl (2.times.75 mL), brine (75 mL), dried (MgSO.sub.4), and
evaporated to provide N-alkyl-aminoacid methyl ester carbamoyl
chloride I-5 (2 steps).
Step 4
[0137] An aliquot of N-alkyl-aminoacid methyl ester carbamoyl
chloride I-5 (4 mmol) dissolved in DCM (4 mL) is added to a
0.degree. C. solution of amine A-5 (5.3 mmol) in pyridine (4 mL).
After 30 minutes, the reaction mixture is diluted with ether (100
mL), washed with 10% KHSO.sub.4 (2.times.25 mL), brine (25 mL),
dried (NaSO.sub.4) and evaporated to provide the desired urea
I-6.
[0138] To N-[(2-carboxypyrrolidin-1-carbonyl)amino]aminoacid methyl
ester I-6 (200 .mu.mol) in dioxane (2 mL). The solution is diluted
with 50% aqueous hydroxylamine (1 mL), and the reaction stirred for
24-36 hours. The crude reaction mixture is then purified by
preparative reverse-phase (C18) HPLC to afford
N-hydroxy-2-[(2-amidopyrrolidin)amino]acetamide I-7.
General Procedure J
Synthesis of
1-[2-S-(oxazol-2-yl)-pyrrolidin-1-yl]-2-R/S-mercaptomethyl-alkan-1-one
##STR00013##
[0139] Step 1:
2-butyl-1-(2-S-oxazol-2-yl-pyrrolidin-1-yl)-propenone
[0140] To a solution of 2-butyl acrylic acid D-2 (1.2 mmol) in DMF
(10 mL) is added bicyclic pyrrolidine F-5 (1.2 mmol), HATU (1.2
mmol) and DIEA (2.5 mmol). The mixture is stirred overnight, then
concentrated and purified on silica gel (EtOAc/hexane 1:4) to
afford of the desired amide J-1.
Step 2: Thiolacetic Acid
S-{2-R/S-[2-S-(oxazol-2-yl)-pyrrolidine-1-carbonyl]-hexyl}ester
[0141] A solution of J-1 (3 mmol) in thiolacetic acid (15 mL) is
heated at 90.degree. C. for 3 hours then cooled to rt. It is
diluted with EtOAc and washed with saline, cold aq. NaHCO.sub.3,
dried over Na.sub.2SO.sub.4 and concentrated under reduced
pressure. The residue is purified on silica gel column using
solvent gradient consisting of 5-20% acetone in DCM to afford
J-2.
Step 3:
1-[2-S-(5-tert-butyl-oxazole-2-yl)-pyrrolidin-1-yl]-2-R/S-mercapto-
methyl-hexan-1-one
[0142] J-2 (1 mmol) is dissolved in MeOH (5 mL) under Argon with
stirring. Degassed 2 M NaOH solution (6 mmol) is added, and the
mixture is stirred for 3 hours. The reaction mixture is acidified
with IR-120(H+) resin until pH 2. The resin is removed by
filtration and filtrate is concentrated under reduced pressure to
give the title J-3 as a clear oil.
General Procedure K
Synthesis of
2-S-hydroxy-3-R-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-alkanoic
Acid
##STR00014##
[0143] Step 1:
5-R-{1-[2-S-(5-tert-butyl-oxazol-2-yl)-pyrrolidine-1-carbonyl]-pentyl}-2,-
2-dimethyl-[1,3]dioxolan-4-one
[0144] To a DMF solution (7 mL) of bicyclic amine salt (0.852 g,
2.4 mmol, 1.1 equiv) Hunig's base (2.1 mL, 12 mmol, 5.5 equiv.) is
added and the mixture cooled to 0.degree. C. This is followed by
the addition of the acetonide (500 mg, 2.17 mmol, 1.0 equiv.), and
HATU (0.913 g, 2.4 mmol, 1.1 equiv.) at 0.degree. C. The resulting
mixture is stirred at rt for 16 hours. The mixture is partitioned
between excess EtOAc and 10% citric acid. The organic layer is
washed with brine and saturated NaHCO.sub.3, dried over anhydrous
Na.sub.2SO.sub.4, filtered and concentrated to give K-1. The
residue is carried forward as is to the final step.
[0145] ES-MS: calcd. for C.sub.22H.sub.34N.sub.2O.sub.5 (406.25).
found: 407.5 [M+H].
Step 2:
3-R-[2-S-(5-tert-butyl-oxazol-2-yl)-pyrrolidine-1-carbonyl]-2-S-hy-
droxy-heptanoic Acid
[0146] The acetonide K-1 is dissolved in 10% (95:5
TFA:H.sub.2O)/DCE (10 mL) and the reaction stirred at rt for 7
hours. The final product is purified by preparative HPLC which upon
lyophylization yielded a final compound K-2 as a colorless
powder.
[0147] The following examples are to illustrate the invention but
should not be interpreted as a limitation thereon.
EXAMPLE 1
2(S)-hydroxy-3(R)-[2(R/S)-pyridin-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00015##
[0149] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 2-pyrrolidin-2-yl-pyridine A-5 according to
General Procedure B.
[0150] 1H NMR (DMSO): .delta. 8.6-8.5 (m, 1H), 7.97-7.92 (t, J=7.4
& 7.1 Hz, 1H), 7.5-7.3 (m, 2H), 5.14-4.94 (m, 1H), 3.96-3.44
(m, 3H), 3.1-2.9 (m, 1H), 2.3-1.9 (m, 6H), 1.3-1.1 (m, 4H),
0.87-0.9 (m, 3H). ES-MS: calcd. for C.sub.17H.sub.25N.sub.3O.sub.4
(335.40). found: 336.5 [M+1].
EXAMPLE 2
2(S)-hydroxy-3(R)-[3(R/S)-pyridin-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00016##
[0152] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 2-pyrrolidin-3-yl-pyridine A-5 according to
General Procedure B.
[0153] 1H NMR (DMSO): .delta. 8.85-8.8 (m, 1H), 8.21-8.14 (m, 2H),
7.8-7.6 (m, 4H), 4.4-3.9 (m, 4H), 3.8-3.6 (m, 2H), 3.09-3.06 (d,
J=9.34 Hz, 1H), 2.5-2.2 (m, 2H), 1.63 (bs, 2H), 1.38 (bs, 4H),
1.06-0.96 (m, 3H). ES-MS: calcd. for C.sub.17H.sub.25N.sub.3O.sub.4
(335.40). found: 336.5 [M+1].
EXAMPLE 3
2(S)-hydroxy-3(R)-[2-pyridin-3-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide (Isomer 1)
##STR00017##
[0155] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 3-pyrrolidin-2-yl-pyridine A-5 according to
General Procedure B.
[0156] 1H NMR (DMSO): .delta. 8.61 (s, 2H), 7.99-7.97 (d, J=7.97
Hz, 1H), 7.67-7.63 (m, 1H), 5.11-5.07 (m, 1H), 4.02-3.57 (m, 3H),
2.99-2.93 (m, 1H), 2.49-2.24 (m, 2H), 1.96-1.76 (m, 4H), 1.37-1.0
(m, 4H), -0.98-0.78 (m, 3H). ES-MS: calcd. for
C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5 [M+1].
EXAMPLE 4
2(S)-hydroxy-3(R)-[2-pyridin-3-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide (Isomer 2)
##STR00018##
[0158] The title compound is the other isomer isolated from the
reaction as described in Example 3.
[0159] 1H NMR (DMSO): .delta. 8.86-8.76 (m, 2H), 8.32-8.3 (d,
J=7.97 Hz, 1H), 7.9-7.88 (m, 1H), 5.4-5.32 (m, 1H), 4.3-3.74 (m,
3H), 3.22-3.16 (t, J=9.34 Hz, 1H), 2.52-2.46 (m, 2H), 2.26-1.98 (m,
3H), 1.63-1.38 (m, 5H), 1.07-1.01 (m, 3H). ES-MS: calcd. for
C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5 [M+1].
