U.S. patent application number 15/071955 was filed with the patent office on 2016-06-30 for aminopyridine derived compounds as lrrk2 inhibitors.
This patent application is currently assigned to H. Lundbeck A/S. The applicant listed for this patent is H. Lundbeck A/S, Vernalis (R&D) Ltd.. Invention is credited to I-Jen Chen, Laurent David, Gitte Kobberoe Mikkelsen, Garrick Paul Smith, Stephen Watson, Douglas Stewart Williamson.
Application Number | 20160184317 15/071955 |
Document ID | / |
Family ID | 51062163 |
Filed Date | 2016-06-30 |
United States Patent
Application |
20160184317 |
Kind Code |
A1 |
Mikkelsen; Gitte Kobberoe ;
et al. |
June 30, 2016 |
Aminopyridine Derived Compounds as LRRK2 Inhibitors
Abstract
The present invention is directed to aminopyridine derived
compounds of formula (A) ##STR00001## The compounds are considered
useful for the treatment of diseases associated with LRRK2 such a
Lewy body dementia, Parkinson's disease or cancer.
Inventors: |
Mikkelsen; Gitte Kobberoe;
(Ballerup, DK) ; David; Laurent; (Malmo, SE)
; Watson; Stephen; (Hertfordshire, GB) ; Smith;
Garrick Paul; (Valby, DK) ; Williamson; Douglas
Stewart; (Winnersh, GB) ; Chen; I-Jen;
(Winnersh, GB) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
H. Lundbeck A/S
Vernalis (R&D) Ltd. |
Valby
Winnersh |
|
DK
GB |
|
|
Assignee: |
H. Lundbeck A/S
Valby
DK
Vernalis (R&D) Ltd.
Winnersh
GB
|
Family ID: |
51062163 |
Appl. No.: |
15/071955 |
Filed: |
March 16, 2016 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14758884 |
Jul 1, 2015 |
9321748 |
|
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PCT/EP2013/078105 |
Dec 30, 2013 |
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15071955 |
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61777120 |
Mar 12, 2013 |
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61748142 |
Jan 2, 2013 |
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Current U.S.
Class: |
514/236.2 ;
514/255.05; 514/318; 514/339 |
Current CPC
Class: |
C07D 403/14 20130101;
A61K 31/497 20130101; A61K 31/4545 20130101; C07D 401/14 20130101;
C07D 413/04 20130101; A61K 31/5377 20130101; A61P 43/00 20180101;
C07D 413/14 20130101; A61P 25/28 20180101; A61P 37/00 20180101;
A61P 1/04 20180101; A61P 35/00 20180101; A61P 25/16 20180101; C07D
401/04 20130101; A61K 31/4439 20130101 |
International
Class: |
A61K 31/5377 20060101
A61K031/5377; A61K 31/4545 20060101 A61K031/4545; A61K 31/497
20060101 A61K031/497; A61K 31/4439 20060101 A61K031/4439 |
Foreign Application Data
Date |
Code |
Application Number |
Jan 2, 2013 |
DK |
PA201300004 |
Mar 11, 2013 |
DK |
PS201300136 |
Claims
1. A method of treating Parkinson's disease or Lewy body dementia,
wherein said method comprises administering to a subject in need
thereof a pharmaceutical composition that comprises a
therapeutically effective amount of a compound of Formula (A):
##STR00068## or a pharmaceutically acceptable salt thereof;
wherein: R1 represents triazolyl or oxadizolyl, said triazolyl or
oxadizolyl may optionally be substituted with 1 R5 group, X
represents N or CH when R1 represents triazolyl or when R1
represents oxadiazolyl X represents CH, R5 represents
C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl or a 5-6 membered
heterocyclic ring with 1 or 2 heteroatom(s), said cycloalkyl or
heterocyclic ring are optionally substituted with 1 or 2
C.sub.1-C.sub.3 alkyl or 1 or 2 C.sub.1-C.sub.3 alkoxy, R2, R3 and
R4 each independently represents hydrogen, halogen, C.sub.1-C.sub.6
alkyl, C.sub.1-C.sub.6 alkoxy, O(CH.sub.2).sub.nCF.sub.3,
(CH.sub.2).sub.nR6, (C.dbd.O)R6 or (CH.sub.2).sub.n(C.dbd.O)R6,
n=0, 1, 2 or 3, or R2 and R3 or R3 and R4 may together with the
atom they are attached to form a 9-10 membered bicyclic
heterocyclic ring with 1 or 2 heteroatom(s), said bicyclic
heterocyclic ring may optionally be substituted with 1 or 2
C.sub.1-C.sub.6 alkyl or 1 or 2 C.sub.1-C.sub.6 alkoxy, R6
represents a 5-6 membered heterocyclic ring with 1, 2 or 3
heteroatom(s), said heterocyclic ring may optionally be substituted
with 1 or 2 C.sub.1-C.sub.3 alkyl or 1 or 2 C.sub.1-C.sub.3
alkoxy.
