U.S. patent application number 15/751765 was filed with the patent office on 2018-08-30 for 2-amino-3-fluoro-3-(fluoromethyl)-6-methyl-6-phenyl-3,4,5,6-tetrahydropyri- dines as bace1 inhibitors.
The applicant listed for this patent is H. Lundbeck A/S. Invention is credited to Karsten JUHL, Mauro MARIGO, Lena TAGMOSE.
Application Number | 20180244645 15/751765 |
Document ID | / |
Family ID | 56615973 |
Filed Date | 2018-08-30 |
United States Patent
Application |
20180244645 |
Kind Code |
A1 |
JUHL; Karsten ; et
al. |
August 30, 2018 |
2-AMINO-3-FLUORO-3-(FLUOROMETHYL)-6-METHYL-6-PHENYL-3,4,5,6-TETRAHYDROPYRI-
DINES AS BACE1 INHIBITORS
Abstract
The present invention provides compounds of Formula (I) for the
treatment of neurodegenerative or cognitive diseases. The method
further provides for piperidine-2-one and piperidine-2-thione
intermediates to compounds of formula (I). ##STR00001##
Inventors: |
JUHL; Karsten; (Greve,
DK) ; TAGMOSE; Lena; (Lyngby, DK) ; MARIGO;
Mauro; (Skovlunde, DK) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
H. Lundbeck A/S |
Valby |
|
DK |
|
|
Family ID: |
56615973 |
Appl. No.: |
15/751765 |
Filed: |
August 10, 2016 |
PCT Filed: |
August 10, 2016 |
PCT NO: |
PCT/EP2016/069029 |
371 Date: |
February 9, 2018 |
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07B 2200/05 20130101;
C07D 401/12 20130101; C07D 413/12 20130101; C07D 417/12 20130101;
A61P 25/28 20180101; A61P 43/00 20180101; C07D 211/72 20130101;
C07D 211/76 20130101; C07B 59/002 20130101 |
International
Class: |
C07D 401/12 20060101
C07D401/12; C07B 59/00 20060101 C07B059/00 |
Foreign Application Data
Date |
Code |
Application Number |
Aug 12, 2015 |
DK |
PA201500465 |
Claims
1-16. (canceled)
17. A compound or pharmaceutically acceptable salt thereof, wherein
the compound is ##STR00074##
18. (canceled)
19. A pharmaceutical composition comprising a compound or
pharmaceutically acceptable salt thereof according to claim 17 and
a pharmaceutically acceptable carrier.
20. A method of treating a disease selected from the group
consisting of Alzheimer's disease, preclinical Alzheimer's disease,
prodromal Alzheimer's disease, mild cognitive impairment, Down's
syndrome and cerebral amyloid angiopathy, the method comprising
administering a therapeutically effective amount of a compound or
pharmaceutically acceptable salt thereof according to claim 17 to a
patient in need thereof.
21-22. (canceled)
23. The method of claim 20, wherein the disease is Alzheimer's
disease, preclinical Alzheimer's disease, prodromal Alzheimer's
disease, or mild cognitive impairment.
24-27. (canceled)
28. A method of treating familial Alzheimer's disease or sporadic
Alzheimer's disease, comprising administrating a therapeutically
effective amount of a compound, or pharmaceutically acceptable salt
thereof, according to claim 17 to a patient in need thereof.
29. A method of treating preclinical Alzheimer's disease or
prodromal Alzheimer's disease, comprising administrating a
therapeutically effective amount of a compound, or pharmaceutically
acceptable salt thereof, according to claim 17 to a patient in need
thereof.
30. A method of treating mild cognitive impairment, comprising
administrating a therapeutically effective amount of a compound, or
pharmaceutically acceptable salt thereof, according to claim 17 to
a patient in need thereof.
31. A method of treating Down's syndrome or cerebral amyloid
angiopathy, comprising administrating a therapeutically effective
amount of a compound, or pharmaceutically acceptable salt thereof,
according to claim 17 to a patient in need thereof.
Description
FIELD OF THE INVENTION
[0001] The present invention provides compounds which are BACE1
inhibitors. Separate aspects of the invention are directed to
pharmaceutical compositions comprising said compounds and uses of
the compounds to treat neurodegenerative and cognitive
disorders.
BACKGROUND ART
[0002] Dementia is a clinical syndrome characterized by deficits in
multiple areas of cognition that cannot be explained by normal
aging, a noticeable decline in function, and an absence of
delirium. In addition, neuropsychiatric symptoms and focal
neurological findings are usually present. Dementia is further
classified based on etiology. Alzheimer's disease (AD) is the most
common cause of dementia, followed by mixed AD and vascular
dementia, Lewy body dementia (DLB), and fronto-temporal
dementia.
[0003] .beta.-Amyloid deposits and neurofibrillary tangles are
considered to be major pathologic characterizations associated with
AD which is characterized by the loss of memory, cognition,
reasoning, judgment, and orientation. Also affected, as the disease
progresses, are motor, sensory and linguistic abilities until
global impairment of multiple cognitive functions occurs.
.beta.-Amyloid deposits are predominantly an aggregate of A.beta.
peptide, which in turn is a product of the proteolysis of amyloid
precursor protein (APP) as part of the .beta.-amyloidogenic
pathway. A.beta. peptide results from the cleavage of APP at the
C-terminals by one or more .gamma.-secretases and at the N-terminus
by .beta.-secretase 1 (BACE1) also known as aspartyl protease 2.
BACE1 activity is correlated directly to the generation of A.beta.
peptide from APP. Studies indicate that the inhibition of BACE1
impedes the production of A.beta. peptide. Further, BACE1
co-localizes with its substrate APP in Golgi and endocytic
compartments (Willem M, et al. Semin.Cell Dev. Biol, 2009, 20,
175-182). Knock-out studies in mice have demonstrated the absence
of amyloid peptide formation while the animals are healthy and
fertile (Ohno M, et al. Neurobiol. Dis., 2007, 26, 134-145).
Genetic ablation of BACE1 in APP-overexpressing mice has
demonstrated absence of plaque formation, and the reversal of
cognitive deficits (Ohno M, et al. Neuron; 2004, 41, 27-33). BACE1
levels are elevated in the brains of sporadic AD patients (Hampel
and Shen, Scand. J. Clin. Lab. Invest. 2009, 69, 8-12).
[0004] These convergent findings indicate that the inhibition of
BACE1 may be a therapeutic target for the treatment of AD as well
as neurodegenerative or cognitive disorders for which the reduction
of A.beta. deposits is beneficial.
[0005] AstraZeneca announced the discovery of AZD3839, a potent
BACE1 inhibitor clinical candidate for the treatment of AD
(Jeppsson, F., et al. J. Biol. Chem., 2012, 287, 41245-41257) in
October 2012. The effort which led to the discovery of AZD3839 was
further described in Ginman, T., et al. J. Med. Chem., 2013, 56,
4181-4205. The Ginman publication describes the issues which were
overcome in connection with the discovery and identification of
AZD3839. These issues related to poor blood brain barrier
penetration and P-glycoprotein mediated efflux of the compounds
resulting in lack of brain exposure.
[0006] The Ginman manuscript hypothesized that the differences in
brain exposure would largely be due to the core structures and
Structure Activity Relationship data was provided wherein the in
vitro properties on the reported compounds were given in four
tables according to core sub-types. In Table 6, a series of amidine
containing compounds are described that were considered interesting
from an activity perspective. However, the data suggests that the
amidine containing core did not exhibit a favourable blood brain
barrier permeability profile.
[0007] Researchers from Hoffmann-La Roche and Siena Biotech also
reported the discovery of amidine containing compounds (Woltering,
T. J., et al. Bioorg. Med. Chem. Lett. 2013, 23, 4239-4243). These
compounds (compounds 17 and 18 in the paper) were found not to have
any in vivo effect (lack of A.beta.40 reduction in brain in wild
type mice).
[0008] Contrary to the teachings of Ginman, et al. and Woltering,
T. J., et al., the present inventors have discovered a series of
amidine compounds which are brain penetrating. Accordingly, the
present invention relates to novel compounds having BACE1
inhibitory activity, to their preparation, to their medical use and
to medicaments comprising them.
SUMMARY OF THE INVENTION
[0009] An objective of the present invention is to provide
compounds that inhibit BACE1. Accordingly, the present invention
relates to compounds of Formula I.
##STR00002##
[0010] wherein Ar is selected from the group consisting of phenyl,
pyridyl, pyrimidyl, pyrazinyl, imidazolyl, pyrazolyl, thiazolyl,
oxazolyl, isoxazolyl, and where Ar is optionally substituted with
one or more substituents selected from halogen, CN, C.sub.1-C.sub.6
alkyl, C.sub.2-C.sub.6 alkenyl, C.sub.2-C.sub.6 alkynyl,
C.sub.1-C.sub.6 fluoroalkyl or C.sub.1-C.sub.6 alkoxy; and
[0011] R.sup.1 is one or more hydrogen, halogen, C.sub.1-C.sub.3
fluoroalkyl or C.sub.1-C.sub.3 alkyl;
[0012] R.sup.2 is hydrogen or fluoro
[0013] or a pharmaceutically acceptable salt thereof.
[0014] In one embodiment the present invention provides compounds
of Formula I or a pharmaceutically acceptable salt thereof for use
in therapy.
[0015] The present invention further provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable carrier.
[0016] In one embodiment the invention provides the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
in the manufacture of a medicament for the treatment of
neurodegenerative or cognitive disorder.
[0017] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof for use in
a method for the treatment of a neurodegenerative or cognitive
disorder.
[0018] The present invention provides a method of treating a
neurodegenerative or cognitive disorder comprising administering a
therapeutically effective amount of a compound of Formula I or a
pharmaceutically acceptable salt thereof to a patient in need
thereof.
DETAILED DESCRIPTION OF THE INVENTION
[0019] One embodiment the present invention provides compounds of
Formula Ia or Ib
##STR00003##
[0020] In one embodiment R.sup.1 is halogen, and in particularly
R.sup.1 is fluoro.
[0021] In one embodiment R.sup.2 is hydrogen.
[0022] In one embodiment R.sup.2 is fluoro.
[0023] In one embodiment Ar is pyridyl.
[0024] In one embodiment Ar is pyrimidyl.
[0025] In one embodiment Ar is pyrazinyl.
[0026] In one embodiment Ar is oxazolyl.
[0027] In one embodiment Ar is imidazolyl.
[0028] In one embodiment Ar is thiazolyl.
[0029] In one embodiment Ar is pyrazolyl.
[0030] In one embodiment Ar is isoxazolyl
[0031] In one embodiment Ar is phenyl.
[0032] In one embodiment, Ar is substituted by one or more
substituents selected from the group consisting of F, Cl, Br, CN,
C.sub.1-C.sub.3 alkyl, C.sub.1-C.sub.3 haloalkyl or C.sub.1-C.sub.3
alkoxy.
[0033] In one embodiment, a compound of the present invention is
selected from the group consisting of
[0034]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-methoxypicolinamide;
[0035]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-(methoxy-d3)picolinamide;
[0036]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-cyano-3-methylpicolinamide;
[0037]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-chloropicolinamide;
[0038]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-methoxypyrazine-2-carboxamide;
[0039]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-2-methyloxazole-4-carboxamide;
[0040]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-4-bromo-1-methyl-1H-imidazole-2-carb-
oxamide;
[0041]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-4-methylthiazole-2-carboxamide;
[0042]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-2-(difluoromethyl)oxazole-4-carboxam-
ide;
[0043]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-1-(difluoromethyl)-1H-pyrazole-3-car-
boxamide;
[0044]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-(difluoromethyl)pyrazine-2-carboxa-
mide;
[0045]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-4-chlorobenzamide;
[0046]
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-fluoropicolinamide;
[0047]
N-(3-((2S,5R)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-te-
trahydropyridin-2-yl)-4-fluorophenyl)-5-methoxypicolinamide;
[0048]
N-[3-[(2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-fluoro-pyridine-2-carboxamide;
[0049]
N-[3-[(2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-methoxy-pyridine-2-carboxamide;
[0050]
N-[3-[(2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-methoxy-pyrazine-2-carboxamide;
[0051]
N-[3-[(2S,5R)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-fluoro-pyridine-2-carboxamide;
[0052]
N-[3-[(2S,5R)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-methoxy-pyridine-2-carboxamide;
[0053]
N-[3-[(2S,5R)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydr-
opyridin-2-yl]-4,5-difluorophenyl]-5-methoxy-pyrazine-2-carboxamide;
[0054]
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyr-
idin-2-yl]-4-fluorophenyl]-5-fluoro-pyridine-2-carboxamide;
[0055]
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyr-
idin-2-yl]-4-fluorophenyl]-5-methoxy-pyridine-2-carboxamide;
[0056]
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyr-
idin-2-yl]-4-fluorophenyl]-5-methoxy-pyrazine-2-carboxamide;
[0057]
N-[3-[(2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyr-
idin-2-yl]-4-fluorophenyl]-5-fluoro-pyridine-2-carboxamide;
[0058]
N-[3-[(2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyr-
idin-2-yl]-4-fluorophenyl]-5-methoxy-pyridine-2-carboxamide and
[0059]
N-[3-R2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyri-
din-2-yl]-4-fluorophenyl]-5-methoxy-pyrazine-2-carboxamide;
[0060] or a pharmaceutically acceptable salt thereof.
[0061] As used herein, the term "C.sub.1-C.sub.6 alkyl" refers to a
straight chained or branched saturated hydrocarbon having from one
to six carbon atoms inclusive. Examples of C.sub.1-C.sub.6 alkyl
include, but are not limited to, methyl, ethyl, 1-propyl, 2-propyl,
1-butyl, 2-butyl, 2-methyl-2-propyl, 2-methyl-1-propyl, n-pentyl
and n-hexyl. Similarly, the term "C.sub.1-C.sub.3 alkyl" refers to
a straight chained or branched saturated hydrocarbon having from
one to three carbon atoms inclusive. Examples of such substituents
include, but are not limited to, methyl, ethyl and n-propyl.
[0062] Likewise, the term "C.sub.1-C.sub.6 alkoxy" refers to a
straight chained or branched saturated alkoxy group having from one
to six carbon atoms inclusive with the open valency on the oxygen.
Examples of C.sub.1-C.sub.6 alkoxy include, but are not limited to,
methoxy, ethoxy, n-butoxy, t-butoxy and n-hexyloxy. The
"C.sub.1-C.sub.6 alkoxy" is optionally substituted with one or more
fluorine atoms.
[0063] As used herein, the term "C.sub.1-C.sub.6 fluoroalkyl"
refers to a straight chained or branched saturated hydrocarbon
having from one to six carbon atoms inclusive substituted with one
or more fluorine atoms. Examples of C.sub.1-C.sub.6 fluoroalkyl
include, but are not limited to, trifluoromethyl, pentafluoroethyl,
1-fluoroethyl, monofluoromethyl, difluoromethyl, 1,2-difluoroethyl
and 3,4 difluorohexyl. Similarly, the term "C.sub.1-C.sub.3
fluoroalkyl" refers to a straight chained or branched saturated
hydrocarbon having from one to three carbon atoms inclusive
substituted with one or more fluorine atoms per carbon atom.