EXAMPLE 5
2(S)-hydroxy-3(R)-[2-pyridin-4-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide (Isomer 1)
##STR00019##
[0161] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 4-pyrrolidin-2-yl-pyridine A-5 according to
General Procedure B.
[0162] 1H NMR (DMSO): .delta. 8.89-8.87 (d, J=4.7 Hz, 2H),
7.77-7.75 (d, J=5.8 Hz, 2H), 5.3-5.25 (m, 1H), 4.23-3.8 (m, 3H),
3.33-3.15 (m, 1H), 2.52-2.43 (m, 2H), 2.13-1.86 (m, 3H), 1.56-1.26
(m, 5H), 1.07-1.05 (m, 3H). ES-MS: calcd. for
C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5 [M+1].
EXAMPLE 6
2(S)-hydroxy-3(R)-[2-pyridin-4-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide (Isomer 2)
##STR00020##
[0164] The title compound is the other isomer isolated from the
reaction as described in Example 5.
[0165] 1H NMR (DMSO): .delta. 8.89-8.85 (t, J=6.32 & 6.6 Hz,
2H), 7.92-7.68 (m, 2H), 5.39-5.36 (dd, 1H), 4.25-3.8 (m, 3H),
3.25-3.2 (t, J=7.96 & 8.8 Hz, 1H), 2.56-2.47 (m, 2H), 2.12-1.62
(m, 3H), 1.46-1.38 (m, 5H), 1.07-1.01 (m, 3H). ES-MS: calcd. for
C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5 [M+1].
EXAMPLE 7
2(S)-hydroxy-3(R)-[3(R/S)-pyridin-4-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00021##
[0167] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 4-pyrrolidin-3-yl-pyridine A-5 according to
General Procedure B.
[0168] 1H NMR (DMSO): .delta. 8.94 (dd, 2H), 8.06-8.02 (t, J=4.12
& 6.32 Hz, 2H), 4.2-3.53 (m, 6H), 3.1-3.02 (dd, 1H), 2.7-2.1
(m, 2H), 1.66-1.28 (m, 6H), 1.05-0.96 (m, 3H). ES-MS: calcd. for
C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5 [M+1].
EXAMPLE 8
2(S)-hydroxy-3(R)-[2(R/S)-phenyl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00022##
[0170] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid A-5 and
commercially available 2-phenyl-pyrrolidine B-5 according to
General Procedure B.
[0171] 1H NMR (DMSO): .delta. 7.6-7.3 (m, 5H), 5.6-5.57 (d, J=7.14
Hz, 1H), 5.3-5.24 (m, 1H), 4.21-3.72 (m, 4H), 3.2-3.14 (t, J=7.96
& 10.44 Hz, 1H), 2.4-2.33 (m, 1H), 2.3-1.85 (m, 4H), 1.6-1.36
(m, 5H), 1.04-0.97 (m, 3H). ES-MS: calcd. for
C.sub.18H.sub.26N.sub.2O.sub.4 (334.42). found: 335.5 [M+1].
EXAMPLE 9
2(S)-hydroxy-3(R)-[3(R/S)-phenyl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00023##
[0173] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 3-phenyl-pyrrolidine A-5 according to
General Procedure B.
[0174] 1H NMR (DMSO): .delta. 7.56-7.4 (m, 5H), 4.4-3.4 (m, =6H),
3.09-3.08 (d, J=3.85 Hz, 1H), 2.49-1.97 (m, 3H), 1.64-1.34 (m, 5H),
1.05-1.02 (m, 3H). ES-MS: calcd. for C.sub.18H.sub.26N.sub.2O.sub.4
(334.42). found: 335.5 [M+1].
EXAMPLE 10
2(S)-hydroxy-3(R)-[3(R/S)-pyridin-3-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00024##
[0176] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
commercially available 3-pyrrolidin-3-yl-pyridine A-5 according to
General Procedure B.
[0177] 1H NMR (DMSO): .delta. 8.89-8.76 (m, 2H), 8.26-8.2 (m, 1H),
7.79-7.74 (t, J=4.7 & 7.7 Hz, 1H), 4.1-3.5 (m, 6H), 3.09-3.07
(d, J=7.42 Hz, 1H), 2.54-1.37 (m, 8H), 1.05-0.97 (m, 3H). ES-MS:
calcd. for C.sub.17H.sub.25N.sub.3O.sub.4 (335.40). found: 336.5
[M+1].
EXAMPLE 11
2(S)-hydroxy-3(R)-[2(S)-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00025##
[0179] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
2-(S)-pyrrolidin-2-yl-oxazole A-5 (synthesis is described in
Procedure E) according to General Procedure B.
[0180] 1H NMR (CDCl.sub.3): .delta. 7.8 (s, 1H), 7.3 (s, 1H),
5.45-5.43 (t, J=3.9 &3.8 Hz, 1H), 4.45 (d, J=2.5 Hz, 1H),
4.1-3.96 (m, 2H), 3.54-3.51 (t, J=5.5 & 2.2 Hz, 1H), 2.5-2.3
(m, 4H), 1.95-1.91 (m, 2H), 1.57-1.53 (m, 4H), 1.12-1.08 (t, J=6.04
& 7.14 Hz, 3H). ES-MS: calcd. for
C.sub.15H.sub.23N.sub.3O.sub.5 (325.36). found: 326.4 [M+1].
EXAMPLE 12
2(S)-hydroxy-3(R)-[2(S)-(5-methyl-oxazol-2-yl-pyrrolidine-1-carbonyl)-hept-
anoic Acid Hydroxamide
##STR00026##
[0182] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
5-methyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1.dbd.H,
R.sub.2.dbd.CH.sub.3; preparation is described in Procedure F)
according to General Procedure B.
[0183] 1H NMR (CDCl.sub.3): .delta. 7.46 (s, 1H), 5.36-5.33 (m,
1H), 4.45-4.42 (m, 1H), 4.15-3.91 (m, 2H), 3.58-3.40 (m, 1H), 2.46
(s, 3H), 2.40-2.21 (m, 4H), 1.93-1.83 (m, 2H), 1.51-1.49 (m, 4H),
1.08 (t, J=6.9 Hz, 3H). ES-MS: calcd. for
C.sub.16H.sub.25N.sub.3O.sub.5 (339.39). found: 340.6 [M+1].
EXAMPLE 13
2(R)-fluoro-3-(R)-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00027##
[0185] The title compound is prepared according to General
Procedure C.sub.2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic
acid B-5 is coupled with 5-methyl-2-(S)-pyrrolidine-2-yl-oxazole
A-5 (prepared as described in Procedure E) to give B-6 which is
then treated further as described in Procedure C to afford the
title compound as one of the isomer.
[0186] 1H NMR (DMSO): .delta. 8.16 (s, 1H), 7.3 (s, 1H), 5.22-5.18
(dd, 1H), 5.07-4.9 (dd, J=7.97 & 8.24 Hz, 1H), 3.9-3.65 (m,
2H), 3.4-3.35 (m, 1H), 2.37-2.07 (m, 4H), 1.78-1.76 (d, J=6.04 Hz,
2H), 1.42 (bs, 4H), 1.03 (bs, 3H). ES-MS: calcd. for
C.sub.15H.sub.22FN.sub.3O.sub.4 (327.56). found: 350.5 [M+23].
EXAMPLE 14
2(S)-fluoro-3-(R)-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid Hydroxamide
##STR00028##
[0188] The title compound is the other isomer isolated from the
reaction as described in Example 13.