2. The method of claim 1, wherein said method is a method of
treating Parkinson's disease.
3. The method of claim 2, wherein said Parkinson's disease is
idiopathic Parkinson's disease or sporadic Parkinson's disease.
4. The method of claim 1, wherein said method is a method of
treating Lewy body dementia.
5. The method according to claim 1, wherein said heteroatom(s) of
said compound of Formula (A) are independently selected from N, O
or S.
6. The method according to claim 1, wherein said halogen(s) of said
compound of Formula (A) are independently fluoro, chloro, bromo or
iodo.
7. The method according to claim 1, wherein said C.sub.1-C.sub.6
alkyl group of said compound of Formula (A) is methyl, ethyl,
propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl or
isopentyl.
8. The method according to claim 1, wherein said C.sub.1-C.sub.3
alkyl group of said compound of Formula (A) is methyl, ethyl,
propyl, or isopropyl.
9. The method according to claim 1, wherein said C.sub.1-C.sub.6
alkoxy group of said compound of Formula (A) is methoxy, ethoxy,
propoxy, butoxy, isobutoxy, t-butoxy, pentoxy, or isopropoxy.
10. The method according to claim 1, wherein said C.sub.1-C.sub.3
alkoxy group of said compound of Formula (A) is methoxy, ethoxy,
propoxy or isopropoxy.
11. The method according to claim 1, wherein R1 of said compound of
Formula (A) is: ##STR00069## wherein * denotes the attachment
point.
12. The method according to claim 1, wherein R5 of said compound of
Formula (A) is selected from the group consisting of
C.sub.1-C.sub.3 alkyl, C.sub.3-C.sub.6 cycloalkyl, ##STR00070##
wherein * denotes the attachment point.
13. The method according to claim 1, wherein R2, R3 and/or R4 of
said compound of Formula (A) is independently selected from the
group consisting of: ##STR00071## wherein * denotes the attachment
point.
14. The method according to claim 1, wherein R2 and R3 or R3 and R4
of said compound of Formula (A), together with the atom they are
attached to form a bicyclic heterocyclic ring selected from the
group consisting of: ##STR00072## wherein * denotes the attachment
point
15. The method according to claim 1, wherein said compound of
Formula (A) is selected from the group consisting of: (1)
5-(1H-Indol-5-yl)-3-(5-isopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine-
; (2)
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1H-indol-5-yl)-pyridin-2--
ylamine; (3)
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1-methyl-2,3-dihydro-1H-indol--
5-yl)-pyridin-2-ylamine; (4)
3-[5-(1-Methyl-piperidin-4-yl)-[1,3,4]oxadiazol-2-yl]-5-p-tolyl-pyridin-2-
-ylamine; (5)
5-(1-Methyl-2,3-dihydro-1H-indol-5-yl)-3-(5-piperidin-4-yl-[1,3,4]oxadiaz-
ol-2-yl)-pyridin-2-ylamine; (6)
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(4-morpholin-4-ylmethyl-phenyl)-
-pyridin-2-ylamine; (7)
5-(3-Methoxy-phenyl)-3-(5-piperidin-4-yl-[1,3,4]oxadiazol-2-yl)-pyridin-2-
-ylamine; (8)
5-(3-Methoxy-phenyl)-3-[5-(1-methyl-piperidin-4-yl)-[1,3,4]oxadiazol-2-yl-
]-pyridin-2-ylamine; (9)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-1H-indol-5-yl)-pyridin-
-2-ylamine; (10)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indol-5-
-yl)-pyridin-2-ylamine; (11)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indol-5-
-yl)-pyrazin-2-ylamine; (12)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-ylmethyl-phenyl)--
pyridin-2-ylamine; (13)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-yl-phenyl)-pyridi-
n-2-ylamine; (14)
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-[4-(2-morpholin-4-yl-ethyl)-phen-
yl]-pyridin-2-ylamine; (15)
5-(3-fluoro-4-(morpholinomethyl)phenyl)-3-(1-isopropyl-1H-1,2,3-triazol-4-
-yl)pyridin-2-amine; (16)
(4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-fluoroph-
enyl)(morpholino)methanone; (17)
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl-2-methoxy-phenyl]-morph-
olino-methanone; (18)
3-(1-isopropyltriazol-4-yl)-5-[3-methoxy-4-(morpholinomethyl)phenyl]pyrid-
in-2-amine; (19)
5-[4-fluoro-3-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine; (20)
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methyl-phenyl]-morph-
olino-methanone; (21)
3-(1-Isopropyltriazol-4-yl)-5-[3-methyl-4-(morpholinomethyl)phenyl]pyridi-
n-2-amine; (22)
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-chloro-phenyl]-morph-
olino-methanone; (23)
5-[3-Chloro-4-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine; (24)
3-(1-Isopropyltriazol-4-yl)-5-[3-(morpholinomethyl)phenyl]pyridin-2-amine-
; (25)
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morpholin-
o-methanone; (26)
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-5-chloro-phenyl]-morph-
olino-methanone; (27)
5-[3-Chloro-5-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine; (28)
[4-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morpholino-meth-
anone; (29)
3-(5-cyclopropyl-1,3,4-oxadiazol-2-yl)-5-[3-(morpholinomethyl)phenyl]pyri-
din-2-amine; and (30)
2-[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-1-morpholino--
ethanone.