[0064] The term "halogen" refers to fluoro, chloro, bromo and
iodo.
[0065] The term "C.sub.2-C.sub.6 alkenyl" refers to a branched or
unbranched alkenyl group having from two to six carbon atoms and
one double bond, including but not limited to ethenyl, propenyl,
and butenyl.
[0066] The term "C.sub.2-C.sub.6 alkynyl" shall mean a branched or
unbranched alkynyl group having from two to six carbon atoms and
one triple bond, including but not limited to ethynyl, propynyl and
butynyl.
[0067] The phrase "therapeutically effective amount" when applied
to a compound of the invention is intended to denote an amount of
the compound that is sufficient to ameliorate, palliate, stabilize,
reverse, slow or delay the progression of a disorder or disease
state, or of a symptom of the disorder or disease. In an
embodiment, the method of the present invention provides for
administration of combinations of compounds. In such instances, the
"therapeutically effective amount" is the amount of a compound of
the present invention in the combination sufficient to cause the
intended biological effect.
[0068] The term "treatment" or "treating" as used herein means
ameliorating or reversing the progress or severity of a disease or
disorder, or ameliorating or reversing one or more symptoms or side
effects of such disease or disorder. "Treatment" or "treating", as
used herein, also means to inhibit or block, as in retard, arrest,
restrain, impede or obstruct, the progress of a system, condition
or state of a disease or disorder. For purposes of this invention,
"treatment" or "treating" further means an approach for obtaining
beneficial or desired clinical results, where "beneficial or
desired clinical results" include, without limitation, alleviation
of a symptom, diminishment of the extent of a disorder or disease,
stabilized (i.e., not worsening) disease or disorder state, delay
or slowing of a disease or disorder state, amelioration or
palliation of a disease or disorder state, and remission of a
disease or disorder, whether partial or total.
[0069] The present invention is based on the discovery that
compounds of Formula I are inhibitors of BACE1, and as such, are
useful for the treatment of disorders which pathological
characteristics comprise .beta.-amyloid deposits and
neurofibrillary tangles, such as neurodegenerative or cognitive
disorders.
[0070] The compounds of the present invention are, as discussed
above, expected to be useful in the treatment of Alzheimer's
disease due to their effects on .beta.-amyloid deposits and
neurofibrillary tangles. This includes familial Alzheimer's disease
where patients carry mutations on specific genes intimately
involved in the production of A.beta. peptide. It is, however,
important to note that aggregates of A.beta. peptide is not limited
to familial Alzheimer's disease but is similarly an important
pathophysiological characteristics of the more common sporadic
Alzheimer's disease [Mol Cell Neurosci, 66, 3-11, 2015].
[0071] The compounds of the present invention are also believed to
be useful in the treatment of early-stage Alzheimer's disease, i.e.
disease stages where the biological and structural changes have
started but the clinical manifestations of the disease have not yet
become evident or are not yet well developed. Early-stage
Alzheimer's disease may, in fact, start years before any clinical
manifestation of the disease becomes manifest. Early-stage
Alzheimer's disease includes prodromal Alzheimer's disease,
preclinical Alzheimer's disease and mild cognitive impairment.
Although mild cognitive impairment may be unrelated to Alzheimer's
disease it is often a transitional stage to Alzheimer's disease or
due to Alzheimer's disease. Preclinical and prodromal Alzheimer's
disease are asymptomatic stages, and they are typically diagnosed
by the presence of Alzheimer's disease related biomarkers. In this
context the compounds of the present invention are believed to be
useful in slowing down the progression of early-stage Alzheimer's
disease, such as mild cognitive impairment to Alzheimer's disease.
The compounds of the present invention are also believed to be
useful in the treatment of memory loss, attention deficits and
dementia associated with Alzheimer's disease.
[0072] Other diseases, in addition to the continuum of Alzheimer's
disease, are characterized by .beta.-amyloid deposits and
neurofibrillary tangles. This includes e.g. Trisomy 21 also known
as Down's syndrome. Patients suffering from Down's syndrome have an
extra chromosome 21 which chromosome contains the gene for the
amyloid precursor protein (APP). The extra chromosome 21 leads to
overexpression of APP, which leads to increased levels of A.beta.
peptide, which eventually causes the markedly increased risk of
developing Alzheimer's disease seen in Down's syndrome patients
[Alzheimer's & Dementia, 11, 700-709, 201]. Cerebral amyloid
angiopathy is also characterized by .beta.-amyloid deposits and
neurofibrillary tangles in blood vessels of the central nervous
system [Pharmacol Reports, 67, 195-203, 2015] and is as such
expected to be treatable with compounds of the present
invention.
[0073] In one embodiment, the present invention provides a method
of treating a disease selected from Alzheimer's disease (familial
or sporadic), preclinical Alzheimer's disease, prodromal
Alzheimer's disease, mild cognitive impairment, Down's syndrome and
cerebral amyloid angiopathy, the method comprising the
administration of a therapeutically effective amount of a compound
of Formula I or a pharmaceutically acceptable salt thereof to a
patient in need thereof. The compounds of the invention are
believed to be useful in the treatment of a disease is selected
from Alzheimer's disease (familial or sporadic), preclinical
Alzheimer's disease, prodromal Alzheimer's disease and mild
cognitive impairment.
[0074] The present invention further provides a method of
inhibiting BACE1 in a patient comprising administering to a patient
in need thereof a therapeutically effective amount of a compound of
Formula I or a pharmaceutically acceptable salt thereof.
[0075] The present invention also provides a method of inhibiting
.beta.-secretase mediated cleavage of amyloid precursor protein
comprising administering to a patient in need of such treatment a
therapeutically effective amount a compound of Formula I or a
pharmaceutically acceptable salt thereof.
[0076] In further embodiments, the present invention provides the
use of a compound of Formula I or a pharmaceutically acceptable
salt thereof for the manufacture of a medicament for the treatment
of a disease selected from Alzheimer's disease (familial or
sporadic), preclinical Alzheimer's disease, prodromal Alzheimer's
disease, mild cognitive impairment, Down's syndrome or cerebral
amyloid angiopathy.
[0077] The present invention also provides the use of a compound of
Formula I or a pharmaceutically acceptable salt thereof for the
manufacture of a medicament for the inhibition of BACE1. The
present invention further provides the use of a compound of Formula
I or a pharmaceutically acceptable salt thereof for the manufacture
of a medicament for the inhibition of production or accumulation of
A.beta. peptide.
[0078] In one embodiment, the present invention provides a compound
of Formula I or a pharmaceutically acceptable salt thereof for use
in a method for the treatment of a disease selected form
Alzheimer's disease (familial or sporadic), preclinical Alzheimer's
disease, prodromal Alzheimer's disease, mild cognitive impairment,
Down's syndrome or cerebral amyloid angiopathy.
[0079] In one embodiment, the present invention relates to a
compound of Formula I or a pharmaceutically acceptable salt thereof
for use in a method for inhibiting of BACE1 or in a method for
inhibiting of production or accumulation of A.beta. peptide.
[0080] The compounds of the present invention are as demonstrated
in the examples potent inhibitors of BACE1 and capable of lowering
the level of A.beta. peptide in rat brain and plasma, and said
compounds are thus believed to be useful in the treatment of
neurodegenerative and cognitive disorders which pathological
characteristics comprise A.beta. deposits and neurofibrilary
tangles, such as e.g. Alzheimer's disease. It may be beneficial to
combine a compound of the present invention with another treatment
paradigm useful in the treatment of such disease, e.g. Alzheimer's
disease.
[0081] Tau proteins are abundant in neurons. Tau proteins are
soluble, highly phosphorylation labile and bind to tubulin
providing regulation and modulation of tubulin assembly, i.e.
eventually the microtubular structure and stability. Tau proteins
can only associate with tubulin in the most de-phosphorylated
state, and phosphorylation/de-phosphorylation acts as a switch
controlling the tubulin association. Phosphorylated Tau constitutes
an important part of the neurofibrillary tangles which are one of
the hallmarks of Alzheimer's disease. The so-called Tau hypothesis
suggests targeting these pathological tangles, a main constituent
of which is phosphorylated Tau protein, as a treatment paradigm for
Alzheimer's disease. In particular, immunotherapies, both active
and passive, have been suggested as a way to target Tau
neurofibrillary tangles. In active immunotherapy, a pathogenic
antigen is injected into the patient and the innate immune system
elicits an immune response. This triggers the maturation of B-cells
generating high affinity antibodies against the administered
antigen. In a passive immunotherapy, the triggering of the innate
immune system is circumvented by infusing a specific antibody
against the antigen. It is suggested that the inherent clearance
system then removes antibody bound ligand. Substantial evidence for
the efficacy of both active and passive immunotherapy targeting
phosphorylated Tau protein as a treatment for Alzheimer's disease
exists [Alzheimer's & Dementia, 7(4, suppl) S480-481; J
Neurosci 30, 16559-16556, 2010; J Neurosci, 27, 9115-9129,
2007].
[0082] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) a
compound useful in active or passive Tau immunotherapy to a patient
in need thereof. Said compound useful in passive Tau immunotherapy
may be an antibody directed against phosphorylated Tau protein.
Said compound useful in active Tau immunotherapy may be a fragment
of the Tau protein amino acid sequence which upon injection in a
patient elicits antibodies against phosphorylated Tau protein in
said patient. The administration according to this embodiment of
the invention may be simultaneous, or there may be a time gap
between the administration of the two components.
[0083] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and a compound useful in active or passive Tau immunotherapy in the
manufacture of a medicament for the treatment of neurodegenerative
or cognitive disorder, e.g. Alzheimer's disease.
[0084] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and a
compound useful in active or passive Tau immunotherapy for use in a
method for the treatment of a neurodegenerative or cognitive
disorder, e.g. Alzheimer's disease.
[0085] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and a compound useful in
active or passive Tau immunotherapy and a pharmaceutically
acceptable carrier.
[0086] Another paradigm to treat neurodegenerative and cognitive
disorder, e.g. Alzheimer's disease is to target the A.beta.
peptides. It has been suggested that this can be achieved by either
passive or active immunotherapy targeting A.beta. peptides [J
Neurosci, 34, 11621-11630, 2014; J Neurosci 33, 4923-4934, 2013].
In combination with compounds of the present invention this would
attempt to target the same pathological mechanism via two different
routes. Anti-A.beta. antibodies (either injected directly into the
patient or generated in the patient as a result of active
immunotherapy) clear A.beta. deposits in the brain, while further
accumulation of A.beta. peptide is blocked or reduced by the
compounds of the present invention.
[0087] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) a
compound useful in active or passive A.beta. peptide immunotherapy
to a patient in need thereof. Said compound useful in passive
A.beta. peptide immunotherapy may be an anti-A.beta. peptide
antibody, such as gantenerumab, solanezumab, aducanumab or
crenezumab. Said compound useful in active A.beta. peptide
immunotherapy may be a fragment of the A.beta. peptide amino acid
sequence which upon injection into a patient elicits anti-A.beta.
peptide antibodies in said patient. The administration according to
this embodiment of the invention may be simultaneous, or there may
be a time gap between the administration of the two components.
[0088] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and a compound useful in active or passive A.beta. peptide
immunotherapy in the manufacture of a medicament for the treatment
of neurodegenerative or cognitive disorder, e.g. Alzheimer's
disease.
[0089] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and a
compound useful in active or passive A.beta. peptide immunotherapy
for use in a method for the treatment of a neurodegenerative or
cognitive disorder, e.g. Alzheimer's disease.
[0090] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and a compound useful in
active or passive A.beta. peptide immunotherapy and a
pharmaceutically acceptable carrier.
[0091] The NMDA (N-Methyl-D-Aspartate) receptor antagonist
memantine and the acetylcholine esterase inhibitors donepezil,
rivastigmine and galantamine are approved drugs for the treatment
of Alzheimer's disease.
[0092] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) an
NMDA receptor antagonist or an acetylcholine esterase inhibitor to
a patient in need thereof. The administration according to this
embodiment of the invention may be simultaneous, or there may be a
time gap between the administration of the two components.
[0093] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and an NMDA receptor antagonist or an acetylcholine esterase
inhibitor in the manufacture of a medicament for the treatment of
neurodegenerative or cognitive disorder, e.g. Alzheimer's
disease.
[0094] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and an NMDA
receptor antagonist or an acetylcholine esterase inhibitor for use
in a method for the treatment of a neurodegenerative or cognitive
disorder, e.g. Alzheimer's disease.
[0095] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and an NMDA receptor
antagonist or an acetylcholine esterase inhibitor and a
pharmaceutically acceptable carrier.
[0096] Seizures or epileptiform activity are also associated with
Alzheimer's disease, including early stages of Alzheimer's disease,
and treatment of said epileptic activity, which seeks to normalise
hippocampal hyperactivity, may form part of an Alzheimer's disease
treatment paradigm [JAMA Neurol, 70, 1158-1166, 2013; J Neurosci
Res, 93, 454, 465, 2015; Neuron, 74, 647-474, 2012;
Neurepsychpharm, 35, 1016-1025, 2010; CNS Neurosci Ther, 19,
871-881, 2013]. Useful antiepileptics include NMDA receptor
antagonists and ion channel modulators, such as topiramate,
levetiracetam and lamotrigine.
[0097] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) an
antiepileptic to a patient in need thereof. The administration
according to this embodiment of the invention may be simultaneous,
or there may be a time gap between the administration of the two
components.
[0098] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and an antiepileptic in the manufacture of a medicament for the
treatment of neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease.
[0099] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and an
antiepileptic for use in a method for the treatment of a
neurodegenerative or cognitive disorder, e.g. Alzheimer's
disease.
[0100] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and an antiepileptic and a
pharmaceutically acceptable carrier.
[0101] Emerging evidence suggests that inflammation has a causal
role in Alzheimer's disease pathogenesis and that neuroinflammation
is not a passive system activated by emerging .beta.-amyloid
deposits and neurofibrilary tangles, but also contributes to
pathogenesis itself [Lancet Neurol, 14, 388-405, 2015; J Alz Dis,
44, 385-396, 2015; Neurol, 84, 2161-2168, 2015]. It follows from
this that anti-inflammatory drugs, such as NSAID (non-steriod
anti-inflammatory drugs), TNF.alpha. inhibitors, such as etanercept
and p38 MAP kinase inhibitors, such as VX-745
(5-(2,6-Dichlorophenyl)-2-((2,4-difluorophenyl)thio)-6H-pyrimido[1-
,6-b]pyridazin-6-one) may be useful in the treatment of Alzheimer's
disease.
[0102] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) an
anti-inflammatory drug to a patient in need thereof. The
administration according to this embodiment of the invention may be
simultaneous, or there may be a time gap between the administration
of the two components.