[0189] 1H NMR (DMSO): .delta. 8.16 (s, 1H), 7.3 (s, 1H), 5.33-5.29
(dd, 1H), 4.97-4.78 (dd, J=9.89 & 10.44 Hz, 1H), 3.94-3.7 (m,
2H), 3.38-3.32 (m, 1H), 2.42-2.08 (m, 6H), 1.54-1.38 (m, 4H),
1.02-0.99 (m, 3H). ES-MS: calcd. for
C.sub.15H.sub.22FN.sub.3O.sub.4 (327.56). found: 328.5 [M+1].
EXAMPLE 15
N-hydroxy-N-[2-(2-oxazol-2-yl-pyrrolidine-1-carbonyl)-hexyl]-formamide
##STR00029##
[0191] The title compound is prepared from
2-[(benzyloxy-formyl-amino)-methyl]-hexanoic acid D-7 (R=n-butyl)
and 2-(S)-pyrrolidine-2-yl-oxazole E-5 (prepared as described in
Procedure E) according to General Procedure D.
[0192] 1H NMR (DMSO-d.sub.6): .delta. 9.89 (s, 1H), 8.16 (s, 1H),
7.97 (s, 1H), 7.29 (s, 1H), 5.28-5.21 (m, 1H), 3.88-3.45 (m, 4H),
3.31-3.02 (m, 1H), 2.41-2.27 (m, 1H), 2.20-2.03 (m, 4H), 1.57-1.41
(m, 5H), 1.02 (bs, 3H). ES-MS: calcd. for
C.sub.15H.sub.23N.sub.3O.sub.4 (309.36). found: 310.6 [M+H], 332.5
M+Na].
EXAMPLE 16
S-hydroxy-N-[3(R)-(2-R/S-pyridin-2-yl-pyrrolidine-1-carbonyl)-hexyl]-forma-
mide (Isomer 1)
##STR00030##
[0194] The title compound is prepared from
2-[(benzyloxy-formyl-amino)-methyl]-hexanoic acid D-7 (R=n-butyl)
and commercially available 2-pyrrolidin-2-yl-pyridine A-5 according
to General Procedure D.
[0195] 1H NMR (D.sub.2O): .delta. 8.55 (d, J=4.9, 1H), 7.99-7.96
(m, 1H), 7.78 (s, 1H), 7.37-7.28 (m, 2H), 5.02 (dd, J=3.6, 3.6,
1H), 3.80-3.63 (m, 1H), 3.61-3.52 (m, 1H), 3.32-3.25 (m, 1H),
2.37-2.09 (m, 1H), 1.98-1.85 (m, 2H), 1.48-1.35 (m, 1H), 1.36-1.19
(m, 4H), 0.85-0.81 (t, J=11.8, 3H). ES-MS: calcd. for
C.sub.18H.sub.27N.sub.5O.sub.4 (319.2). found: 320.5 [M+H].
EXAMPLE 17
N-hydroxy-N-[3(R)-(2-R/S-pyridin-2-yl-pyrrolidine-1-carbonyl)-hexyl]-forma-
mide (Isomer 2)
##STR00031##
[0197] The title compound is the other isomer isolated from the
reaction as described in Example 16.
[0198] 1H NMR (D.sub.2O): .delta. 8.59 (d, J=5.8, 1H), 8.41 (dd,
J=7.7, 7.7 1H), 7.82-7.75 (m, 1H), 7.59 (s, 1H), 7.56-7.49 (m, 1H),
5.22 (dd, J=4.7, 8.8, 1H), 4.05-3.89 (m, 1H), 3.87-3.80 (m, 1H),
3.72-3.62 (m, 1H), 3.54 (dd, J=3.9, 14.8, 1H), 3.41-3.28 (m, 1H),
2.53-2.44 (m, 2H), 2.12-1.94 (m, 4H), 1.80-1.45 (m, 2H), 1.40-1.23
(m, 4H), 0.89 (dd, J=5.5, 6.9, 3H). ES-MS: calcd. for
C.sub.18H.sub.27N.sub.5O.sub.4 (319.2). found: 320.5 [M+H].
EXAMPLE 18
2(S)-hydroxy-3(R)-[2-S-(4,5-dimethyl-thiaazol-2-yl-pyrrolidine-1-carbonyl]-
-heptanoic Acid Hydroxamide
##STR00032##
[0200] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
2-(S)-pyrrolidin-2-yl-(4,5-dimethyl-thiaazole) G-4
(R.sub.2.dbd.R.sub.3.dbd.CH.sub.3, synthesis is described in
Procedure G) according to General Procedure B.
[0201] 1H NMR (CDCl.sub.3): .delta. 5.41-5.37 (dd, J=2.5 Hz, 1H),
4.04-4.01 (m, 1H), 3.97 (d, J=9 Hz, 1H), 3.85-3.78 (m, 1H),
3.14-3.08 (m, 1H), 2.44 & 2.39 (each s, 2.times.3H), 2.32-2.11
(m, 4H), 1.61-1.39 (m, 6H), 1.03 (t, J=6.0 Hz, 3H). ES-MS: calcd.
for C.sub.17H.sub.27N.sub.3O.sub.4S (369.17). found: 370.3
[M+1].
EXAMPLE 19
Synthesis of
2(S)-hydroxy-3(R)-[2-S-(4-phenyl-thiaazol-2-yl-pyrrolidine-1-carbonyl]-he-
ptanoic Acid Hydroxamide
##STR00033##
[0203] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
2-(S)-pyrrolidin-2-yl-(4-phenyl-thiaazole) G-4
(R.sub.2.dbd.C.sub.6H.sub.5, R.sub.3.dbd.H, synthesis is described
in Procedure G) according to General Procedure B.
[0204] 1H NMR (CDCl.sub.3): .delta.8.18-8.10 (m, 3H), 7.64-7.49 (m,
3H), 5.59-5.58 (dd, J=2.8 Hz, 1H), 4.10-4.01 (m, 1H), 3.95 (d, J=7
Hz, 1H), 3.91-3.89 (m, 1H), 3.18-3.14 (m, 1H), 2.41-2.19 (m, 4H),
1.64-1.37 (m, 6H), 1.01 (t, J=6.0 Hz, 3H). ES-MS: calcd. for
C.sub.21H.sub.27N.sub.3O.sub.4S (417.52). found: 418.5 [M+1].
EXAMPLE 20
2(S)-hydroxy-3(R)-[2-S-(4-methyl-thiaazol-2-yl-pyrrolidine-1-carbonyl]-hep-
tanoic Acid Hydroxamide
##STR00034##
[0206] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
2-(S)-pyrrolidin-2-yl-(4-methyl-thiaazole) G-4
(R.sub.2.dbd.CH.sub.3, R.sub.3.dbd.H, preparation is described in
Procedure G) according to General Procedure B.
[0207] 1H NMR (CDCl.sub.3): .delta. 7.49-7.6 (dd, 1H), 7.05 (bs,
1H), 5.71-5.69 (d, J=6.04 Hz, 1H), 4.45 (bs, 1H), 4.08-3.94 (d,
2H), 3.51 (bs, 1H), 2.67-2.63 (t, 3H), 2.39 bs, 4H), 1.96 (bs, 2H),
1.57 (bs, 4H), 1.12-1.10 (d, 3H). ES-MS: calcd. for
C.sub.16H.sub.25N.sub.3O.sub.4S (355.47). found: 356.3 [M+1].
EXAMPLE 21
2(S)-hydroxy-3(R)-[2-S-(benzimidazol-2-yl-pyrrolidine-1-carbonyl]-heptanoi-
c Acid Hydroxamide
##STR00035##
[0209] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid B-5 and
2-(S)-pyrrolidin-2-yl-(benzimidazole) G-4 (synthesis is described
in Procedure G) according to General Procedure B.