16. The method of claim 15, wherein said method is a method of
treating Parkinson's disease.
17. The method of claim 16, wherein said Parkinson's disease is
idiopathic Parkinson's disease or sporadic Parkinson's disease.
18. The method of claim 15, wherein said method is a method of
treating Lewy body dementia.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of, and claims the
benefit of, U.S. patent application Ser. No. 14/758,884 (filed Jul.
1, 2015; pending), which application is a .sctn.371 U.S. National
Stage Application of PCT International Application No.
PCT/EP2013/078105, (filed Dec. 30, 2013, now expired), which claims
the benefit of U.S. Provisional Patent Applications No. 61/748,142
(filed Jan. 2, 2013, now expired) and 61/777,120 (filed Mar. 12,
2013, now expired) and the benefit of Danish Applications No.
PA201300136 (filed Mar. 11, 2013) and PA201300004 (filed Jan. 2,
2013). Each of these applications is hereby incorporated by
reference in its entirety.
REFERENCE TO SEQUENCE LISTING
[0002] This application includes one or more Sequence Listings
pursuant to 37 C.F.R. 1.821 et seq., which are disclosed in both
paper and computer-readable media, and which paper and
computer-readable disclosures are herein incorporated by reference
in their entireties.
FIELD OF THE INVENTION
[0003] The present invention relates to aminopyridine derivatives
which are LRRK2 inhibitors and thus useful in therapy and to
pharmaceutical composition comprising said compounds.
BACKGROUND OF THE INVENTION
[0004] Parkinson's disease is a neurodegenerative disease. It is
the second most common neurodegenerative disease after Alzheimer's
disease and affects more than 1% of the population above the age of
65. Parkinson's disease is clinically characterised by resting
tremor, bradykinesia and muscular rigidity. Pathologically, the
disease is characterised by loss of dopaminergic neurons with the
consequent decrease in dopamine levels in the brain and by
aggregation of the protein .alpha.-synuclein in the dopaminergic
neurons. These aggregations called Lewy-bodies are composed of
insoluble .alpha.-synuclein phosporylated at serine-129 and
ubiquitin. Current Parkinson's disease therapeutic intervention
strategies aim at increasing the dopamine levels in areas
innervated by dopaminergic neurons in the brain. Levadopa is a
precursor of dopamine, and it is therapeutically used to increase
dopamine levels. Carbidopa is an inhibitor of the enzyme
aromatic-L-amino-acid decarboxylase also known as DOPA
decarboxylase, and it is often co-administered with levadopa to
increase the fraction of levadopa which reaches the clinically
relevant regions in the brain. Monoamine oxidase B inhibitors are
administered to increase the levels of dopamine by blocking the
metabolism of dopamine As an alternative, dopamine agonists are
administered to stimulate dopaminergic neurons, an effect similar
to that obtained by increasing the dopamine levels. Although these
therapies provide significant symptomatic benefit to the patient,
they are also associated with adverse side effects and often become
ineffective after prolonged treatment Importantly, neither of the
existing therapies addresses the underlying and disease causing
problem, i.e. the progressive loss or inactivation of dopaminergic
neurons.