[0103] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and anti-inflammatory drug in the manufacture of a medicament for
the treatment of neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease.
[0104] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and an
anti-inflammatory drug for use in a method for the treatment of a
neurodegenerative or cognitive disorder, e.g. Alzheimer's
disease.
[0105] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and an anti-inflammatory
drug and a pharmaceutically acceptable carrier.
[0106] In addition, efficacy in the treatment of Alzheimer's
disease has been demonstrated for Tau protein aggregation
inhibitors, such as TRX-0237, also known as Methylene Blue, and
SSRIs (Selective Serotonin Reuptake Inhibitor), such as citalopram
[Behav Pharmacol, 26, 353-368, 2015; Sci Transl Med, 6(236re4),
2014].
[0107] In one embodiment the invention provides a method for the
treatment of a neurodegenerative or cognitive disorder, e.g.
Alzheimer's disease, the method comprising the administration of a
therapeutically effect amount of two components (1) a compound of
Formula I or a pharmaceutically acceptable salt thereof and (2) Tau
protein aggregation inhibitor or an SSRI to a patient in need
thereof. The administration according to this embodiment of the
invention may be simultaneous, or there may be a time gap between
the administration of the two components.
[0108] In one embodiment, the invention relates to the use of a
compound of Formula I or a pharmaceutically acceptable salt thereof
and a Tau protein aggregation inhibitor or an SSRI in the
manufacture of a medicament for the treatment of neurodegenerative
or cognitive disorder, e.g. Alzheimer's disease.
[0109] In one embodiment, the invention provides a compound of
Formula I or a pharmaceutically acceptable salt thereof and a Tau
protein aggregation inhibitor or an SSRI drug for use in a method
for the treatment of a neurodegenerative or cognitive disorder,
e.g. Alzheimer's disease.
[0110] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and a Tau protein
aggregation inhibitor or an SSRI drug and a pharmaceutically
acceptable carrier.
[0111] In one embodiment, a mammal is a human.
[0112] In one embodiment, the patient is a human patient.
[0113] Pharmaceutically Acceptable Salts
[0114] The compounds of this invention are generally used as the
free base or as a pharmaceutically acceptable salt thereof.
Pharmaceutically acceptable salts of a compound of Formula I are
prepared e.g. in a conventional manner by treating a solution or
suspension of a free base of Formula I with a molar equivalent of a
pharmaceutically acceptable acid. Representative examples of
suitable organic and inorganic acids are described below. Such
salts include pharmaceutically acceptable acid addition salts. Acid
addition salts include salts of inorganic acids as well as organic
acids.
[0115] Representative examples of suitable inorganic acids include
hydrochloric, hydrobromic, hydroiodic, phosphoric, sulfuric,
sulfamic, nitric acids and the like.
[0116] Representative examples of suitable organic acids include
formic, acetic, trichloroacetic, trifluoroacetic, propionic,
benzoic, cinnamic, citric, fumaric, glycolic, itaconic, lactic,
methanesulfonic, maleic, malic, malonic, mandelic, oxalic, picric,
pyruvic, salicylic, succinic, methane sulfonic, ethanesulfonic,
tartaric, ascorbic, pamoic, bismethylene salicylic,
ethanedisulfonic, gluconic, citraconic, aspartic, stearic,
palmitic, EDTA, glycolic, p-aminobenzoic, glutamic,
benzenesulfonic, p-toluenesulfonic acids, theophylline acetic
acids, as well as the 8-halotheophyllines (for example,
8-bromotheophylline and the like). Further examples of
pharmaceutically acceptable inorganic or organic acid addition
salts include the pharmaceutically acceptable salts listed in S. M.
Berge, et al., J. Pharm. Sci., 1977, 66, 2.
[0117] Furthermore, the compounds of this invention may exist in
unsolvated as well as in solvated forms with pharmaceutically
acceptable solvents such as water, ethanol and the like.
[0118] The compounds of the present invention may have one or more
asymmetric centres and it is intended that any optical isomers
(i.e. enantiomers or diastereomers), as separated, pure or
partially purified optical isomers and any mixtures thereof
including racemic mixtures, i.e. a mixture of stereoisomeres, are
included within the scope of the invention.
[0119] In this context is understood that when specifying the
enantiomeric form, then the compound is in enantiomeric excess,
e.g. essentially in a pure form. Accordingly, one embodiment of the
invention relates to a compound of the invention having an
enantiomeric excess of at least 60%, at least 70%, at least 80%, at
least 85%, at least 90%, at least 96%, preferably at least 98%.
[0120] Racemic forms may be resolved into the optical antipodes by
known methods, for example, by separation of diastereomeric salts
thereof with an optically active acid, and liberating the optically
active amine compound by treatment with a base.
[0121] Separation of such diastereomeric salts can be achieved,
e.g. by fractional crystallization. The optically active acids
suitable for this purpose may include, but are not limited to d- or
l-tartaric, mandelic or camphorsulfonic acids. Another method for
resolving racemates into the optical antipodes is based upon
chromatography on an optically active matrix. The compounds of the
present invention may also be resolved by the formation and
chromatographic separation of diastereomeric derivatives from
chiral derivatizing reagents, such as, chiral alkylating or
acylating reagents, followed by cleavage of the chiral auxiliary.
Any of the above methods may be applied either to resolve the
optical antipodes of the compounds of the invention per se or to
resolve the optical antipodes of synthetic intermediates, which can
then be converted by methods described herein into the optically
resolved final products which are the compounds of the
invention.
[0122] Additional methods for the resolution of optical isomers,
known to those skilled in the art, may be used. Such methods
include those discussed by J. Jaques, A. Collet and S. Wilen in
Enantiomers, Racemates, and Resolutions, John Wiley and Sons, New
York, 1981. Optically active compounds can also be prepared from
optically active starting materials.
[0123] Pharmaceutical Compositions
[0124] The present invention further provides a pharmaceutical
composition comprising a compound of Formula I or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable carrier. The present invention also provides a
pharmaceutical composition comprising a specific compound disclosed
in the Experimental Section or a pharmaceutically acceptable salt
thereof and a pharmaceutically acceptable carrier.
[0125] The compounds of the invention may be administered alone or
in combination with pharmaceutically acceptable carriers or
excipients, in either single or multiple doses. The pharmaceutical
compositions according to the invention may be formulated with
pharmaceutically acceptable carriers or diluents as well as any
other known adjuvants and excipients in accordance with
conventional techniques such as those disclosed in Remington: The
Science and Practice of Pharmacy, 22.sup.th Edition, Gennaro, Ed.,
Mack Publishing Co., Easton, Pa., 2013.
[0126] Pharmaceutical compositions for oral administration include
solid dosage forms such as capsules, tablets, dragees, pills,
lozenges, powders and granules. Where appropriate, the compositions
may be prepared with coatings such as enteric coatings or they may
be formulated so as to provide controlled release of the active
ingredient such as sustained or prolonged release according to
methods well known in the art. Liquid dosage forms for oral
administration include solutions, emulsions, suspensions, syrups
and elixirs. Pharmaceutical compositions for parenteral
administration include sterile aqueous and nonaqueous injectable
solutions, dispersions, suspensions or emulsions as well as sterile
powders to be reconstituted in sterile injectable solutions or
dispersions prior to use. Other suitable administration forms
include, but are not limited to, suppositories, sprays, ointments,
creams, gels, inhalants, dermal patches and implants.
[0127] Typical oral dosages range from about 0.01 to about 100
mg/kg body weight per day.
[0128] Suitable pharmaceutical carriers include inert solid
diluents or fillers, sterile aqueous solutions and various organic
solvents. Examples of solid carriers include lactose, terra alba,
sucrose, cyclodextrin, talc, gelatin, agar, pectin, acacia,
magnesium stearate, stearic acid and lower alkyl ethers of
cellulose. Examples of liquid carriers include, but are not limited
to, syrup, peanut oil, olive oil, phospholipids, fatty acids, fatty
acid amines, polyoxyethylene and water. Similarly, the carrier or
diluent may include any sustained release material known in the
art, such as glyceryl monostearate or glyceryl distearate, alone or
mixed with a wax. The pharmaceutical compositions formed by
combining the compounds of Formula I or a pharmaceutically
acceptable salt thereof and a pharmaceutically acceptable carrier
are readily administered in a variety of dosage forms suitable for
the disclosed routes of administration. The formulations may
conveniently be presented in unit dosage form by methods known in
the art of pharmacy.
[0129] If a solid carrier is used for oral administration, the
preparation may be tabletted, placed in a hard gelatin capsule in
powder or pellet form or it may be in the form of a troche or
lozenge. The amount of solid carrier will vary widely but will
range from about 25 mg to about 1 g per dosage unit. If a liquid
carrier is used, the preparation may be in the form of a syrup,
emulsion, soft gelatin capsule or sterile injectable liquid such as
an aqueous or non-aqueous liquid suspension or solution.
[0130] Process and Key Intermediates
[0131] Key intermediates in the preparation of a compounds of the
invention are
(6S)-3-Fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-2-one-
,
(6S)-6-(2,3-Difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiperidin--
2-one, and
(6S)-3-Fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-
-2-one. Accordingly, one aspect of the invention is directed to a
compound selected from the group consisting of
##STR00004##
[0132]
(6S)-3-fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-
-2-one and salts thereof;
##STR00005##
[0133]
(6S)-6-(2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiper-
idin-2-one, and salts thereof, and
##STR00006##
[0134]
(6S)-3-fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-2-o-
ne , and salts thereof.
[0135]
(6S)-3-Fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-
-2-one and salts thereof,
(6S)-6-(2,3-Difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiperidin-2-
-one and salts thereof, and
(6S)-3-Fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-2-one
and salts thereof are central intermediates to the compounds of the
invention. Coupling to an aromatic selected from phenyl, pyridyl,
pyrimidyl, pyrazinyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl,
isoxazolyl,according to the invention requires functionalization of
the phenyl ring of the above phenyl piperidinones. As shown in the
Experimental section, the key intermediates
(6S)-3-Fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-2-one
and salts thereof,
(6S)-6-(2,3-Difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiperidin-2-
-one and salts thereof, and
(6S)-3-Fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-2-one
and salts thereof were converted to further key intermediates of
the invention, namely
(3S,6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one,
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one,
(3S,6S)-6-(2,3-Difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidin-2-one,
(3R,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidin-2-one and
(6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piperidin-2-
-one. Accordingly, a further aspect of the invention relates to
compounds selected from the group consisting of
##STR00007##
[0136]
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-meth-
ylpiperidin-2-one
##STR00008##
[0137]
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-meth-
ylpiperidin-2-one
##STR00009##
[0138]
(3R,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidin-2-one
##STR00010##
[0139]
(3S,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidin-2-one and
##STR00011##
[0140]
(6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piper-
idin-2-one and salts thereof.
[0141] As can be seen in greater detail from the experimental
section below, the phenyl amino group on an intermediate provides
for the key coupling step:
##STR00012##
[0142] Compounds of formula XVI and salts thereof, and their
immediate precursors, compounds of formula XVa and XVb and salts
thereof are therefore also key intermediate compounds of the
invention.
##STR00013##
[0143] Accordingly, a further aspect of the invention is directed
to compounds selected from the group consisting of
##STR00014##
[0144]
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-meth-
ylpiperidine-2-thione
##STR00015##
[0145]
(3R,6S)-3-fluoro-6-((R)-2-fluoro-5-nitrocyclohexa-2,4-dien-1-yl)-3--
(fluoromethyl)-6-methylpiperidine-2-thione
##STR00016##
[0146]
(3S,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione
##STR00017##
[0147] (3R,6S)-6-(2,
3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiperidine-2--
thione and
##STR00018##
[0148]
(6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piper-
idine-2-thione and salts thereof.
[0149] The intermediate compounds involved in the coupling step are
a further aspect of the invention, namely compounds selected from
the group consisting of
##STR00019##
[0150]
(3S,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-meth-
ylpiperidine-2-thione
##STR00020##
[0151]
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-meth-
ylpiperidine-2-thione
##STR00021##
[0152]
(3S,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione
##STR00022##
[0153]
(3R,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione and
##STR00023##
[0154]
(6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piper-
idine-2-thione and salts thereof.
[0155] Experimental Section
[0156] The compounds of the present invention of general formula I,
wherein R.sup.1 and Ar are as defined above can be prepared by the
methods outlined in the following reaction schemes 1-5 and in the
examples. In the described methods, it is possible to make use of
variants or modifications, which are themselves known to chemists
skilled in the art or could be apparent to the person of ordinary
skill in this art. Furthermore, other methods for preparing
compounds of the invention will be readily apparent to the person
skilled in the art in light of the following reaction schemes and
examples.
[0157] It may be necessary to incorporate protection and
deprotection strategies for substituents such as amino, amido, keto
and hydroxyl groups in the synthetic methods described below to
synthesize the compounds of Formula I. Methods for protection and
deprotection of such groups are well known in the art, and may be
found in T. Green, et al., Protective Groups in Organic Synthesis,
1991, 2.sup.nd Edition, John Wiley & Sons, New York.
[0158] For compounds, which can exist as a mixture or equilibrium
between two or more tautomers, only one tautomer is represented in
the schemes, although it may not be the most stable tautomer. For
compounds, which can exist in enantiomeric, stereoisomeric or
geometric isomeric forms their geometric configuration is
specified; otherwise the structure represents a mixture of
stereoisomers.
[0159] Analytical LC-MS data was obtained using the following
methods.
[0160] Method A:
[0161] LC-MS was run on Waters Aquity UPLC-MS consisting of Waters
Aquity including column manager, binary solvent manager, sample
organizer, PDA detector (operating at 254 nm), ELS detector, and
SQ-MS equipped with APPI-source operating in positive ion mode.
[0162] LC-conditions: The column was Acquity UPLC BEH C18 1.7
.mu.m; 2.1.times.150 mm operating at 60.degree. C. with 0.6 ml/min
of a binary gradient consisting of water+0.05% trifluoroacetic acid
(A) and acetonitrile+5% water+0.03% trifluoroacetic acid (B).
Gradient: 0.00 min: 10% B; 3.00 min: 99.9% B; 3.01 min: 10% B; 3.60
min: 10% B. Total run time: 3.60 min.
[0163] Method B:
[0164] LC-MS was run on Waters Acquity UPLC-MS consisting of Waters
Acquity including column manager, binary solvent manager, sample
organizer, PDA detector (operating at 254 nm), ELS detector, and
TQ-MS equipped with APPI-source operating in positive ion mode.
[0165] LC-conditions: The column was Acquity UPLC BEH C18 1.7
.mu.m; 2.1.times.50 mm operating at 60.degree. C. with 1.2 ml/min
of a binary gradient consisting of water+0.05% trifluoroacetic acid
(A) and acetonitrile+5% water+0.05% trifluoroacetic acid (B).
Gradient: 0.00 min: 10% B; 1.00 min: 100% B; 1.01 min: 10% B; 1.15
min: 10% B. Total run time: 1.15 min.