[0210] 1H NMR (DMSO): .delta. 7.78-7.66 (m, 2H), 7.51-7.37 (m, 2H),
5.41-5.37 (m, 1H), 5.25-5.21 (t, J=6.32 Hz, 1H), 3.8-3.52 (m, 2H),
3.02-2.9 (m, 1H), 2.38-2.06 (m, 4H), 1.47-1.03 (m, 6H), 0.87-0.70
(m, 3H). ES-MS: calcd. for C.sub.19H.sub.26N.sub.4O.sub.4 (374.44).
found: 375.3 [M+1].
EXAMPLE 22
N-hydroxy-2-[N-(2-cyclopentylethyl)-N-[2-S-(4-phenyl)-thiaazol-2-yl-pyrrol-
idine]-acetamide
[0211] The title compound is prepared from I-5 and
2-(S)-pyrrolidin-2-yl-(5-phenyl-thiaazole) G-4
(R.sub.2.dbd.C.sub.6H.sub.5, R.sub.3.dbd.H) according to General
Procedure I-4.
[0212] 1H NMR (D.sub.6 DMSO): .delta. 7.93-7.91 (m, 3H), 8.41 (dd,
J=7.1, 7.1 2H), 7.35-7.30 (m, 1H), 5.31 (dd, J=6.9, 6.9 1H), 3.89
(d, J=16.2 1H), 3.62 (d, J=16.2 1H), 3.58-3.40 (m, 2H), 3.30-3.15
(m, 2H), 2.42-2.39 (m, 1H), 1.96-1.83 (m, 3H), 1.69-1.60 (m, 3H),
1.54-1.43 (m, 6H), 1.07-0.96 (m, 2H). ES-MS: calcd. for
C.sub.23H.sub.30N.sub.4O.sub.3S (442.2). found: 465.3 [M+Na].
EXAMPLE 23
N-hydroxy-2-[N-(2-cyclopentylethyl)-N-[2-(2-oxazol-2-yl-pyrrolidine-1-carb-
onyl)-hexyl]-acetamide
[0213] The title compound is prepared from I-5 and
5-methyl-2-(S)-pyrrolidine-2-yl-oxazole E-5 (prepared as described
in Procedure E) according to General Procedure I-4.
[0214] 1H NMR (D.sub.6 DMSO): .delta. 7.97 (s, 1H), 7.09 (s, 1H),
5.07 (dd, J=7.1, 7.1 1H), 3.81 (d, J=16.2 1H), 3.47-3.39 (m, 2H),
3.25-2.99 (m, 2H), 2.26-2.22 (m, 1H), 1.98-1.81 (m, 3H), 1.69-1.60
(m, 3H), 1.78-1.42 (m, 7H), 1.04-1.02 (m, 2H). ES-MS: calcd. for
C.sub.17H.sub.26N.sub.4O.sub.4 (350.2). found: 373.4 [M+Na].
EXAMPLE 24
2(S)-hydroxy-3(R)-[2(S)-(5-tert-butyl-oxazol-2-yl-pyrrolidine-1-carbonyl)--
heptanoic Acid Hydroxamide
##STR00036##
[0216] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
5-tert-butyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1.dbd.H,
R.sub.2=tert-butyl) according to General Procedure B.
[0217] 1H NMR (DMSO): .delta. 6.87-6.85 (d, J=5.22 Hz, 1H),
5.25-5.21 (dd, J=3.02 & 2.74 Hz, 1H), 4.04-3.96 (m, 2H),
3.86-3.79 (m, 1H), 3.13-3.07 (t, J=9.06 & 9.91 Hz), 2.35-2.28
(m, 2H), 2.26-2.05 (m, 2H), 1.59-1.37 (m, 6H), 1.013-0.997 (t,
J=4.8 & 6.32 Hz, 3H). ES-MS: calcd. for
C.sub.19H.sub.31N.sub.3O.sub.5 (381.47). found: 382.4 [M+H].
[0218] 5-tert-butyl-2-(S)-pyrrolidine-2-yl-oxazole is prepared as
described:
1-amino-pinacolone
[0219] To a solution of 1-bromo-pinacolone (5.4 g, 1 equiv.) in DMF
(30 mL) is added sodium azide (4 g, 5 equiv.) at 0.degree. C.,
stirred at 0.degree. C. for 1 hour and then brought to rt for 1
hour. The reaction mixture is diluted with EtOAc (150 mL) and
washed with cold water, dried over sodium sulfate. This azido
compound is treated with 10% Pd--C in ethanol-conc HCl to give
corresponding the title compound.
5-tert-butyl-2-S-pyrrolidine-2-yl-oxazole
[0220] A solution of Z-L-Pro chloride (1 equiv.) in DCM is treated
with 1-amino-pinacolone (1.2 equiv.) in pyridine at rt for 5 hours.
After usual work up, the resulting amide intermediate is treated
with POCl.sub.3 at 70.degree. C. for 2 hours to provide the
Z-N-protected bicyclic compound. Treatment with HBr-AcOH affords
title compound.
[0221] 1H NMR (DMSO): .delta. 9.51 (bs, 1H), 5.04 (bs, 1H), 3.49
(bs, 2H), 2.54-2.21 (m, 4H), 1.45 (bs, 9H). ES-MS: calcd. for
C.sub.11H.sub.18N.sub.2O (194.14). found: 195.3 [M+H].
EXAMPLE 25
2(S)-hydroxy-3(R)-[2(S)-(5-phenyl-oxazol-2-yl-pyrrolidine-1-carbonyl)-hept-
anoic Acid Hydroxamide
##STR00037##
[0223] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
5-phenyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1.dbd.H,
R.sub.2=phenyl) according to General Procedure B.
[0224] 1H NMR (DMSO): .delta. 7.86-7.83 (m, 1H), 7.77 (bs, 1H),
7.76-7.62 (m, 3H), 7.56-7.52 (m, 1H), 5.6-5.58 (d, J=6.32 Hz, 1H),
5.32-5.29 (d, J=6.59 Hz, 1H), 4.07-3.92 (m, 2H), 3.14-3.11 (m, 1H),
2.42-2.18 (m, 4H), 1.56-1.37 (m, 6H), 0.882 (bs, 3H). ES-MS: calcd.
for C.sub.21H.sub.27N.sub.3O.sub.5 (401.46). found: 402.2
[M+H].
[0225] The amine is prepared following the same procedure as
described in the Example 24 from 2-bromoacetophenone
##STR00038##
EXAMPLE 26
2(S)-hydroxy-3(R)-[2(S)-(5-iso-butyl-oxazol-2-yl-pyrrolidine-1-carbonyl)-h-
eptanoic Acid Hydroxamide
##STR00039##
[0227] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
5-iso-butyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1.dbd.H,
R.sub.2=iso-butyl) according to General Procedure B.
[0228] 1H NMR (DMSO): .delta. 6.93 (bs, 1H), 5.23-5.19 (dd, J=3.57
& 3.02 Hz, 1H), 4.05-3.95 (m, 2H), 3.83-3.78 (dd, J=6.87 &
7.14 Hz, 1H), 3.13-3.07 (t, J=9.34 & 8.42 Hz, 1H), 2.7-2.57 (m,
2H), 2.47-1.97 (m, 5H), 1.58-1.35 (m, 6H), 1.13-0.99 (m, 9H).
ES-MS: calcd. for C.sub.19H.sub.31N.sub.3O.sub.5 (381.47). found:
382.3 [M+H].
[0229] Bromination of 4-methyl-2-butanone provided the desired
bromo compound (major) along with other positional isomer in low
yield. Amine is prepared from this bromide following the same
procedure as described in the Example 24.
EXAMPLE 27
2(S)-hydroxy-3(R)-[2(S)-(4,5-dimethyl-oxazol-2-yl-pyrrolidine-1-carbonyl)--
heptanoic Acid Hydroxamide
##STR00040##
[0231] The title compound is prepared from
2-(2,2-di-methyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
4,5-di-methyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1=Me,
R.sub.2=Me) according to General Procedure B.
[0232] 1H NMR (DMSO): .delta. 5.17-5.13 (dd, J=3.57 & 3.02 Hz,
1H), 4.06-3.92 (m, 2H), 3.86-3.82 (dd, J=7.42 & 4.12 Hz, 1H),
3.12-3.06 (t, J=8.79 & 8.06 Hz, 1H), 2.56-2.26 (m, 2H), 2.35
(bs, 3H), 2.18-2.05 (m, 2H), 2.14 (bs, 3H), 1.57-1.35 (m, 6H),
1.04-1.02 (d, J=6.043 Hz, 3H). ES-MS: calcd. for
C.sub.17H.sub.27N.sub.3O.sub.5 (353.41). found: 354.63 [M+H].
[0233] 4,5-di-methyl-2-(S)-pyrrolidin-2-yl-oxazole is prepared as
follows:
2-amino-butan-3-one
[0234] To a solution of di-tert-butyl imino carboxylate (28 g, 1
equiv.) in DMF is added Cs.sub.2CO.sub.3 (134 g, 3 equiv.) and
tetrabutylammonium iodide (154 g, 3 equiv.) under Argon atmosphere.
After 1 hour, 2-chloro-butan-3-one (40 mL, 3 equiv.) is added to
reaction mixture and the mixture stirred at rt for 72 hours,
diluted with EtOAc and filtered through Celite to remove the
inorganic material. The filtrate is washed with aq. NaHCO.sub.3,
water, 10% aq. citric acid, dried over Na.sub.2SO.sub.4 and
concentrated under reduced pressure. The residue is purified on
silica gel chromatography to give Boc protected compound. Treatment
of this intermediate with 4 M HCl for 16 hours and evaporation of
the solvent provides 2-amino-butan-3-one.
4,5-di-methyl-2-S-pyrrolidine-2-yl-oxazole
##STR00041##
[0236] A solution of Z-L-Pro chloride (1 equiv.) in DCM is treated
with 2-amino-butan-3-one (1.5 equiv.) in pyridine at rt for 5
hours. After usual work up, the resulting amide intermediate is
treated with POCl.sub.3 at 70.degree. C. for 2 hours providing the
Z-N-protected bicyclic intermediate. Treatment of this material
with HBr-AcOH affords title compound.
[0237] 1H NMR (DMSO): .delta. 4.99-4.94 (t, J=7.32 & 6.7 Hz,
1H), 3.49-3.42 (m, 2H), 2.56-2.1 (m, 10H). ES-MS: calcd. for
C.sub.9H.sub.14N.sub.2O (166.11). found: 167.3 [M+H].
EXAMPLE 28
2-S-hydroxy-3-R-(2-S-oxazol-2-yl-pyrrolidine-1-carbonyl)-heptanoic
Acid
##STR00042##
[0239] The title compound is prepared from
2-(2,2-dimethyl-4-oxo-1,3-dioxolan-5-yl)hexanoic acid and
5-tert-butyl-2-(S)-pyrrolidin-2-yl-oxazole F-5 (R.sup.1.dbd.H,
R.sub.2=tert-butyl) according to General Procedure K followed by
the treatment with 90% aq. TFA-DCE.
[0240] 1H NMR (DMSO): .delta. 6.48 (s, 1H), 5.21-5.18 (dd, J=3.65
& 3.3 Hz, 1H), 4.15-3.92 (m, 3H), 3.80-3.74 (m, 1H), 2.92-2.86
(m, 2H), 2.32-2.28 (m, 1H), 2.14-2.03 (m, 3H), 2.02-1.36 (m, 15H),
1.012-0.973 (t, J=6.6 & 5.22 Hz, 3H). ES-MS: calcd. for
C.sub.19H.sub.30N.sub.2O.sub.5 (366.46). found: 367.5 [M+H].
EXAMPLE 29
1-[2-S-(5-tert-butyl-oxazole-2-yl)-pyrrolidin-1-yl]-2-R/S-mercaptomethyl-h-
exan-1-one
##STR00043##
[0242] The title compound is prepared from 2-butyl acrylic acid
(D-2) and 5-tert-butyl-2-(S)-pyrrolidin-2-yl-oxazole F-5
(R.sup.1.dbd.H, R.sub.2=tert-butyl) according to General Procedure
J followed by the treatment with thiolacetic acid at 90.degree. C.
for 3 hours to give S-acetyl compound. The removal of acetyl group
with 2N-sodium hydroxide in MeOH provided the isomeric mixture of
title this compound.
[0243] 1H NMR (CDCl.sub.3): .delta. 6.62 (s, 0.6H), 6.55 (s, 0.4H),
4.09-4.04 (m, 0.6H), 3.72-3.74 (m, 2H), 3.62-3.53 (m, 0.4H),
2.95-2.73 (m, 2H), 2.61-2.40 (m, 0.6H), 2.38-2.30 (m, 0.4H)
2.27-2.00 (m, 6H), 1.71-1.64 (m, 4H), 1.56-1.44 (m, 1H, SH), 1.26
(bs, 9H), 0.89 (t, J=7 Hz, 3H), 0.74 (t, J=7 Hz, 3H). ES-MS: calcd.
for C.sub.18H.sub.30N.sub.2O.sub.2S (338.51). found: 339.5
[M+H].
[0244] Preferred compounds according to the invention are, e.g.,
the compounds of Examples 11-15.
[0245] The compounds of the invention, e.g., the compounds of
formula (I), in free form or in pharmaceutically acceptable salt
from or a prodrug thereof, exhibit valuable pharmacological
properties, e.g., as anti-infectious agents, e.g., as indicated in
in vitro and in vivo tests and are therefore indicated for
therapy.
A. Inhibition of Peptide Deformylase Activity
[0246] The PDF/FDH coupled assay (see Lazennec et al., Anal.
Biochem., Vol. 224, pp. 180-182 (1997)) is used. In this coupled
assay, the formate released by PDF from its substrate fMAS is
oxidized by the coupling enzyme FDH, reducing one molecule of
NAD.sup.+ to NADH, which causes an increase in absorption at 340
nM. All assays are carried out at rt in a buffer of 50 mM HEPES, pH
7.2, 10 mM NaCl, 0.2 mg/mL BSA, in half-area 96-well microtiter
plates (Corning). The reaction is initiated by adding a mixture of
0.5 U/mL FDH, 1 mM NAD.sup.+, and fMAS at the desired
concentration. To determine IC.sub.50 values, PDF is pre-incubated
for 10 minutes with varying concentrations of actinonin, and the
deformylation reaction is initiated by the addition of reaction
mixture containing 4 mM fMAS. The initial reaction velocity, y, is
measured as the initial rate of absorption increase at 340 nM using
a SpectraMax plate reader (Molecular Devices, Sunnyvale, Calif.).
The inhibitor concentration [In] at which 50% of the enzyme
activity is inhibited, IC.sub.50, is calculated using the following
formula:
y=y.sub.o/(1+[In]/IC.sub.50)
where y.sub.o is the reaction velocity in the absence of inhibitor.
Solving this equation for IC.sub.50 at the [In] when y=y.sub.o/2
yields IC.sub.50. The IC.sub.50 is calculated based on a nonlinear
least-square regression fit using a commercial software package
(Deltapoint, Inc., Chicago, Ill.).
[0247] Using this assay, the IC.sub.50 of various compounds are
determined. The IC.sub.50 for the various compounds is determined
against deformylase enzyme containing nickel and zinc as the metal
ion. The IC.sub.50 values of preferred compounds of formula (I)
determined for the zinc-containing deformylase ranged from about
0.585 .mu.M to about 0.004 .mu.M. The IC.sub.50 values of preferred
compounds of formula (I) determined for the nickel-containing
deformylase ranged from about 0.06 .mu.M to about 0.0001 .mu.M.