[0166] Method C:
[0167] An Agilent 1200 LCMS system with ELS detector was used.
Column: Agilent TC-C18 5 .mu.m; 2.1.times.50 mm; Column
temperature: 50.degree. C.; Solvent system: A=water/trifluoroacetic
acid (99.9:0.1) and B=acetonitrile/trifluoroacetic acid
(99.95:0.05); Method: Linear gradient elution with A:B=99:1 to
0:100 in 4.0 minutes and with a flow rate of 0.8 mL/minute.
[0168] Method D:
[0169] An Agilent 1200 LCMS system with ELS detector was used.
Column: XBridge ShieldRP18, 5 .mu.m, 50.times.2.1mm; Column
temperature: 40.degree. C.; Solvent system: A=water/conc. NH.sub.3
(aq) (99.95:0.05) and B=acetonitrile; Method: Linear gradient
elution with A:B=95:5 to 0:100 in 3.4 minutes and with a flow rate
of 0.8 mL/min.
[0170] .sup.1H NMR spectra were recorded at 600 MHz on a Bruker
Avance AV-III-600 instrument or at 400 MHz on a Bruker Avance
AV-III-400 instrument or a Varian 400 instrument. Chemical shift
values are expressed in ppm-values relative. The following
abbreviations are used for multiplicity of NMR signals: s=singlet,
d=doublet, t=triplet, q=quartet, dd=double doublet, ddd=double
double doublet, dt=double triplet, br=broad, and m=multiplet.
[0171] Compounds of general formula XIII may be prepared as shown
in Scheme 1.
##STR00024## ##STR00025##
[0172] where R.sup.1 and R.sup.2 are as defined for Formula I,
R.sup.3 is hydrogen or a nitro group and R.sup.4 is as an alkyl
group such as methyl or ethyl
[0173] Compounds of general formula IV (Scheme 1) may be prepared
by reacting compounds of formula II with a sulfinamide such as III
in the presence of a Lewis acid/drying agent such as titanium
tetraethoxide. Treatment of compounds of general formula IV with
compounds of general formula V such as ethyl bromoacetate in the
presence of Zn powder or in the presence of diethyl zinc and
tris(triphenylphosphine)rhodium(I) chloride gives compounds of
general formula VI (Hilpert, H. et al J. Med. Chem. 2013, 56,
3980-3995). Hydrolysis of compounds of general formula VI with an
aqueous base such as sodium hydroxide in water gives compounds of
general formula VII. Activation of the acid group of compounds of
general formula VII with a coupling reagent such as
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) in the presence
of a catalyst such as 4-dimethylaminopyridine (DMAP) followed by
reaction with Meldrum's acid (2,2-dimethyl-1,3-dioxane-4,6-dione)
acid gives compounds of general formula VIII. Reduction of
compounds of general formula VIII with a reducing reagent such as
sodium borohydride in a solvent such as acetic acid gives compounds
of general formula IX. Compounds of general formula IX can be
fluorinated with a fluorination reagent such as
N-fluorodibenzenesulfonamide (NFSi) in the precense of a base such
as 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) to give compounds of
general formula X. Treating compounds of general formula X with an
acid such as hydrochloric acid in methanol will cleave the
sulfinamide bond and transesterification will occur on the two
ester moieties. Treatment of the resulting intermediate with a base
such as potassium carbonate in methanol gives compounds of general
formula XI as a mixture of two diastereomers. The ester moiety of
compounds of general formula XI can be reduced with a reducing
reagent such as sodium borohydride to give compounds of general
formula XII. The mixture of two diastereomers of compounds of
general formula XII can be converted to compounds of general
formula XIII as a mixture of two diastereomers by treatment with
reagents such as nonafluorobutanesulfonyl fluoride (NfF) and a base
such as triethylamine followed by treatment with a reagent such as
tetra-N-butylammonium fluoride (TBAF). The mixture of two
diastereomers of compounds of general formula XIII when R.sup.3 is
a nitro group can be separated by chromatography to give compounds
of general formulae XIVa and XIVb (Scheme 2)
##STR00026##
[0174] where R.sup.1 and R.sup.2 are as defined under formula I and
R.sup.3 is a nitro group.
[0175] Compounds of general formulae XIVa and XIVb may be prepared
as shown in Scheme 3.
##STR00027##
[0176] where R.sup.1 and R.sup.2 are as defined under formula I and
R.sup.3 is hydrogen Treatment of a mixture of two diastereomers of
compounds of general formula XIII with nitric acid in sulfuric acid
and trifluoroacetic acid gives compounds of general formulae XIVa
and XIVb which can be separated by chromatography (Scheme 3).
[0177] Compounds of general formulae XVIa and XVIb may be prepared
as shown in Scheme 4.
##STR00028##
[0178] where R.sup.1 and R.sup.2 are as defined under formula I
[0179] Treatment of compounds of general formula XIVa (Scheme 4)
with a reagent such as Lawesson's reagent
(2,4-bis(4-methoxyphenyl)-1,3,2,4-dithiadiphosphetane-2,4-disulfide)
gives compounds of general formula XVa. Reduction of the nitro
group of compounds of general formula XVa gives compounds of
general formula XVIa. The same procedures can be used to prepare
compounds of general formula XVIb starting from compounds of
general formula XIVb.
[0180] Compounds of general Formula I may be prepared as shown in
Scheme 5.
##STR00029##
[0181] where R.sup.1, R.sup.2 and Ar are as defined under Formula
I.
[0182] Compounds of general formula XIX (Scheme 5) may be prepared
by reacting compounds of general formula XVI with a carboxylic acid
chloride of general formula XVII or by reaction with a carboxylic
acid of general formula XVIII using procedures known to chemists
skilled in the art. Treatment of compounds of general formula XIX
with ammonia gives compounds of general formula I. In some cases,
the addition of and oxidizing reagent such as tert-butyl
hydroperoxide might be necessary to facilitate the reaction.
[0183] Preparation Of Intermediates
INTERMEDIATE:
(R)-N-(1-(2-Fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide
##STR00030##
[0185] To a solution of 1-(2-fluorophenyl)ethanone (25.0 g, 181
mmol) in THF (500 mL) was added Ti(OEt).sub.4 (82.6 g, 362.0 mmol)
and (R)-2-methylpropane-2-sulfinamide (26.3 g, 217 mmol). The
mixture was stirred at 85.degree. C. for 14 hours. The reaction
mixture was quenched with water (200 mL) and then filtered and the
filtrate was extracted with ethyl acetate (3.times.200 mL). The
combined organic layers were washed with brine (100 mL), dried over
Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure.
The residue was combined with four similar batches (25 g reactant
1-(2-fluorophenyl)ethanone for each batch), the mixture was
purified by column chromatography (silica gel, petroleum
ether/ethyl acetate=10/1 to 5:1).
(R)-N-(1-(2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide
(135.0 g, 77.3% yield) was obtained.
[0186]
(R)-N-(1-(2,3-Difluorophenyl)ethylidene)-2-methylpropane-2-sulfinam-
ide was prepared in a similar way from
1-(2,3-difluorophenyl)ethan-1-one
[0187]
(R)-N-(2-Fluoro-1-(2-fluorophenyl)ethylidene)-2-methylpropane-2-sul-
finamide was prepared in a similar way from
2-fluoro-1-(2-fluorophenyl)ethan-1-one
INTERMEDIATE: 2-(2-Fluorophenyl)-2,2-dimethoxyethan-1-ol
##STR00031##
[0189] To a mixture of potassium hydroxide (91.4 g, 1.63 mol) in
methanol (1 L) was added a solution of
1-(2-fluorophenyl)ethan-1-one (50 g, 362 mmol) in methanol (300 mL)
in a dropwise manner at 0.degree. C. Then bis(acetoxy)iodobenzene
(175 g, 543 mmol) was added in portions. After stirring at
0.degree. C. for 4 hours, the reaction was quenched with the
addition of water (500 mL). The mixture was concentrated to remove
methanol and the aqueous phase was extracted with ethyl acetate
(700 mL, three times), the combined organic phases were washed with
brine (300 mL), dried over Na.sub.2SO.sub.4 and concentrated. The
crude product was used in the next step directly without further
purification.
INTERMEDIATE: 1-(2-Fluorophenyl)-2-hydroxyethan-1-one
##STR00032##
[0191] 2-(2-Fluorophenyl)-2,2-dimethoxyethan-1-ol (crude, 362 mmol)
was dissolved in THF (450 mL) and water (150 mL). Then p-toluene
sulfonic acid (125 g, 726 mmol) was added portionwise at room
temperature. After the addition, the mixture was stirred under
reflux for 5 hours. Water (150 mL) and sat. NaHCO.sub.3 was added
to quench the reaction, the mixture was extracted with ethyl
acetate (500 mL, three times). The combined organic layers were
washed with brine and dried over Na.sub.2SO.sub.4. After removal of
the solvents under reduced pressure, the residue was purified by
column chromatography with petroleum ether:ethyl acetate=20:1 to
give 1-(2-fluorophenyl)-2-hydroxyethan-1-one (42 g, 76% yield, two
steps).
INTERMEDIATE: 2-Fluoro-1-(2-fluorophenyl)ethan-1-one
##STR00033##
[0193] To a solution of 1-(2-fluorophenyl)-2-hydroxyethan-1-one (10
g, 64.88 mmol) in dichloromethane (200 mL) was added
Et.sub.3N(HF).sub.3 (10.46 g, 227.8 mmol) and
CF.sub.3(CF.sub.2).sub.3SO.sub.2F (29.4 g, 97.32 mmol) dropwise at
0.degree. C., the solution was stirred at room temperature for 12
hours. TLC (petroleum ether:ethyl acetate=10:1) showed no starting
material. The mixture was poured into a saturated solution of
NaHCO.sub.3 and ice, extracted with dichloromethane (200 mL three
times), the combined organic layers were washed with brine, then
dried over Na.sub.2SO.sub.4 and concentrated under reduced
pressure. The mixture was purified by column chromatography on
silica gel (eluted with petroleum ether:ethyl acetate=1:0-10:1) to
afford 2-fluoro-1-(2-fluorophenyl)ethan-1-one (6 g, yield:
59%).
INTERMEDIATE: Ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2-fluorophenyl)butanoate
##STR00034##
[0195] To a solution of
(R)-N-(1-(2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide
(20.0 g, 82.9 mmol) and Rh(PPh.sub.3).sub.3Cl (2.3 g, 2.5 mmol) in
dry THF (300 mL) was added dropwise Et.sub.2Zn (1 M, 161.6 mL) at
-78.degree. C. The mixture was stirred at -78.degree. C.-0.degree.
C. for 2 hours. The reaction mixture was quenched by NH.sub.4Cl
(sat. aq. 100 mL) at 0.degree. C., and then diluted with water (200
mL) and extracted with ethyl acetate (2.times.200 mL). The combined
organic layers were washed with brine (50 mL) dried over
Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure.
The crude product was purified by flash chromatography on silica
gel with petroleum ether/ethyl acetate=4/1. Ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2-fluorophenyl)butanoate
(23.3 g, 85.3% yield) was obtained as a yellow oil. .sup.1H NMR
(CDCl.sub.3 400MHz): .delta. 7.53-7.49 (m, 1H), 7.27-7.25 (m, 1H),
7.14-7.12 (m, 1H), 7.05-6.99 (m, 1H), 5.15 (s, 1H), 4.02 (q, J. 7.2
Hz, 2H), 3.38 (d, J.16.0 Hz, 1H), 3.10 (d, J.17.6 Hz, 1H), 1.85 (s,
3H), 1.31 (s, 9H), 1.13 (t, J=7.2 Hz, 3H).
[0196] Ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2,3-difluorophenyl)butanoate
was prepared in a similar way from
(R)-N-(1-(2,3-difluorophenyhethylidene)-2-methylpropane-2-sulfinamide
[0197] Ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-4-fluoro-3-(2-fluorophenyl)butanoat-
e was prepared in a similar way from
((R)-N-(2-fluoro-1-(2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinam-
ide
INTERMEDIATE:
(S)-3-(((R)-tert-Butylsulfinyl)amino)-3-(2-fluorophenyl)butanoic
acid
##STR00035##
[0199] To a solution of ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2-fluorophenyl)butanoate
(18.3 g, 55.6 mmol) in THF (180 mL) and H.sub.2O (60 mL) was added
LiOH.H.sub.2O (2.8 g, 66.7 mmol). The mixture was stirred at
18.degree. C. for 18 hours. The reaction mixture was adjusted pH to
3-4 by adding KHSO.sub.4 (sat. aq.). The resulting mixture was
extracted with ethyl acetate (3.times.200 mL). The combined organic
layers were washed with brine (50 mL), dried over Na.sub.2SO.sub.4
and concentrated.
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2-fluorophenyl)butanoic
acid (16.7 g, crude) was obtained and was used into the next step
without further purification.
[0200]
(S)-3-(((R)-tert-Butylsulfinyl)amino)-3-(2,3-difluorophenyl)butanoi-
c acid was prepared in a similar way from ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2,3-difluorophenyl)butanoate
[0201]
(S)-3-(((R)-tert-butylsulfinyl)amino)-4-fluoro-3-(2-fluorophenyl)bu-
tanoic acid was prepared in a similar way from ethyl
(S)-3-(((R)-tert-butylsulfinyl)amino)-4-fluoro-3-(2-fluorophenyl)butanoat-
e
INTERMEDIATE:
(R)--N--((S)-4-(2,2-Dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide
##STR00036##
[0203] To a solution of
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2-fluorophenyl)butanoic
acid (10.0 g, 33.2 mmol) in dry THF (100 mL) was added
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) (7.6 g, 39.8
mmol) and 4-dimethylaminopyridine (DMAP) (4.9 g, 39.8 mmol). The
mixture was stirred at 20.degree. C. for 5 min, then Meldrum's acid
(2,2-dimethyl-1,3-dioxane-4,6-dione) (4.8 g, 33.2 mmol) was added.
The mixture was stirred at 20.degree. C. for 18 hours. Water (150
mL) was added, the resulting mixture was extracted with ethyl
acetate (3.times.150 mL). The combined organic layers were washed
with brine (50 mL), dried over Na.sub.2SO.sub.4 and concentrated.
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide (14.2 g, crude)
was obtained and was used into the next step without further
purification.