B. Assay for Testing Antimicrobial Activity
[0248] Minimum inhibitory concentrations (MICs) are determined
using the microdilution method in 96-well format plates. Compounds
are suspended in DMSO at 5 or 10 mg/mL and stored at 4.degree. C.
until used. They are diluted in Mueller-Hinton Broth (MHB) or
Trypticase Soy Broth (TSB) and used for MIC determination. The
range of concentrations tested is 64-0.0625 .mu.g/mL final
concentration using a two-fold dilution system.
[0249] The inoculum is prepared from cells grown on Trypticase Soy
Agar (TSA) and incubated overnight at 35.degree. C., 5-10 colonies
are used to inoculate MHB or TSB broths, and the culture is
incubated overnight at 35.degree. C. The overnight culture is
diluted 1:10, incubated for 1 hour at 35.degree. C., diluted to the
appropriate inoculum size and applied to the wells containing broth
and test compound. Inoculum sizes are 2.times.10.sup.4 CFU/mL.
[0250] Plates are incubated at 35.degree. C. for 48 hours and MIC
are recorded after 18 hours of incubation for bacteria. MIC is
defined as the lowest concentration of compound that does not
produce visible growth after incubation.
[0251] Minimum inhibitory concentration for various preferred
compounds of formula (I) ranged from about 0.125 .mu.g/mL to about
2.0 .mu.g/mL against H. influenza (four strains), from about 0.25
.mu.g/mL to about greater than 64 .mu.g/mL against S. aureus (four
strains), from about 2 .mu.g/mL to about 16 .mu.g/mL against S.
pneumonia (three strains), and from about 0.125 .mu.g/mL to about 2
.mu.g/mL against M. catarrhalis. The deformylase enzyme is obtained
from E. coli.
C. Demonstration of Selective Inhibition of PDF Compared to
MMP-7
[0252] As noted previously, inhibitors which are selective for PDF
over MMPs are desirable in order to avoid side effects.
[0253] In order to test the compounds of the invention for possible
inhibitory effects on MMPs, the following assay for MMP-7 is
used.
MMP-7 Assay
[0254] Matrilysin activity is assayed using a thio-peptide
(Pro-Leu-Gly-S-Leu-Leu-Gly) as substrate. Upon enzyme hydrolysis,
the thiolate is released as a product. The thiolate thus generated
reacts with DTNB (dithionitrobenzene), giving rise to a yellow
color which is monitored at 405 nM. The assay is carried out at rt;
the assay buffer contains 50 mM Tricine, pH 7.5, 0.2 M NaCl, 10 mM
CaCl.sub.2, and 0.05% Brij, in a half-area 96-well microtiter
plate. The reaction is initiated by adding a mixture of 200 TM DTNB
and 100 TM thiopeptide in buffer. To determine IC.sub.50 values,
MMP-7 is pre-incubated for 10 minutes with varying concentrations
of compounds, and the hydrolysis initiated by the addition of
reaction mixture containing thiopeptide and DTNB. The reaction rate
is recorded as the absorbance increase in OD.sub.405 over 30
minutes using a SpectraMax plate reader (Molecular Devices,
Sunnyvale, Calif.). The inhibitor concentration [In] at which 50%
of the enzyme activity is inhibited, IC.sub.50, is calculated using
the following formula:
y=y.sub.o/(1+[In]/IC.sub.50)
where y.sub.o is the reaction velocity in the absence of inhibitor.
Solving this equation for IC.sub.50 at the [In] when y=y.sub.o/2
yields IC.sub.50.
[0255] Using this assay, the IC.sub.50 of various compounds are
determined. The IC.sub.50 of various preferred compounds of formula
(I) against MMP-7 is about 100 .mu.M, whereas the IC.sub.50 of
these same compounds against zinc-containing PDF ranged from about
0.004 .mu.M to about 0.585 .mu.M, and against nickel-containing PDF
ranged from about 0.001 .mu.M to about 0.006 .mu.M. Accordingly, it
can be seen that the compounds provided by the invention have
superior selectivity for PDF as compared to their activity against
MMP-7. Similar selectivity of the compounds for PDF over MMP-1,
MMP-2, MMP-3, MMP-9, MMP-13, MT-MMP-1 and tissue necrosis factor
converting enzyme is observed. Similar selectivity is also observed
over other metalloproteinases such as angiotensin converting
enzyme.
D. Mouse Septicemia Model for Determining In Vivo Efficacy
[0256] CD1 female out-bred mice (Charles River Laboratories)
weighing 18-22 g each are injected intraperitoneally with 0.5 mL of
a suspension containing 5.times.10.sup.7 cfu of S. aureus (Smith
strain) in 7% hog gastric mucosa (mucin). The mice are treated
either s.c., i.v. or p.o., 1 hour and 5 hours after infection. Six
groups of six mice each are given different dosage levels
representing two-fold dilutions of each compound (range of 100-0.1
mg/kg). Vancomycin is used as the control antibiotic and is
administered s.c. Compounds are formulated in PBS and untreated
controls are dosed with vehicle alone.
[0257] Deaths in each group are monitored daily for 6 days and
cumulative mortality is used to determine PD.sub.50, which are
calculated using the method of Reed and Muench. The ED.sub.50
(s.c.) in mice against S. aureus for several preferred compound of
formula (I) ranged from about 3.45 mg/kg to greater than 10 mg/kg.
The PD.sub.50 (p.o.) in mice against S. aureus for these same
compounds of formula (I) ranged from 7.06 mg/kg to about 10.83
mg/kg.
E. Pharmacokinetics Study of PDF Inhibitors in Mice
[0258] The pharmacokinetics of PDF compounds are determined in CD1
female out-bred mice (Charles River Laboratories) weighing 20-25 g.
PDF compounds are formulated in 20% cyclodextrin (Aldrich) and
filtered through 0.22 .mu.M filter for sterilization. Either single
compound or mixtures of 4-6 compounds as a cassette are
administered i.v. and p.o. at 10 mL/kg. The dose ranged from 3-15
mg/kg for each compound. At 0.083, 0.25, 0.5, 1, 2, 4 and 7 hours
after dosing, serum samples are collected via cardiac puncture
under anesthesia. Groups of four mice are used for each time point.
The serum samples are stored at -80.degree. C. until analysis.
[0259] The serum protein is precipitated by addition of
acetonitrile. The samples after protein precipitation are analyzed
by LC/MS/MS method. Standard curve is obtained for each compound
and used for determination of compound concentration in serum. The
pharmacokinetics parameters including T.sub.max, C.sub.max,
t.sub.1/2 and AUC, are calculated according to standard method. The
oral bioavailability is calculated as the ratio of AUC of p.o.
administration versus the AUC administered i.v. The preferred
compounds of formula (I) exhibited oral bioavailability greater
than 35%.
[0260] The compounds of the present invention are, therefore,
useful to inhibit bacteria or for the treatment and/or prevention
of infectious disorders caused by a variety of bacterial or
prokaryotic organisms. Examples include, but are not limited to,
Gram positive and Gram negative aerobic and anaerobic bacteria
including Staphylococci, e.g., S. aureus and S. epidermidis;
Enterococci, e.g., E. faecalis and E. faecium; Streptococci, e.g.,
S. pneumoniae; Haemophilus, e.g., H. influenza; Moraxella, e.g., M.
catarrhalis; Bacteroides, e.g., Bacteroides fragilis, Clostridium,
e.g., Clostridium difficile, Niesseria, e.g., N. meningitidis and
N. gonorrhoae, Legionella and Escherichia, e.g., E. coli. Other
examples include Mycobacteria, e.g., M. tuberculosis; intracellular
microbes, e.g., Chlamydia and Rickettsiae; and Mycoplasma, e.g., M.
pneumoniae; Pseudomonas, e.g., P. aeruginosa; Helicobacter pylori;
and parasites, e.g., Plasmodium falciparum.
[0261] As used herein, an "infectious disorder" is any disorder
characterized by the presence of a microbial infection, such as the
presence of bacteria. Such infectious disorders include, for
example, central nervous system infections, external ear
infections, infections of the middle ear, such as acute otitis
media, infections of the cranial sinuses, eye infections,
infections of the oral cavity, such as infections of the teeth,
gums and mucosa, upper respiratory tract infections, lower
respiratory tract infections, genitourinary infections,
gastrointestinal infections, gynecological infections, septicemia,
bone and joint infections, skin and skin structure infections,
bacterial endocarditis, burns, antibacterial prophylaxis of
surgery, antibacterial prophylaxis in immunosuppressed patients,
such as patients receiving cancer chemotherapy, or organ transplant
patients and chronic diseases caused by infectious organisms, e.g.,
arteriosclerosis.
[0262] The compounds may be used to treat a subject to treat,
prevent, and/or reduce the severity of an infection. Subjects
include animals, plants, blood products, cultures and surfaces such
as those of medical or research equipment, such as glass, needles,
surgical equipment and tubing, and objects intended for temporary
or permanent implantation into an organism. Preferred animals
include mammals, e.g., mice, rats, cats, dogs, cows, sheep, pigs,
horses, swine, primates, such as rhesus monkeys, chimpanzees,
gorillas and most preferably humans. Treating a subject includes,
but is not limited to, preventing, reducing and/or eliminating the
clinical symptoms caused by an infection of a subject by a
microorganism; preventing, reducing and/or eliminating an infection
of a subject by a microorganism; or preventing, reducing and/or
eliminating contamination of a subject by a microorganism. The
microorganism involved is preferably a prokaryote, more preferably
a bacterium.
[0263] For the above uses the required dosage will of course vary
depending on the mode of administration, the particular condition
to be treated and the effect desired. The compositions may contain,
for example, from about 0.1% by weight to about 99% by weight,
e.g., from about 10-60% by weight, of the active material,
depending on the method of administration. Where the compositions
comprise dosage units, each unit will contain, for example, from
about 1-1000 mg, e.g., 1-500 mg of the active ingredient. The
dosage as employed for adult human treatment will range, for
example, from about 1-3000 mg/day, for instance, 1500 mg/day
depending on the route and frequency of administration. Such a
dosage corresponds to about 0.015-50 mg/kg/day. Suitably the dosage
is, for example, from about 5-20 mg/kg/day. Suitable unit dosage
forms for oral administration comprise ca. 0.25-1500 mg active
ingredient.
[0264] A "pharmaceutically acceptable carrier" means an excipient
that is useful in preparing a pharmaceutical composition that is
generally safe, non-toxic and neither biologically nor otherwise
undesirable, and includes an excipient that is acceptable for
veterinary use as well as human pharmaceutical use. A
"pharmaceutically acceptable carrier" as used in the specification
and claims includes both one and more than one such carriers.
[0265] The compounds may be administered by any conventional route,
e.g., locally or systemically e.g., orally, topically,
parenterally, subdermally or by inhalation and may be used for the
treatment of bacterial infection in a subject such as animals,
preferably, mammals, more preferably, humans.
[0266] The compositions may be administered by any conventional
route known in the art, e.g., subdermally, by inhalation, orally,
topically or parenterally, and may be used for the treatment of
bacterial infection in a subject, such as animals, preferably,
mammals, more preferably, humans.
[0267] The compounds of the invention may be formulated for
administration in any convenient way for use in human or veterinary
medicine, by analogy with other antibiotics. Such methods are known
in the art (see, e.g., Remington's Pharmaceutical Sciences, Easton,
Pa.: Mack Publishing Co.) and are not described in detail
herein.
[0268] The compositions may be in any form known in the art,
including but not limited to tablets, capsules, powders, granules,
lozenges, creams or liquid preparations, such as oral or sterile
parenteral solutions or suspensions. The compounds may also be
administered in liposome formulations. The compounds may also be
administered as prodrugs, where the prodrug administered undergoes
biotransformation in the treated mammal to a form which is
biologically active.
[0269] The topical formulations of the present invention may be
presented as, for instance, ointments, creams or lotions,
solutions, salves, emulsions, plasters, eye ointments and eye or
ear drops, impregnated dressings and aerosols and may contain
appropriate conventional additives, such as preservatives, solvents
to assist drug penetration and emollients in ointments and
creams.
[0270] The formulations may also contain compatible conventional
carriers, such as cream or ointment bases and ethanol or oleyl
alcohol for lotions. Such carriers may be present, for example,
from about 1% up to about 99% of the formulation. For example, they
may form up to about 80% of the formulation.
[0271] Tablets and capsules for oral administration may be in unit
dose presentation form, and may contain conventional excipients,
such as binding agents, for example, syrup, acacia, gelatin,
sorbitol, tragacanth or polyvinylpyrollidone; fillers, for example,
lactose, sugar, maize-starch, calcium phosphate, sorbitol or
glycine; tabletting lubricants, for example, magnesium stearate,
talc, polyethylene glycol or silica; disintegrants, for example,
potato starch; or acceptable wetting agents, such as sodium lauryl
sulphate. The tablets may be coated according to methods well-known
in standard pharmaceutical practice.
[0272] Oral liquid preparations may be in the form of, for example,
aqueous or oily suspensions, solutions, emulsions, syrups or
elixirs or may be presented as a dry product for reconstitution
with water or another suitable vehicle before use. Such liquid
preparations may contain conventional additives, such as suspending
agents, for example, sorbitol, methyl cellulose, glucose syrup,
gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, aluminium
stearate gel or hydrogenated edible fats; emulsifying agents, for
example, lecithin, sorbitan monooleate, or acacia; non-aqueous
vehicles (which may include edible oils), for example, almond oil,
oily esters, such as glycerine, propylene glycol, or ethyl alcohol;
preservatives, for example, methyl or propyl p-hydroxybenzoate or
sorbic acid, and, if desired, conventional flavoring or coloring
agents.
[0273] For parenteral administration, fluid unit dosage forms are
prepared utilizing the compound and a sterile vehicle, water being
preferred. The compound, depending on the vehicle and concentration
used, may be either suspended or dissolved in the vehicle or other
suitable solvent. In preparing solutions, the compound may be
dissolved in water for injection and filter sterilized before
filling into a suitable vial or ampoule and sealing.
Advantageously, agents such as a local anesthetic preservative and
buffering agents may be dissolved in the vehicle. To enhance the
stability, the composition may be frozen after filling into the
vial and the water removed under vacuum. The dry lyophilized powder
is then sealed in the vial and an accompanying vial of water for
injection may be supplied to reconstitute the liquid prior to use.
Parenteral suspensions are prepared in substantially the same
manner except that the compound is suspended in the vehicle instead
of being dissolved and sterilization can be accomplished by
filtration. The compound may be sterilized by exposure to ethylene
oxide before suspending in the sterile vehicle. Advantageously, a
surfactant or wetting agent is included in the composition to
facilitate uniform distribution of the compound.
[0274] The compounds of the invention, e.g., the compounds of
formula (I), may be administered in free form or in
pharmaceutically acceptable salt form, e.g., as indicated above.
Such salts may be prepared in conventional manner and exhibit the
same order of activity as the free compounds.
[0275] In accordance with the foregoing the present invention
further provides: [0276] 1.1. A method for treating and/or
preventing an infectious disorder in a subject, such as a human or
other animal subject, comprising administering to the subject an
effective amount of a compound of the invention, e.g., of formula
(I), a pharmaceutically acceptable salt thereof or a prodrug
thereof. [0277] 1.2. A method for inhibiting PDF in a subject
comprising administering to the subject an effective PDF inhibiting
amount of a compound of the invention, e.g., of formula (I), a
pharmaceutically acceptable salt thereof or a prodrug thereof.