[0204]
(R)--N--((S)-2-(2,3-Difluorophenyl)-4-(2,2-dimethyl-4,6-dioxo-1,3-d-
ioxan-5-yl)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide was
prepared in a similar way from
(S)-3-(((R)-tert-butylsulfinyl)amino)-3-(2,3-difluorophenyl)butanoic
acid
[0205]
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-1-fluoro-2--
(2-fluorophenyl)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide was
prepared in a similar way from
(S)-3-(((R)-tert-butylsulfinyl)amino)-4-fluoro-3-(2-fluorophenyl)butanoic
acid
INTERMEDIATE:
(R)--N--((S)-4-(2,2-Dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)butan-2-yl)-2-methylpropane-2-sulfinamide
##STR00037##
[0207] To a solution of
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide (14.2 g, 33.2
mmol) in acetic acid (150 mL) was added NaBH.sub.4 (6.3 g, 165.9
mmol) in 5 portions with 15 minute intervals. Then the mixture was
stirred at 20.degree. C. for 18 hours. NH.sub.4Cl (sat. aq. 100 mL)
was added to quench the reaction, then the mixture was
concentrated. Water (150 mL) was added and the mixture was
extracted with ethyl acetate (3.times.170 mL), the combined organic
layers were washed with brine (100 mL), dried over Na.sub.2SO.sub.4
and concentrated.
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)butan-2-yl)-2-methylpropane-2-sulfinamide (13.7 g, crude) was
obtained and was used into the next step without further
purification.
[0208]
(R)--N--((S)-2-(2,3-Difluorophenyl)-4-(2,2-dimethyl-4,6-dioxo-1,3-d-
ioxan-5-yl)butan-2-yl)-2-methylpropane-2-sulfinamide was prepared
in a similar way from
(R)--N--((S)-2-(2,3-difluorophenyl)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan--
5-yl)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide
[0209]
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-1-fluoro-2--
(2-fluorophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide was
prepared in a similar way from
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-1-fluoro-2-(2-flu-
orophenyl)-4-oxobutan-2-yl)-2-methylpropane-2-sulfinamide
INTERMEDIATE:
(R)--N--((S)-4-(5-Fluoro-2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-flu-
orophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide
##STR00038##
[0211] To a solution of
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-fluorophenyl-
)butan-2-yl)-2-methylpropane-2-sulfinamide (5.0 g, 12.1 mmol) in
anhydrous THF (75 mL) was added N-fluorodibenzenesulfonamide (NFSi)
(4.6 g, 14.5 mmol) and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU)
(1.8 g, 12.1 mmol) at 0.degree. C. The mixture was stirred at
0.degree. C.-20.degree. C. for 1 hour. The mixture was filtered and
the filtrate was concentrated.
(R)--N--((S)-4-(5-fluoro-2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-flu-
orophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide (5.2 g, crude)
was obtained and was used into the next step without further
purification.
[0212]
(R)--N--((S)-2-(2,3-Difluorophenyl)-4-(5-fluoro-2,2-dimethyl-4,6-di-
oxo-1,3-dioxan-5-yl)butan-2-yl)-2-methylpropane-2-sulfinamide was
prepared in a similar way from
(R)--N--((S)-2-(2,3-difluorophenyl)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan--
5-yl)butan-2-yl)-2-methylpropane-2-sulfinamide
[0213]
(R)--N--((S)-1-fluoro-4-(5-fluoro-2,2-dimethyl-4,6-dioxo-1,3-dioxan-
-5-yl)-2-(2-fluorophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide
was prepared in a similar way from
(R)--N--((S)-4-(2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-1-fluoro-2-(2-flu-
orophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide
INTERMEDIATE: Methyl
(6S)-3-fluoro-6-(2-fluorophenyl)-6-methyl-2-oxopiperidine-3-carboxylate
##STR00039##
[0215] To a solution of
(R)--N--((S)-4-(5-fluoro-2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-2-(2-flu-
orophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide (5.2 g, 12.1
mmol) in MeOH (50 mL) was added HCl/MeOH (4 M, 30.3 mL). The
mixture was stirred at 20.degree. C. for 18 hours. The mixture was
concentrated and redissolved in dry MeOH (60 mL). Triethylamine was
added (2.9 g, 29.1 mmol). The mixture was stirred at 80.degree. C.
for 18 hours. The mixture was concentrated. Then anhydrous THF (100
mL) was added and the mixture was stirred at 20.degree. C. for 5
min. The mixture was filtered and the residue was washed with
anhydrous THF (2.times.30 mL) and the combined filtrates were
concentrated. Methyl
(6S)-3-fluoro-6-(2-fluorophenyl)-6-methyl-2-oxopiperidine-3-carboxylate
(3.4 g, crude) was obtained and was used into the next step without
further purification.
[0216] Methyl
(6S)-6-(2,3-difluorophenyl)-3-fluoro-6-methyl-2-oxopiperidine-3-carboxyla-
te was prepared in a similar way from
(R)--N--((S)-2-(2,3-difluorophenyl)-4-(5-fluoro-2,2-dimethyl-4,6-dioxo-1,-
3-dioxan-5-yl)butan-2-yl)-2-methylpropane-2-sulfinamide
[0217] Methyl
(6S)-3-fluoro-6-(fluoromethyl)-6-(2-fluorophenyl)-2-oxopiperidine-3-carbo-
xylate was prepared in a similar way from
(R)--N--((S)-1-fluoro-4-(5-fluoro-2,2-dimethyl-4,6-dioxo-1,3-dioxan-5-yl)-
-2-(2-fluorophenyl)butan-2-yl)-2-methylpropane-2-sulfinamide
INTERMEDIATE:
(6S)-3-Fluoro-6-(2-fluorophenyl)-3-(hydroxymethyl)-6-methylpiperidin-2-on-
e
##STR00040##
[0219] To a solution of methyl
(6S)-3-fluoro-6-(2-fluorophenyl)-6-methyl-2-oxopiperidine-3-carboxylate
(3.4 g, 12.1 mmol) in anhydrous MeOH (50 mL) was added NaBH.sub.4
(3.7 g, 96.9 mmol) in 5 portions with 15 minute intervals. The
mixture was stirred at 20.degree. C. for 17 hours. The mixture was
concentrated, then water (200 mL) was added and the mixture was
extracted with ethyl acetate (3.times.150 mL). The combined organic
layers were washed with brine (50 mL), dried over Na.sub.2SO.sub.4
and concentrated. The crude product was purified by flash
chromatography on silica gel with petroleum ether:ethyl
acetate=1:1.about.1:2 to give
(6S)-3-fluoro-6-(2-fluorophenyl)-3-(hydroxymethyl)-6-methylpiperidin-2-on-
e (2.5 g, 80.9% yield).
[0220]
(6S)-6-(2,3-Difluorophenyl)-3-fluoro-3-(hydroxymethyl)-6-methylpipe-
ridin-2-one was prepared in a similar way from methyl
(6S)-6-(2,3-difluorophenyl)-3-fluoro-6-methyl-2-oxopiperidine-3-carboxyla-
te
[0221]
(6S)-3-Fluoro-6-(fluoromethyl)-6-(2-fluorophenyl)-3-(hydroxymethyl)-
piperidin-2-one was prepared in a similar way from methyl
(6S)-3-fluoro-6-(fluoromethyl)-6-(2-fluorophenyl)-2-oxopiperidine-3-carbo-
xylate
INTERMEDIATE:
(6S)-3-Fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-2-one
##STR00041##
[0223] To a solution of
(6S)-3-fluoro-6-(2-fluorophenyl)-3-(hydroxymethyl)-6-methylpiperidin-2-on-
e (2.8 g, 11.0 mmol) in anhydrous THF (60 mL) was added
nonafluorobutanesulfonyl fluoride (NfF) (9.9 g, 32.9 mmol) and
triethylamine (4.4 g, 43.9 mmol). The mixture was stirred at
20.degree. C. for 2.5 hours. A solution of tetra-N-butylammonium
fluoride (TBAF) (1 M, 13.2 mL) in THF was added. The mixture was
stirred at 50.degree. C. for 16 h. Water (150 mL) was added. The
mixture was extracted with ethyl acetate (3.times.100 mL). The
combined organic phase was washed with brine (30 mL), dried over
MgSO.sub.4 and concentrated in vacuo. The crude material was
purified via flash chromatography on silica gel with petroleum
ether:ethyl acetate=2:1 to give
(6S)-3-fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-2-one
(2.4 g, 85.1% yield).
[0224]
(6S)-6-(2,3-Difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiper-
idin-2-one was prepared in a similar way from
(6S)-6-(2,3-difluorophenyl)-3-fluoro-3-(hydroxymethyl)-6-methylpiperidin--
2-one
[0225]
(6S)-3-Fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-2-o-
ne was prepared in a similar way from
(6S)-3-fluoro-6-(fluoromethyl)-6-(2-fluorophenyl)-3-(hydroxymethyl)piperi-
din-2-one
INTERMEDIATES:
(3S,6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one and
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one
##STR00042##
[0227]
(6S)-3-Fluoro-3-(fluoromethyl)-6-(2-fluorophenyl)-6-methylpiperidin-
-2-one (1.4 g, 5.4 mmol) was suspended in TFA (9 mL). The mixture
was cooled to 0.degree. C. and concentrated H.sub.2SO.sub.4 (4.2 g,
41.9 mmol) (98%) was added. Finally, HNO.sub.3 (1.7 g, 16.3 mmol)
(60%) was added dropwise. After addition, the black brown mixture
was stirring at 0.degree. C. for 2 h. The mixture was poured onto
100 g ice and basified to pH>11 using 5 M NaOH (aq.). The
suspension was extracted with ethyl acetate (150 mL). The phases
were separated and the aqueous layer was extracted with ethyl
acetate (2.times.100 mL). The combined organics were washed with a
solution of saturated aqueous NH.sub.4Cl (50 mL) and water (50 mL),
dried over MgSO.sub.4, filtered, and concentrated under reduced
pressure. The residue was purified by column chromatography (silica
gel, petroleum ether/ethyl acetate=2/1) to give
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one and
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one.
(3S,6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one (900 mg) .sup.1H NMR (CDCl.sub.3 400MHz): .delta.
8.42-8.40 (m, 1H), 8.26-8.22 (m, 1H), 7.56 (brs, 1H), 7.31-7.26 (m,
1H), 4.78 (d, J=19.6 Hz, 1H), 4.67 (d, J=19.6 Hz, 1H), 2.61-2.57
(m, 1H), 2.38-2.30 (m, 1H), 2.20-2.19 (m, 1H), 2.19-2.16 (m, 1H),
1.78 (s, 3H).
[0228]
(3R,6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-meth-
ylpiperidin-2-one (600 mg).sup.1H NMR (CDCl.sub.3 400 MHz): .delta.
8.29-8.22 (m, 2H), 7.72 (brs, 1H), 7.31-7.26 (m, 1H), 4.72-4.47 (m,
2H), 2.62-2.58 (m, 1H), 2.30-2.25 (m, 2H), 2.19-1.80 (s, 3H),
1.80-1.66 (m, 1H).
[0229]
(3S,6S)-6-(2,3-Difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidin-2-one and
(3R,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidin-2-one were prepared in a similar way from
(6S)-6-(2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpiperidin-2-
-one
[0230]
(6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piper-
idin-2-one was prepared in a similar way from
(6S)-3-fluoro-3,6-bis(fluoromethyl)-6-(2-fluorophenyl)piperidin-2-one
INTERMEDIATE:
(3R,6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione
##STR00043##
[0232] To a solution of
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one (900 mg, 3.0 mmol) in anhydrous toluene (18 mL) was
added
2,4-bis(4-methoxyphenyl)-1,3,2,4-dithiadiphosphetane-2,4-dithione
(Lawesson's reagent) (723 mg, 1.8 mmol). The mixture was stirred at
80.degree. C. for 2 hours. The mixture was concentrated. The crude
product was purified by flash chromatography with petroleum
ether:ethyl acetate=3:1 to give
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione (900 mg, 94.9% yield).
[0233]
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-meth-
ylpiperidine-2-thione was prepared in a similar way from
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridin-2-one
[0234]
(3R,6S)-6-(2,3-Difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione was prepared in a similar way from
(3S,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidin-2-one
[0235]
(3S,6S)-6-(2,3-Difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione was prepared in a similar way from
(3R,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidin-2-one
[0236]
(6S)-3-Fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piper-
idine-2-thione was prepared in a similar way from
(6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piperidin-2-
-one
INTERMEDIATE:
(3R,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione
##STR00044##
[0238] To a solution of
(3R,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione (900 mg, 2.8 mmol) in EtOH (18.0 mL) and H.sub.2O
(4.5 mL) was added Fe (790 mg, 14.15 mmol) and NH.sub.4Cl (756 mg,
14.2 mmol).The mixture was stirred at 20.degree. C. for 5 hours.
The mixture was filtered and the residue was washed with EtOH (30
mL). The combined filtrates were concentrated. The residue was
dispersed in ethyl acetate (30 mL) and then filtered. The filter
cake was washed with ethyl acetate (2.times.15 mL). The combined
organic layers were concentrated. The crude product was purified by
flash chromatography with petroleum ether:ethyl acetate=3:1 to give
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione (712 mg, 87% yield). .sup.1H NMR (600 MHz,
CDCl.sub.3) .delta. 8.42 (s, 1H), 6.88 (dd, J=11.8, 8.6 Hz, 1H),
6.58 (dt, J=7.0, 3.2 Hz, 1H), 6.50 (dd, J=6.8, 2.8 Hz, 1H), 4.95
(ddd, J=49.0, 12.6, 10.9 Hz, 1H), 4.61 (ddd, J=46.7, 28.4, 10.7 Hz,
1H), 3.70 (s, 2H), 2.62-2.54 (m, 1H), 2.39-2.30 (m, 1H), 2.10-2.02
(m, 1H), 1.91 (q, J=14.0 Hz, 1H), 1.70 (s, 3H). [.alpha.].sup.20,
D=-236.degree. (c=0.10, EtOH).
[0239]
(3S,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-meth-
ylpiperidine-2-thione was prepared in a similar way from
(3S,6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione. .sup.1H NMR (600 MHz, CDCl.sub.3) .delta. 8.58 (s,
1H), 6.88 (dd, J=11.7, 8.6 Hz, 1H), 6.58 (dt, J=8.4, 3.2 Hz, 1H),
6.38 (dd, J=6.7, 2.7 Hz, 1H), 4.97 (dt, J=48.8, 10.6 Hz, 1H), 4.60
(ddd, J=46.0, 24.6, 10.3 Hz, 1H), 3.68 (s, 2H), 2.53 (d, J=14.1 Hz,
1H), 2.26-2.17 (m, 1H), 2.13 (td, J=13.9, 2.6 Hz, 1H), 1.87-1.74
(m, 1H), 1.73 (s, 3H). [.alpha.].sup.20, D=-154.degree. (c=0.10,
EtOH).
[0240]
(3R,6S)-6-(5-Amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione was prepared in a similar way from
(3R,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione
[0241]
(3S,6S)-6-(5-Amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6--
methylpiperidine-2-thione was prepared in a similar way from
(3S,6S)-6-(2,3-difluoro-5-nitrophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione
[0242]
(3R,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)pi-
peridine-2-thione and
(3S,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione were prepared in a similar way starting from
(6S)-3-fluoro-6-(2-fluoro-5-nitrophenyl)-3,6-bis(fluoromethyl)piperidine--
2-thione followed by chromatographic separation of the two
diastereomers.
[0243]
(3R,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)pi-
peridine-2-thione: .sup.1H NMR (CDCl.sub.3 400 MHz) .delta. 8.42
(s, 1H), 6.95-6.89 (m, 1H), 6.65-6.62 (m, 1H), 6.54 (dd, J=6.8, 3.2
Hz, 1H), 5.03-4.51 (m, 4H), 3.73 (s, 2H), 2.48-2.39 (m, 2H), 2.22
(t, J=14.0 Hz, 1H), 1.96 (q, J=12.8 Hz, 1H). [.alpha.].sup.20,
D=-204.degree. (c=0.10, EtOH).
(3S,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)p-
iperidine-2-thione: .sup.1H NMR (CDCl.sub.3 400 MHz) .delta. 8.65
(s, 1H), 6.95-6.90 (m, 1H), 6.64-6.62 (m, 1H), 6.41 (dd, J=6.8, 2.8
Hz, 1H), 5.09-4.95 (m, 2H), 4.89-4.52 (m, 2H), 2.41-2.38 (m, 1H),
2.33-2.24 (m, 1H), 2.21-2.14 (m, 1H), 1.92-1.75 (m, 1H).
[.alpha.].sup.20, D=-130.degree. (c=0.10, EtOH).
INTERMEDIATE: Methyl 5-(methoxy-d.sub.3)picolinate
##STR00045##
[0245] Methyl 5-hydroxypicolinate (2.88 g, 18.8 mmol) was dissolved
in dimethylformamide (108 ml) under argon. Potassium carbonate
(7.20 g, 52.1 mmol) was added and the orange suspension was stirred
for 45 minutes at room temperature. lodomethane-d.sub.3 (1.41 ml,
22.6 mmol) was added. The reaction mixture was stirred for 2 hours.
Water was added. The mixture was extracted with ethyl acetate. The
organic phase was washed with brine, dried over MgSO.sub.4 and
concentrated in vacuo and purified by column chromatography on
silica gel (heptane: ethyl acetate) to give methyl
5-(methoxy-d.sub.3)picolinate.
INTERMEDIATE: 5-(Methoxy-d.sub.3)picolinic acid
##STR00046##
[0247] Methyl 5-(methoxy-d.sub.3)picolinate (200 mg, 1.175 mmol)
was dissolved in water (1.5 ml) and 1,4-dioxane (3 ml). Lithium
hydroxide (70.4 mg, 2.94 mmol) was added and the reaction mixture
was stirred for 1 hour. The reaction mixture was evaporated to
about 2 ml and extracted with diethylether. The organic phase was
extracted with 1M NaOH and the combined aqueous phases were
acidified to pH 2 with 6N HCl (aq).
[0248] The mixture was cooled on an icebath and a precipitate was
formed. The precipitate was collected to give
5-(methoxy-d.sub.3)picolinic acid.
[0249] Stereochemistry
[0250] The relative stereochemistry of intermediate
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione was assigned by 2D ROESY (rotating frame nuclear
Overhauser effect spectroscopy) (FIG. 1). nOe (nuclear Overhauser
effect) signals were observed between H(A) (.delta. 4.61) and H(B)
(.delta. 2.10-2.02) and between H(B) (.delta. 2.10-2.02) and H(D)
(.delta. 1.70) and an nOe signal was also observed between H(C)
(.delta. 4.61) and H(E) (.delta. 6.58). Thus, the relative
stereochemistry of intermediate
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione is confirmed since no significant nOe's was
observed between H(A) (.delta. 4.60) and H(B) (.delta. 2.53 or
2.13) in the 2D ROESY of
(3S,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-m-
ethylpiperidine-2-thione. The relative stereochemistries of
(3R,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione,
(3S,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione,
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and
(3S,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione were assigned by analogy.
##STR00047##
[0251] FIG. 1. nOe in ROESY of
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione
[0252] Preparation of Compounds of Formula I
EXAMPLE 1
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-methoxypicolinamide (Compound
1)
##STR00048##
[0254] HATU
(1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate) (600 mg, 1.58 mmol) was added to
5-methoxypicolinic acid (242 mg, 1.58 mmol) in DMF (1 ml). The
reaction mixture was stirred at room temperature for 5 minutes.
(3R,6S)-6-(5-Amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione (350 mg, 1.21 mmol) was added followed by DIPEA
(N,N-diisopropylethylamine) (1.06 ml, 6.07 mmol) and the reaction
mixture was stirred at room temperature for 1 hour. Saturated
ammonium chloride (aq) was added. The mixture was extracted with
ethyl acetate. The organic phase was washed with brine, dried over
magnesium sulfate and concentrated in vacuo. 7M ammonia in methanol
(3 mL, 21 mmol) was added and the reaction mixture was stirred in a
sealed vial at 50.degree. C. overnight. The reaction mixture was
allowed to cool to room temperature and was concentrated in vacuo.
The crude product was purified by flash chromatography on silica
gel (eluent: heptane/ethyl acetate). The product was further
purified by the following procedure: The product was dissolved in
ethyl acetate (50 mL) and washed with a solution of saturated
aqueous NaHCO.sub.3/water (1/1). The organic phase was washed total
of 10 times (using 10 mL each time). The organic phase was dried
over MgSO.sub.4, filtered, and evaporated to give
N-(3-((2S,5S)-6-amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahyd-
ropyridin-2-yl)-4-fluorophenyl)-5-methoxypicolinamide. .sup.1H NMR
(600 MHz, DMSO) .delta. 10.38 (s, 1H), 8.39 (d, J=2.6 Hz, 1H), 8.12
(d, J=8.6 Hz, 1H), 7.84-7.76 (m, 2H), 7.62 (dd, J=8.7, 2.9 Hz, 1H),
7.13 (dd, J=11.9, 8.6 Hz, 1H), 6.10 (s, 2H), 4.94-4.68 (m, 2H),
3.93 (s, 3H), 2.16-2.03 (m, 2H), 1.97-1.87 (m, 1H), 1.64-1.53 (m,
1H), 1.46 (s, 3H). LC-MS (m/z) 407.1 (MH.sup.+); t.sub.R=0.51
(Method B)
EXAMPLE 2
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-(methoxy-d.sub.3)picolinamide
(Compound 2)
##STR00049##
[0256] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-(methoxy-d.sub.3)picolinic acid .sup.1H NMR
(600 MHz, DMSO) .delta. 10.35 (s, 1H), 8.39 (dd, J=2.9, 0.5 Hz,
1H), 8.12 (dd, J=8.7, 0.5 Hz, 1H), 7.84-7.77 (m, 2H), 7.61 (dd,
J=8.7, 2.9 Hz, 1H), 7.12 (dd, J=11.9, 8.6 Hz, 1H), 6.24 (s, 2H),
4.96-4.74 (m, 2H), 2.16-2.05 (m, 2H), 1.99-1.89 (m, 1H), 1.66-1.55
(m, 1H), 1.47 (s, 3H). LC-MS (m/z) 410.2 (MH.sup.+); t.sub.R=0.5
(Method B)
EXAMPLE 3
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-cyano-3-methylpicolinamide
(Compound 3)
##STR00050##
[0258] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-cyano-3-methylpicolinic acid .sup.1H NMR (600
MHz, DMSO) .delta. 10.72 (s, 1H), 8.98 (s, 1H), 8.40 (s, 1H),
7.84-7.79 (m, 1H), 7.67 (dd, J=7.4, 2.6 Hz, 1H), 7.19-7.13 (m, 1H),
4.97-4.72 (m, 2H), 2.51 (s, 3H), 2.17-2.09 (m, 2H), 1.98-1.89 (m,
1H), 1.62-1.52 (m, 1H), 1.49 (s, 3H). LC-MS (m/z) 416.1 (MH.sup.+);
t.sub.R=0.5 (Method A)
EXAMPLE 4
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-chloropicolinamide (Compound
4)
##STR00051##
[0260] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-chloropicolinic acid .sup.1H NMR (600 MHz,
DMSO) .delta. 10.61 (s, 1H), 8.78 (d, J=1.9 Hz, 1H), 8.20 (dd,
J=8.4, 2.2 Hz, 1H), 8.15 (d, J=8.4 Hz, 1H), 7.86-7.82 (m, 1H),
7.82-7.78 (m, 1H), 7.14 (dd, J=11.7, 8.8 Hz, 1H), 6.17 (s, 2H),
4.97-4.66 (m, 2H), 2.16-2.05 (m, 2H), 1.97-1.89 (m, 1H), 1.65-1.55
(m, 1H), 1.47 (s, 3H). LC-MS (m/z) 411.1 (MH.sup.+); t.sub.R=0.53
(Method A)
EXAMPLE 5
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-methoxypyrazine-2-carboxamide
(Compound 5)
##STR00052##
[0262] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-methoxypyrazine-2-carboxylic acid .sup.1H NMR
(600 MHz, DMSO) .delta. 10.45 (s, 1H), 8.88 (d, J=1.0 Hz, 1H), 8.41
(d, J=1.0 Hz, 1H), 7.83 (dd, J=7.3, 2.4 Hz, 1H), 7.78-7.74 (m, 1H),
7.13 (dd, J=11.8, 8.8 Hz, 1H), 6.12 (s, 2H), 4.96-4.68 (m, 2H),
4.02 (s, 3H), 2.26-2.04 (m, 2H), 1.96-1.89 (m, 1H), 1.64-1.53 (m,
1H), 1.46 (s, 3H). LC-MS (m/z) 408.1 (MH.sup.+); t.sub.R=0.48
(Method A)
EXAMPLE 6
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-2-methyloxazole-5-carboxamide
(Compound 6)
##STR00053##
[0264] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 2-methyloxazole-4-carboxylic acid .sup.1H NMR
(600 MHz, DMSO) .delta. 10.09 (s, 1H), 8.62 (s, 1H), 7.76 (dd,
J=7.5, 2.7 Hz, 1H), 7.70-7.65 (m, 1H), 7.10 (dd, J=11.9, 8.7 Hz,
1H), 6.08 (s, 2H), 4.94-4.68 (m, 2H), 2.50 (s, 3H), 2.14-2.04 (m,
2H), 1.95-1.87 (m, 1H), 1.60-1.52 (m, 1H), 1.46 (s, 3H). LC-MS
(m/z) 381.1 (MH.sup.+); t.sub.R=0.43 (Method A)
EXAMPLE 7
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-4-bromo-1-methyl-1H-imidazole-2-carboxamide
(Compound 7)
##STR00054##
[0266] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 4-bromo-1-methyl-1H-imidazole-2-carboxylic acid
.sup.1H NMR (600 MHz, DMSO) .delta. 10.40 (s, 1H), 7.77 (dd,
J=14.3, 9.2 Hz, 1H), 7.70-7.65 (m, 1H), 7.63 (s, 1H), 7.10 (dd,
J=11.8, 8.8 Hz, 1H), 6.10 (s, 2H), 4.95-4.68 (m, 2H), 3.96 (s, 3H),
2.15-2.02 (m, 2H), 1.95-1.85 (m, 1H), 1.64-1.53 (m, 1H), 1.45 (s,
3H). LC-MS (m/z) 460 (MH.sup.+); t.sub.R=0.51 (Method A)
EXAMPLE 8
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-4-methylthiazole-2-carboxamide
(Compound 8)
##STR00055##
[0268] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 4-methylthiazole-2-carboxylic acid .sup.1H NMR
(600 MHz, DMSO) .delta. 10.72 (s, 1H), 7.87 (dd, J=7.4, 2.5 Hz,
1H), 7.73-7.67 (m, 2H), 7.12 (dd, J=11.8, 8.8 Hz, 1H), 6.12 (s,
2H), 4.97-4.68 (m, 2H), 2.50 (s, s 3H), 2.15-2.04 (m, 2H),
1.95-1.87 (m, 1H), 1.63-1.52 (m, 1H), 1.46 (s, 3H). LC-MS (m/z)
397.1 (MH.sup.+); t.sub.R=0.49 (Method A)
EXAMPLE 9
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-2-(difluoromethypoxazole-4-carboxamide
(Compound 9)
##STR00056##
[0270] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 2-(difluoromethyl)oxazole-4-carboxylic acid
.sup.1H NMR (600 MHz, DMSO) .delta. 10.40 (s, 1H), 9.04-8.99 (m,
1H), 7.75 (dd, J=11.8, 6.7 Hz, 1H), 7.71-7.64 (m, 1H), 7.34 (t,
J=51.9 Hz, 1H), 7.12 (dd, J=11.7, 8.9 Hz, 1H), 6.10 (s, 2H),
4.96-4.68 (m, 2H), 2.16-2.03 (m, 2H), 1.96-1.86 (m, 1H), 1.62-1.52
(m, 1H), 1.46 (s, 3H). LC-MS (m/z) 417.1 (MH.sup.+); t.sub.R=0.46
(Method A)
EXAMPLE 10
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-1-(difluoromethyl)-1H-imidazole-4-carboxami-
de (Compound 10)
##STR00057##
[0272] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 1-(difluoromethyl)-1H-pyrazole-3-carboxylic
acid .sup.1H NMR (600 MHz, DMSO) .delta. 10.36 (s, 1H), 8.41 (d,
J=2.5 Hz, 1H), 7.92 (t, J=58.7 Hz, 1H), 7.76 (dd, J=7.1, 2.1 Hz,
1H), 7.71-7.66 (m, 1H), 7.12 (dd, J=11.7, 8.9 Hz, 1H), 7.01 (d,
J=2.5 Hz, 1H), 6.08 (s, 2H), 4.96-4.69 (m, 2H), 2.15-2.03 (m, 2H),
1.97-1.87 (m, 1H), 1.64-1.53 (m, 1H), 1.46 (s, 3H). LC-MS (m/z)
416.1 (MH.sup.+); t.sub.R=0.46 (Method A)
EXAMPLE 11
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-(difluoromethyl)pyrazine-2-carboxamide
(Compound 11)
##STR00058##
[0274] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-(difluoromethyl)pyrazine-2-carboxylic acid
.sup.1H NMR (600 MHz, DMSO) .delta. 10.88 (s, 1H), 9.38 (s, 1H),
9.09 (s, 1H), 7.89 (dd, J=7.3, 2.4 Hz, 1H), 7.83-7.77 (m, 1H), 7.27
(t, J=51.0 Hz, 2H), 7.18-7.13 (m, 1H), 6.16 (s, 2H), 4.96-4.69 (m,
2H), 2.15-2.07 (m, 2H), 1.97-1.88 (m, 1H), 1.62-1.54 (m, 1H), 1.48
(s, 3H). LC-MS (m/z) 428.1 (MH.sup.+); t.sub.R=0.48 (Method A)
EXAMPLE 12
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-4-chlorobenzamide (Compound 12)
##STR00059##
[0276] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 4-chlorobenzoic acid .sup.1H NMR (600 MHz,
DMSO) .delta. 10.42 (s, 1H), 7.98 (d, J=8.3 Hz, 2H), 7.70 (d, J=5.7
Hz, 2H), 7.60 (d, J=8.4 Hz, 2H), 7.13 (dd, J=11.5, 9.1 Hz, 1H),
6.09 (s, 2H), 4.96-4.68 (m, 2H), 2.17-2.05 (m, 2H), 1.98-1.89 (m,
1H), 1.64-1.52 (m, 1H), 1.46 (s, 3H). LC-MS (m/z) 410.1 (MH.sup.+);
t.sub.R=0.54 (Method A)
EXAMPLE 13
N-(3-((2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-fluoropicolinamide (Compound
13)
##STR00060##
[0278] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-fluoropicolinic acid .sup.1H NMR (600 MHz,
DMSO) .delta. 11.11 (s, 1H), 10.92 (s, 1H), 9.88 (s, 1H), 9.73 (s,
1H), 8.75 (d, J=2.2 Hz, 1H), 8.24 (dd, J=8.7, 4.6 Hz, 1H),
8.05-8.01 (m, 1H), 7.99 (dd, J=8.7, 2.8 Hz, 1H), 7.91 (dd, J=7.6,
2.4 Hz, 1H), 7.30 (dd, J=12.1, 8.9 Hz, 1H), 5.19-4.94 (m, 2H), 2.39
(d, J=12.8 Hz, 1H), 2.36-2.29 (m, 1H), 2.15 (t, J=13.8 Hz, 1H),
1.79-1.71 (m, 4H). LC-MS (m/z) 395.1 (MH.sup.+); t.sub.R=0.49
(Method A)
EXAMPLE 14
N-(3-((2S,5R)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-2,3,4,5-tetrahydr-
opyridin-2-yl)-4-fluorophenyl)-5-methoxypicolinamide (Compound
14)
##STR00061##
[0280] Prepared from
(3S,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methylpipe-
ridine-2-thione and 5-methoxypicolinic acid LC-MS (m/z) 407
(MH.sup.+); t.sub.R=0.5 (Method B)
EXAMPLE 15
N-[3-[(2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-fluoro-pyridine-2-carboxamide
(Compound 15)
##STR00062##
[0282] Prepared from
(3R,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-fluoropicolinic acid .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 9.83 (s, 1H), 8.45 (d, J=2.8 Hz, 1H),
8.32 (q, J=4.4 Hz, 1H), 8.11-8.06 (m, 1H), 7.60 (td, J=8.0, 2.8 Hz,
1H), 7.17 (dt, J=5.6, 2.4 Hz, 1H), 4.80-4.55 (m, 2H), 3.49 (brs,
2H), 2.50-2.44 (m, 1H), 2.19-2.12 (m, 1H), 1.93 (td, J=14.0, 2.0
Hz, 1H), 1.80-1.68 (m, 1H), 1.63 (s, 3H). LC-MS (m/z) 413.1
(MH.sup.+); t.sub.R=2.07 min (Method C) [.alpha.].sup..degree.,
D=-10.00 (589 nm, c=0.10, MeOH).