[0278] 2. A compound of the invention, e.g., of formula (I), in
free form or in a pharmaceutically acceptable salt form for use as
a pharmaceutical, e.g., in any method as indicated under 1.1 or 1.2
above. [0279] 3. A pharmaceutical composition, e.g. for use in any
of the methods as in 1.1 or 1.2 above comprising a compound of the
invention, e.g., of formula (I), in free form or pharmaceutically
acceptable salt form, e.g., in association with a pharmaceutically
acceptable diluent or carrier therefor. [0280] 4. A compound of the
invention, e.g., of formula (I), a pharmaceutically acceptable salt
or a prodrug thereof for use as a pharmaceutical or in the
preparation of a pharmaceutical composition for use in any method
as indicated under 1.1 or 1.2 above.
[0281] "Treating" or "treatment" of a disease includes:
(a) preventing the disease, i.e., causing the clinical symptoms of
the disease not to develop in a subject, e.g., a mammal, that may
be exposed to or predisposed to the disease but does not yet
experience or display symptoms of the disease; (b) inhibiting the
disease, i.e., arresting or reducing the development of the disease
or its clinical symptoms; or (c) relieving the disease, i.e.
causing regression of the disease or its clinical symptoms.
[0282] An "effective PDF inhibiting amount" means the amount of a
compound, a pharmaceutically acceptable salt thereof or a prodrug
thereof, that when administered to a subject for treating an
infectious disorder responsive to inhibition of PDF or for
inhibiting PDF, is sufficient to inhibit PDF. The "effective PDF
inhibiting amount" will vary depending on the compound, salt
thereof or prodrug thereof, employed, the microorganism that is
inhibited in the subject, the age, weight, sex, medical condition,
species, disorder and its severity, of the subject to be treated,
and the route of administration, but may nevertheless be readily
determined by one skilled in the art.
[0283] The compounds of the invention, e.g., of formula (I), a
pharmaceutically acceptable salt thereof or prodrug thereof, may be
administered alone or in combination with another therapeutic
agent. Examples of such therapeutic agents include, but are not
limited to, other antibacterial agents, such as .beta.-lactams,
e.g., penicillins; cephalosporins; carbapenems; ketolides;
quinolones, e.g., fluoroquinolones; macrolides, e.g.,
clarithromycin, azithromycin or vancomycin; rifamycins;
monobactams; isoniazid; licosamides; mupirocin; sulfonamides;
phenicols; fosfomycin; glycopeptides; tetracyclines;
streptogramins; chloramphenicol; and oxazolidinone,
anti-inflammatory agents, e.g., corticosteroids or NSAID,
analgesics, e.g., narcotic or non-opioic analgesics.
[0284] In accordance with the foregoing the present invention
provides in a yet further aspect: [0285] 5. A method as defined
above comprising co-administration, e.g., concomitantly or in
sequence, of a therapeutically effective amount of a compound of
the invention, e.g., of formula (I), a pharmaceutically acceptable
salt thereof or a prodrug thereof, and a second drug substance.
[0286] 6. A therapeutic combination, e.g., a kit, comprising: a) a
compound of the invention, e.g., of formula (I), a pharmaceutically
acceptable salt thereof or a prodrug thereof; and b) at least one
second active agent. Component a) and component b) may be used
concomitantly or in sequence. The kit may comprise instructions for
its administration.
[0287] The following are representative pharmaceutical formulations
containing a compound of the invention.
EXAMPLE A
Tablet Formulation
[0288] The following ingredients are mixed intimately and pressed
into single scored tablets.
TABLE-US-00001 Ingredient Quantity per tablet (mg) Compound of this
invention 400 Corn starch 50 Croscarmellose sodium 25 Lactose 120
Magnesium stearate 5
EXAMPLE B
Capsule Formulation
[0289] The following ingredients are mixed intimately and loaded
into a hard-shell gelatin capsule.
TABLE-US-00002 Ingredient Quantity per capsule (mg) Compound of
this invention 200 Lactose, spray-dried 148 Magnesium stearate
2
EXAMPLE C
Suspension Formulation
[0290] The following ingredients are mixed to form a suspension for
oral administration.
TABLE-US-00003 Ingredient Amount Compound of this invention 1.0 g
Fumaric acid 0.5 g Sodium chloride 2.0 g Methyl paraben 0.15 g
Propyl paraben 0.05 g Granulated sugar 25.0 g Sorbitol (70%
solution) 13.00 g Veegum K (Vanderbilt Co.) 1.0 g Flavoring 0.035
mL Colorings 0.5 mg Distilled water q.s. to l00 mL
EXAMPLE D
Injectable Formulation
[0291] The following ingredients are mixed to form an injectable
formulation.
TABLE-US-00004 Ingredient Amount Compound of this invention 0.2-20
mg Sodium acetate buffer solution, 0.4 M 2.0 mL HCl (1 N) or NaOH
(1 N) q.s. to suitable pH Water (distilled, sterile) q.s. to 20
mL
EXAMPLE E
Suppository Formulation
[0292] A suppository of total weight 2.5 g is prepared by mixing
the compound of the invention with Witepsol.RTM. H-15
(triglycerides of saturated vegetable fatty acid; Riches-Nelson,
Inc., NY), and has the following composition:
TABLE-US-00005 Compound of the invention 500 mg Witepsol .RTM. H-15
balance
[0293] The present invention is not limited to the clinical use of
the compounds of the invention, i.e, in the treatment of infection
in a subject. The compounds of the invention are useful to inhibit
bacteria wherever it is desired to inhibit bacteria by contacting
the bacteria with one or more compounds of the invention. Because
of their ability to inhibit bacteria, the compounds of the
invention are particularly useful to prevent contamination of cell
cultures. As used in this context, the term "inhibit" means the
suppression, control, stasis, or kill of bacteria. Eukaryotic
cells, in particular animal cells, are often cultured for various
reasons such as for their ability to produce substances such as
proteins. Examples of such cells include Chinese hamster ovary
cells (CHO cells), African green monkey kidney cells, hybridomoas
constructed by fusing a parent cell (myeloma, etc.) with a useful
substance-producing normal cell (lymphocyte, etc.), and the like.
Typically, the compounds of the invention are incorporated into
cell culture media at a bacteria inhibiting amount, e.g., a
concentration of about 0.0001 mg/mL to about 10 mg/mL, preferably
about 0.0001 mg/mL to about 1 mg/mL, and more preferably about
0.001 mg/mL to about 0.1 mg/mL. Any conventional cell culture
medium known in the art can be used.
[0294] In accordance with the foregoing the present invention
provides in a yet further aspect: [0295] 7. A method for preventing
bacterial contamination of a cell culture medium comprising
incorporating into said cell culture medium a bacteria inhibiting
amount of a compound of the invention, e.g., of formula (I), or a
pharmaceutically acceptable salt thereof. [0296] 8. A cell culture
medium comprising, a bacteria inhibiting amount of a compound of
the invention, e.g., of formula (I), or a pharmaceutically
acceptable salt thereof.
[0297] The foregoing invention has been described in some detail by
way of illustration and example, for purposes of clarity and
understanding. It will be obvious to one of skill in the art that
changes and modifications may be practiced within the scope of the
appended claims. Therefore, it is to be understood that the above
description is intended to be illustrative and not restrictive. The
scope of the invention should, therefore, be determined not with
reference to the above description, but should instead be
determined with reference to the following appended claims, along
with the full scope of equivalents to which such claims are
entitled.
[0298] All patents, patent applications and publications cited in
this application are hereby incorporated by reference in their
entirety for all purposes to the same extent as if each individual
patent, patent application or publication were so individually
denoted.
* * * * *