EXAMPLE 16
N-[3-[(2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-methoxy-pyridine-2-carboxamide
(Compound 16)
##STR00063##
[0284] Prepared from
(3R,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-methoxypicolinic acid .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 9.86 (s, 1H), 8.26 (d, J=2.8 Hz, 1H),
8.22 (d, J=8.4 Hz, 1H), 8.14-8.08 (m, 1H), 7.34 (dd, J=8.8, 3.2 Hz,
1H), 7.15 (dt, J=6.0, 2.0 Hz, 1H), 4.80-4.55 (m, 2H), 3.95 (s, 3H),
3.38 (brs, 2H), 2.50-2.44 (m, 1H), 2.20-2.13 (m, 1H), 1.93 (td,
J=13.8, 2.0 Hz, 1H), 1.80-1.68 (m, 1H), 1.62 (d, J=1.2 Hz, 3H).
LC-MS (m/z) 425.1 (MH.sup.+); t.sub.R=2.10 min (Method C)
[.alpha.].sup.20, D=+13.00 (589 nm, c=0.10, MeOH).
EXAMPLE 17
N-[3-[(2S,5S)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-methoxy-pyrazine-2-carboxamide
(Compound 17)
##STR00064##
[0286] Prepared from
(3R,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-methoxypyrazine-2-carboxylic acid .sup.1H
NMR (400 MHz, CDCl.sub.3): .delta. 9.54 (s, 1H), 9.00 (d, J=1.6 Hz,
1H), 9.15 (d, J=1.6 Hz, 1H), 8.13-8.07 (m, 1H), 7.14 (dt, J=6.0,
2.0 Hz, 1H), 4.81-4.56 (m, 2H), 4.08 (s, 3H), 3.05 (brs, 2H),
2.53-2.46 (m, 1H), 2.20-2.13 (m, 1H), 1.94 (td, J=13.6, 2.4 Hz,
1H), 1.82-1.70 (m, 1H), 1.64 (d, J=1.2 Hz, 3H). LC-MS (m/z) 426.1
(MH.sup.+); t.sub.R=2.07 (Method C) [.alpha.].sup.20, D=+12.00 (589
nm, c=0.10, MeOH).
EXAMPLE 18
N-[3-[(2S,5R)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-fluoro-pyridine-2-carboxamide
(Compound 18)
##STR00065##
[0288] Prepared from
(3S,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-fluoropicolinic acid .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 9.80 (s, 1H), 8.46 (d, J=2.8 Hz, 1H),
8.32 (q, J=4.4 Hz, 1H), 7.98-7.92 (m, 1H), 7.61 (td, J=8.0, 2.8 Hz,
1H), 7.19(dt, J=6.0, 2.4 Hz, 1H), 4.69-4.41 (m, 2H), 3.29 (brs,
2H), 2.19 (d, J=5.6 Hz, 2H), 2.14-2.06 (m, 1H), 1.95-1.84 (m, 1H),
1.66 (s, 3H). LC-MS (m/z) 413.1 (MH.sup.+); t.sub.R=2.07 (Method C)
[.alpha.].sup.20, D=+9.00 (589 nm, c=0.10, MeOH).
EXAMPLE 19
N-[3-[(2S,5R)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-methoxy-pyridine-2-carboxamide
(Compound 19)
##STR00066##
[0290] Prepared from
(3S,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-methoxypicolinic acid .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 9.83 (s, 1H), 8.26 (d, J=2.8 Hz, 1H),
8.22 (d, J=8.4 Hz, 1H), 7.99-7.94 (m, 1H), 7.34 (dd, J=8.4, 2.4 Hz,
1H), 7.17 (dt, J=5.6, 2.0 Hz, 1H), 4.69-4.41 (m, 2H), 3.95 (s, 3H),
3.49 (brs, 2H), 2.18 (d, J=6.0 Hz, 2H), 2.13-2.05 (m, 1H),
1.94-1.83 (m, 1H), 1.65 (s, 3H). LC-MS (m/z) 425.1 (MH.sup.+);
t.sub.R=2.10 (Method C) [.alpha.].sup.20, D=+11.00 (589 nm, c=0.10,
MeOH).
EXAMPLE 20
N-[3-[(2S,5R)-6-Amino-5-fluoro-5-(fluoromethyl)-2-methyl-3,4-dihydropyridi-
n-2-yl]-4,5-difluorophenyl]-5-methoxy-pyrazine-2-carboxamide
(Compound 20)
##STR00067##
[0292] Prepared from
(3S,6S)-6-(5-amino-2,3-difluorophenyl)-3-fluoro-3-(fluoromethyl)-6-methyl-
piperidine-2-thione and 5-methoxypyrazine-2-carboxylic acid .sup.1H
NMR (400 MHz, CDCl.sub.3): .delta. 9.51 (s, 1H), 9.00 (d, J=1.2 Hz,
1H), 8.16 (d, J=1.2 Hz, 1H), 7.98-7.93 (m, 1H), 7.16 (dt, J=5.6,
2.4 Hz, 1H), 4.70-4.43 (m, 2H), 4.08 (s, 3H), 2.80 (brs, 2H), 2.20
(dd, J=8.4, 4.0 Hz, 2H), 2.15-2.09 (m, 1H), 1.94-1.82 (m, 1H), 1.68
(s, 3H). LC-MS (m/z) 426.1 (MH.sup.+); t.sub.R=2.07 (Method C)
[.alpha.].sup.20, D=-9.00 (589 nm, c=0.10, MeOH).
EXAMPLE 21
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2--
yl]-4-fluorophenyl]-5-fluoro-pyridine-2-carboxamide (Compound
21)
##STR00068##
[0294] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-fluoropicolinic acid .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 9.85 (s, 1H), 8.48-8.47 (m, 1H), 8.34 (dd,
J=8.8, 4.4 Hz, 1H), 8.02-7.98 (m, 1H), 7.63-7.58 (m, 2H), 7.10 (dd,
J=11.6, 8.8 Hz, 1H), 4.87-4.82 (m, 1H), 4.77-4.64 (m, 2H), 4.49
(dd, J=48.0, 8.0 Hz 1H), 2.22-2.38 (m, 1H), 2.26-2.19 (m, 2H), 1.77
(br, s, 1H). LC-MS (m/z) 413.1 (MH.sup.+); t.sub.R=1.92 (Method
C)
EXAMPLE 22
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2--
yl]-4-fluorophenyl]-5-methoxy-pyridine-2-carboxamide (Compound
22)
##STR00069##
[0296] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-methoxypicolinic acid .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 9.87 (s, 1H), 8.28-8.23 (m, 2H), 8.01-7.99 (m,
1H), 7.59 (d, J=4.0 Hz, 1H), 7.09 (dd, J=11.8, 8.8 Hz 1H),
4.87-4.82 (m, 1H), 4.76-4.64 (m, 2H), 4.48 (dd, J=47.6, 8.8 Hz,
1H), 3.91 (t, J=9.6 Hz, 2H), 3.95 (s, 3H), 2.43-2.37 (m, 1H),
2.26-2.18 (m, 1H), 1.84-1.77 (m, 1H). LC-MS (m/z) 425.1 (MH.sup.+);
t.sub.R=1.95 (Method C)
EXAMPLE 23
N-[3-[(2S,5S)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2--
yl]-4-fluorophenyl]-5-methoxy-pyrazine-2-carboxamide (Compound
23)
##STR00070##
[0298] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-methoxypyrazine-2-carboxylic acid .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 9.46 (s, 1H), 8.94 (s, 1H), 8.08 (s,
1H), 7.93-7.89 (m, 1H), 7.50 (dd, J=7.2, 2.8Hz, 1 Hz), 7.02 (dd,
J=11.6, 8.8Hz, 1H), 4.78-4.74 (m, 1H), 4.68-4.55 (m, 2H), 4.40 (dd,
J=47.4, 8.8 Hz, 1H), 4.00 (s, 3H), 2.32-2.27 (m, 1H), 2.17-2.09 (m,
2H), 1.74-1.63 (m, 1H). LC-MS (m/z) 426.1 (MH.sup.+); t.sub.R=1.9
(Method C)
EXAMPLE 24
N-[3-[(2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2--
yl]-4-fluorophenyl]-5-fluoro-pyridine-2-carboxamide (Compound
24)
##STR00071##
[0300] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-fluoropicolinic acid .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 9.80 (s, 1H), 8.46 (d, J=2.4 Hz 1H), 8.34-8.31
(m, 1H), 7.87-7.84 (m, 1H), 7.62-7.58 (m, 2H), 7.11-7.06 (m, 1H),
4.79-4.50 (m, 4H), 2.37-2.32 (m, 1H), 2.26-2.21 (m, 2H), 1.87-1.82
(m, 1H). LC-MS (m/z) 413.1 (MH.sup.+); t.sub.R=1.9 (Method C)
EXAMPLE 25
N-[3-R2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2-y-
l]-4-fluoro-phenyl]-5-methoxy-pyridine-2-carboxamide (Compound
25)
##STR00072##
[0302] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-methoxypicolinic acid .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 9.82 (s, 1H), 8.26-8.22 (m, 2H), 7.88-7.86 (m,
1H), 7.58-7.56 (m, 1H), 7.33 (dd, J=8.6, 2.0 Hz 1H), 7.07 (dd,
J=11.6, 9.2 Hz, 1H), 4.79-4.38 (m, 4H), 3.94 (s, 3H), 2.36-2.31 (m,
1H), 2.26-2.10 (m, 2H), 1.90-1.78 (m, 1H). LC-MS (m/z) 425.1
(MH.sup.+); t.sub.R=2.26 (Method D)
EXAMPLE 26
N-[3-[(2S,5R)-6-amino-5-fluoro-2,5-bis(fluoromethyl)-3,4-dihydropyridin-2--
yl]-4-fluorophenyl]-5-methoxy-pyrazine-2-carboxamide (Compound
26)
##STR00073##
[0304] Prepared from
(3R,6S)-6-(5-amino-2-fluorophenyl)-3-fluoro-3,6-bis(fluoromethyl)piperidi-
ne-2-thione and 5-methoxypyrazine-2-carboxylic acid .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 9.50 (s, 1H), 9.01 (s, 1H), 8.16 (s,
1H), 7.88-7.86 (m, 1H), 7.55 (dd, J=6.8, 2.0 Hz, 1H), 7.11-7.06 (m,
1H), 4.80-4.50 (m, 4H), 4.08 (s, 3H), 2.40-2.34 (m, 1H), 2.27-2.12
(m, 2H), 1.92-1.79(m, 1H). LC-MS (m/z) 426.1 (MH.sup.+);
t.sub.R=2.21 (Method D)
[0305] Pharmacological Testing
[0306] BACE1 Binding Assay
[0307] The binding assay was performed as SPA-based assay using a
biotinylated form of human BACE1 recombinantly expressed and
subsequently purified from Freestyle HEK293 cells. The binding
assay was run in a 50 mM sodium acetate buffer, pH 4.5 containing
50 mM NaCl and 0.03% Tween-20 in white clear bottom 384 plates
(Corning #3653). 10 nM (final concentration) radioligand
([.sup.3H]--N-((1S,2R)-1-benzyl-3-cyclopropylamino-2-hydroxy-propyl)-5-(m-
ethanesulfonyl-methyl-amino)-N-((R)-1-phenyl-ethyl)-isophthalamide)
(TRQ11569 purchased from GE Healthcare) was mixed with test
compound at a given concentration, 6 nM (final concentration) human
BACE1 and 25 .mu.g Streptavidin coated PVT core SPA beads
(RPNQ0007, GE Healthcare Life Sciences) in a total volume of 40
.mu.l. Several concentrations of each test compound were tested in
the assay for IC.sub.50 determination. The plates were incubated
for one hour at room temperature and counted in a Wallac Trilux
counter. Total and non-specific binding were determined using
buffer and 1 .mu.M (final concentration) of the high affinity BACE1
reference inhibitor
(S)-6-[3-chloro-5-(5-prop-1-ynyl-pyridin-3-yl)-thiophen-2-yl]-2-imino-3,6-
-dimethyl-tetrahydro-pyrimidin-4-one, respectively. For each test
compound, a IC.sub.50 value (the concentration mediating 50%
inhibition of the specific binding of the radioligand) was
determined from concentration-response curve and used to calculate
the K.sub.i from the equation K.sub.i=IC.sub.50/(1+L/K.sub.d),
where L and K.sub.d are the final concentration of the radioligand
used in the assay and the dissociation constant of the radioligand,
respectively. The K.sub.d of the radioligand was determined from
saturation binding experiments.
TABLE-US-00001 TABLE 1 binding affinity of selected compounds
Compound BACE1 1 22 2 20 3 10 4 11 5 24 6 46 7 67 8 62 9 26 10 22
11 55 12 140 13 40 14 130 15 52 16 27 17 42 18 300 19 130 20 230 21
67 22 28 23 17 24 180 25 160 26 160
[0308] BACE1 Efficacy Assay
[0309] The efficacy assay was performed as a FRET-based assay using
a commercially available BACE1 kit (Life Technologies, P2985). 2
.mu.l test compound at 10 .mu.M (final concentration) and 15 .mu.l
BACE1 enzyme from the kit (final concentration 3 nM) were
preincubated for 15 minutes at room temperature before addition of
15 .mu.l of substrate from the kit (250 nM final concentration) and
incubated for additional 90 minutes at room temperature. The assay
plate was subsequently read in a Pherastar (Ex540/Em590). The
enzyme activity observed in presence of test compound were
normalized to the enzyme activity observed in presence of buffer
and 10 .mu.M (final concentration) of the high affinity BACE1
reference inhibitor
(S)-6-[3-Chloro-5-(5-prop-1-ynyl-pyridin-3-yl)-thiophen-2-yl]-2-
-imino-3,6-dimethyl-tetrahydropyrimidin-4-one, respectively. The
efficacy of the test compounds was evaluated at 10 .mu.M (final
concentration) and defined as the percent inhibition of the enzyme
activity using the equation % inhibition=100%-normalized enzyme
activity in percent.
TABLE-US-00002 TABLE 2 BACE1 activity of selected compounds
Compound No BACE1 inhibition at 10 .mu.M (%) 1 106 2 104 3 105 4
102 5 103 6 106 7 104 9 103 10 103 11 103 12 103 13 100 14 103 15
104 16 104 17 104 18 103 19 104 20 102
[0310] Assessment of A.beta. Levels in Rat Brain and Plasma
Following BACE1 Inhibition. Animals.
[0311] All rat care and experimental procedures were approved by
Lundbeck Veterinary Staff, according to Danish legislature. The
rats were maintained in a barrier facility with a 12/12-h
light/dark cycle and ad libitum food and water access.
[0312] Treatment of Naive Rats.
[0313] Young adult Male Sprague Dawley rats of approximately 250 g
weight were purchased from Charles River and received 0-30 mg/kg of
vehicle (10% HP betaCD+1M MeSO.sub.4, pH 2.5) or test compounds
(dissolved in vehicle) only by oral gavage (p.o). The compounds are
dosed at a volume of 5 ml/kg. Cohorts of 5-10 animals were
established for each treatment condition.
[0314] The animals undergoing treatment were closely monitored by
veterinary staff for any signs of toxicity. Monitoring parameters
included body weight, physical appearance, changes in coat
appearance, occurrence of unprovoked behavior, and blunted or
exaggerated responses to external stimuli.
[0315] Tissue Collection.
[0316] At T=180 minutes after initial dosing the animals were
stunned and decapitated with a guillotine. Trunk-blood was sampled
in EDTA coated tubes after decapitation of the animal. The blood
was centrifuged at 2200G at 4.degree. C. for 15 minutes and the
plasma was collected and frozen at -80.degree. C. The blood was
aliquoted for A.beta. ELISA and pharmacokinetic analysis.
Immediately following sacrifice, the brain was extracted and split
into 2 halves. The right hemibrains were snap frozen on dry ice and
stored at -80.degree. C. The left half was dissected; with the
front forebrain taken for A.beta. ELISA and the remainder used for
DMPK analysis. These samples were also snap frozen on dry ice and
stored at -80.degree. C. until use for analysis.
[0317] Tissue Processing.
[0318] The cortex samples were thawed slightly on wet ice before
they were homogenized with a small volume dispersing instrument
(T10 basic ULTRA-TURRAX.RTM.) which was set at speed 5 for
approximately 5-7 sec. The tissue was processed in a 10 times
volume of the weight, for example 100 mg of tissue was homogenized
in 1000 .mu.L of Homogenization buffer. Homogenization buffer: 50
ml Milli Q water+50 nM NaCl+0.2% Diethylamin (DEA)+1 tablet of
Complete Protease inhibitor cocktail+1 nM 4-(2-aminoethyl)
benzenesulfonyl fluoride hydrochloride irreversible serine protease
inhibitor (AEBSF).
[0319] After homogenization 450 .mu.L aliquots of the samples were
collected into a 1.5 ml Eppendorf tube and placed on wet ice, 0.5%
NP-40 (50 ul) was added to all samples and then they were incubated
on ice for 30 min. After which all samples were sonicated using an
Ultrasonic homogenizer with 20 kHz homogeneous sound (SONOPLUS
HD2070, Bandelin Electronic) 10 pulse set at 12-13% power to
extract all the A.beta. species. The samples were then centrifuged
(Ole Dich 157 MPRF Micro centrifuge) at 20000G for 20 minutes at
4.degree. C. After centrifugation 2854 of the supernatant was
pipetted into 6004 microtubes tubes and neutralized with 154 of 1M
Tris-HCL buffer.
[0320] ELISA Protocol.
[0321] WAKO 294-62501 Human/Rat Abeta amyloid (40) kit was used for
all ELISA analyses. 30 .mu.L plasma samples or 30 .mu.L of the
cortex supernatants generated as described above were placed in 600
.mu.L microtubes tubes on wet ice. To this 30 .mu.L of 8M urea
(AppliChem A1049, 9025) was added to generate a 2-fold dilution.
Both plasma and cortex supernatants were incubated on ice for 30
min. Standard rows were prepared from the standard peptide stock
provided in the kit and standard diluent containing 1.6M Urea (200
.mu.L 8M urea+800 .mu.L of standard diluent) and 0.8M urea (400
.mu.L 8M urea+3600 .mu.L Standard diluent). A serial 2-fold
dilution of A.beta.340 from 100 pmol/ml to 0 pmol/L was prepared
for the assay.
[0322] After incubation with urea, all samples were further diluted
by addition of 5 times standard diluent from the Kit. This was done
by adding 240 .mu.L Standard Diluent to 60 .mu.L sample/urea
mixture, which was then mixed well. 100 .mu.L of each diluted
sample was pipetted into designated wells of the ELISA plate in
duplicates. The plate was then covered and incubated overnight at
4.degree. C. The following day, the ELISA kit was brought to room
temperature before use. The incubated plate was washed 5 times with
the 20.times. washing solution diluted in Milli Q water. 100 .mu.L
HRP-conjugate was applied to each well, and the plate was covered
and incubates at 4.degree. C. for 1 hr. The wash was repeated again
for 5 times. 100 .mu.L 3,3',5,5'-Tetramethylbenzidine (TMB)
solution was applied to each well and the plate was covered and
incubated in the dark at room temperature for 30 minutes. 100 .mu.L
STOP-solution was next applied to each well, and the plate was read
at 450 nm wavelength in a spectrophotometer (Labsystems Multiscan
Ascent) within 30 min of adding the STOP-solution to the wells.
[0323] Concentration of A.beta. in the samples was determined based
on a standard curve generated from standards containing known
concentrations of synthetic A.beta.40. Those skilled in the art
will appreciate that diethylamine (DEA) and urea extractions will
release soluble A.beta., and insoluble A.beta. respectively. Since
the ELISA kit is validated and widely used, it is accepted that as
long as the treatment conditions and assay conditions are the same
for each compound tested, then the assay should yield consistent
robust data for the compounds tested and produce minimal
discrepancies.
[0324] Data Analysis
[0325] To determine the concentration of A.beta.340 in the samples,
the interpolated values of the samples loaded on plates are
multiplied by 20 to account for the dilutions made when the volumes
of DEA, urea and neutralization solution were added up. Values are
calculated as percentage change in A.beta.40 compared to vehicle
treated animals.
[0326] Bioanalysis of Brain and Plasma Samples
[0327] TC (test compound) was determined in plasma and brain
homogenate using UltraPerformance LC.RTM. (UPLC.RTM.)
chromatography followed by tandem-MS (MS/MS) detection.
[0328] Apparatus:
[0329] Tecan Genesis RSP 200; Biomek NXP, Beckman Coulter; Sigma
4K15 centrifuge; Acquity UPLC, Waters; Sciex AP14000 TQ, Applied
Biosystems; MS software: Analyst version 1.4.1
[0330] Chemicals
[0331] Acetonitrile, HPLC-grade, Fluka, No. 34967N; Methanol,
HPLC-grade, Sigma-Aldrich, Lot 9003S; Formic acid, HPLC-grade,
Riedel-de Haen, Lot 51660; Purified water, Millipore Synergy UV
[0332] Sample Preparation
[0333] Brain homogenate was prepared by homogenizing the brain 1:4
(v/v) with water:2-propanol:DMSO (50:30:20 v/v/v) followed by
centrifugation and collection of the supernatant. Calibration
standards and QC samples were prepared using a Hamilton robot. 150
.mu.L of ISTD in acetonitrile (1 ng/mL ISTD) was added to 25 .mu.L
of calibration standards, QC samples and test samples (plasma and
brain homogenate) using a Biomek robot. After centrifugation (6200
g, 4.degree. C., 20 min) 100 .mu.L supernatant from each sample was
transferred to a new plate and mixed with 100 .mu.L water with 0.1%
formic acid using a Biomek robot (method file InVivo transfer).
After a quick centrifugation (6200 g, 4.degree. C., 5 min) the
samples were placed in the auto-sampler.
[0334] UPLC-MS/MS Analysis
[0335] MS/MS detection was done with an Applied Biosystems Sciex
API 4000 instrument in positive-ion electrospray ionisation mode.
TC and ISTD were detected at a parent>daughter mass to charge
ratio (m/z). Nitrogen was used for the nebulizer and collision
gases. The peak area correlated linearly with the plasma and brain
concentration of the analytes in the range of 1.00-1000 ng/mL
plasma and 5.00-5000 ng/g brain (corrected for dilution). If the
plasma/brain sample drug concentration was above 1000 ng/mL or 5000
ng/g, the sample was diluted appropriately in blank plasma/blank
brain homogenate before analysis.
[0336] Chromatographic System
[0337] Analytical columns:
[0338] Waters Acquity UPLC HSS C18 SB (pH 2-8) 1.8 .mu.m,
2.1.times.30 mm.
[0339] Mobile phase A: 0.1% aq. formic acid or 0.1% aq. ammonium
hydroxide
[0340] Mobile phase B: Acetonitrile with 0.1% aq. formic acid or
0.1% aq. ammonium hydroxide.
[0341] Weak wash: Methanol
[0342] Strong wash: Acetonitrile/Isopropanol/formic acid (50/50/2
v/v/v)
[0343] Flow: 0.6 mL/min
[0344] Run time: 3 min.
[0345] To waste: 0-0.5 min
[0346] Temperature: 40.degree. C.
[0347] Gradient:
TABLE-US-00003 Time (min) % A % B 0 98 2 0.01 98 2 1.5 5 95 2 5 95
2.2 98 2 3 98 2
[0348] Compound 1was administered at doses of 10 mg/kg p.o. and 30
mg/kg p.o. and brain and plasma samples were collected at 3 hours
post dose and the following exposures were measured as described
above.
TABLE-US-00004 TABLE 3 Results for compound 11 Dose Exp
Brain/Plasma A.beta.40 reduction (mg/kg) (ng/g) ratio (%) Brain Rat
10 416 2.1 21 Plasma Rat 195 39 Brain Rat 30 1352 2.3 41 Plasma Rat
590 60
TABLE-US-00005 TABLE 4 Results for compound 13 Dose Exp
Brain/Plasma A.beta.40 reduction (mg/kg) (ng/g) ratio (%) Brain Rat
10 588 1.25 43 Plasma Rat 469 38 Brain Rat 30 1592 1.59 59 Plasma
Rat 1000 38
TABLE-US-00006 TABLE 5 Results for compound 5 Dose Exp Brain/Plasma
A.beta.40 reduction (mg/kg) (ng/g) ratio (%) Brain Rat 10 536 1.95
56 Plasma Rat 275 44 Brain Rat 30 1606 2.74 65 Plasma Rat 587
46
[0349] As shown in table 3, table 4, and table 5 compounds of the
present invention are able to penetrate the blood brain barrier and
show efficacy in the CNS.
[0350] MDCK-MDR1 Assay
[0351] The permeability of the test compounds was assessed in
MDCK-MDR1 cells that were cultured to confluency (4-6 days) in a 96
transwell plate. Test compounds were diluted with the transport
buffer (HBSS+1% BSA) to a concentration of 0.5 .mu.M and applied to
the apical or basolateral side of the cell monolayer. Permeation of
the test compounds from A to B direction or B to A direction was
determined in triplicate over a 60-minute incubation time at
37.degree. C. and 5% CO2 with a relative humidity of 95%. Test
compounds were quantified by LC-MS/MS analysis based on the peaks
area ratios of analyte/IS in both the receiver and donor wells of
the transwell plate.
[0352] The apparent permeability coefficient Papp (cm/s) was
calculated using the equation:
Papp=(dCr/dt).times.Vr/(A.times.C0)
[0353] Where dCr/dt is the cumulative concentration of compound in
the receiver chamber as a function of time (.mu.M/s); Vr is the
solution volume in the receiver chamber (0.05 mL on the apical
side; 0.25 mL on the basolateral side); A is the surface area for
the transport, i.e. 0.0804 cm.sup.2 for the area of the monolayer;
C0 is the initial concentration in the donor chamber (.mu.M).
[0354] Compounds are classified Pgp substrates when efflux ratio
(Papp BA/Papp AB) is .gtoreq.2.
TABLE-US-00007 TABLE 6 Efflux ratio of selected compounds Compound
MDCK- MDR1 efflux ratio 1 0.89 2 2.32 3 4.87 4 1.34 5 1.07 6 7 8
2.36 9 8.61 10 10.6 11 1.89 13 1.5 14 1.06 15 1.17 16 1 17 0.95 19
1.18 20 0.77 21 0.97 22 1.08 23 1.06 24 0.88 25 1.09 26 0.92
[0355] As shown in tables 4, the majority of the exemplified
compounds of the present invention have MDCK-MDR1 efflux ratios
below 2 and are thus likely to be able to cross the blood brain
barrier (E Kerns, L Di, Drug-like Properties: Concepts, Structure
Design and Methods (2008) Elsevier).
* * * * *