[0005] Leucine-Rich Repeat Kinase 2 (LRRK2) is a 2527 amino acid
protein involved in catalysing phosphorylation and GTP-GTD
hydrolysis. The NCBI reference sequence for human LRKK2 mRNA is
NM_198578.2. Evidence is mounting showing a relationship between
LRRK2 and the pathogenesis of Parkinson's disease. It has been
shown that LRRK2 phosphorylates .alpha.-synuclein at serine-129,
and as discussed above this phosphorylated form constitutes a
significant part of the Lewy-bodies [BiochemBiophys Res Comm., 387,
149-152, 2009]. Additionally, single nucleotide polymorphisms in
functional domains of LRRK2 have been shown to cause familiar and
sporadic Parkinson's disease. So far at least 6 pathogenic variants
have been identified, i.e. Gly2019Ser, Ile2020Thr, Arg1441Cys,
Arg1441Gly, Arg1441His and Tyr1699Cys [Parkinsonism Rel. Dis.,15,
466-467, 2009; Movement Dis., 25, 2340-2345, 2010; Neuron, 44,
601-607, 2004; and Lancet, 365, 412-415, 2005] Importantly, the
clinical features of Parkinson's disease associated with LRRK2
mutations cannot be distinguished from those featuring in
idiopathic Parkinson's disease. This strongly suggests a common
pathogenic mechanism and that LRKK2 activity is a rate-limiting
factor in Parkinson's disease progression [FEBS Journal, 276,
6436-6444, 2009].
[0006] The most common pathogenic form of LRRK2-associated
Parkinson's disease is found in carriers of the amino acid
substitution Gly2019Ser in the kinase domain of the LRRK2 protein.
Gly2019Ser Parkinson's disease is inherited in an autosomal
dominant fashion suggesting a gain-of-function mutation of the
LRRK2 protein. In support of this notion, biochemical studies have
shown that both the glycine to serine substitution at amino acid
position 2019 as well as isoleucine to threonine substitution at
amino acid position 2020 in the kinase domain lead to an increased
kinase activity of LRRK2 [Proc. Nat. Acad. Sci USA, 102,
16842-16847, 2005]. This suggests a causal involvement of
overactive LRRK2 in the pathogenesis of familiar forms of
Parkinson's disease. Thus, inhibitors of LRRK2, including e.g. the
G2019S and 12020T mutations, could be used as disease modifying
treatment in familiar Parkinson's disease.
[0007] In cellular and animal studies several phosphorylation sites
in the LRRK2 protein have been identified. Most prominent,
phosphorylation of LRRK2 at two conserved residues serine at amino
acid position 910 and serine at amino acid position 935 in human
LRRK2 located just amino terminal to the leucine-rich repeat domain
mediates binding to 14-3-3 proteins. Phosphorylation at serine
residues 910 and 935 were shown to be dependent on an active LRRK2
conformation and further, that LRRK2 kinase inhibitors can inhibit
phosphorylation at these two sites [Biochem J., 430, 405-13, 2010;
J Neurochem., 120:37-45, 2012].
[0008] LRRK2 kinase inhibitors have been shown to
concentration-dependently inhibit p LRRK2-Ser910 and LRRK2-Ser935
phosphorylation in cellular models expressing LRRK2 and
LRRK2-G2019S as well as human LRRK2-expressing lymphoblastoid cells
from PD patients homozygous for the LRRK2 G2019S mutation. In
addition, LRRK2 kinase inhibition dose-dependently inhibits
LRRK2-Ser910 and LRRK2-Ser935 phosphorylation in mouse brain after
in vivo administration of compound. [ACS Med. Chem. Lett., DOI:
10.1021/m1300123a, 2012.].
[0009] Common single nucleotide polymorphisms of LRRK2 have also
been associated with Parkinson's disease [Nat Genet. 2009 December;
41(12):1308-12] [Mov Disorder. Oct. 31, 2012; doi:
10.1002/mds.25226]. A recent genome wide association meta-analysis
study where correction for G2019S carrier status was performed
indicated that common LRRK2 variants with minor allele frequency
(MAF) above 1% also are associated with an increased risk of
Parkinson's disease [Lancet. 377, 641-649, 2011]. Further,
investigations of common exonic polymorphic variants have
highlighted several LRRK2 Parkinson's disease risk variants: in
Caucasians the M1646T mutation, and in the Asian population the
A419V mutation and also the previously found G2385R mutation
[Lancet Neurol. 10, 898-908, 2011]. This indicates that LRRK2
inhibitors also could be useful as disease-modifying treatment in
Parkinson's disease patients carrying common genomic LRRK2 variants
such as M1646T, G2385R and A419V.
[0010] Indeed, as discussed above, as the clinical features of
LRRK2 associated and idiopathic Parkinson's disease are very
similar this also suggests that LRRK2 inhibitors could be useful
for the treatment of sporadic PD.
[0011] As established above, LRRK2 inhibitors may by used in the
treatment of Parkinson's disease and particular mention is made of
Parkinson's disease associated with mutations in LRRK2, such as
Gly2019Ser. Moreover, LRRK2 inhibitors are also expected to be
useful in the treatment of other diseases which are associated with
LRRK2. LRRK2 has been identified as a core component in Lewy bodies
and is thus expected to be useful in the treatment of Lewy body
dementia [Neuropathol. Appl. Neurobiol., 34, 272-283, 2008].
Expression of LRRK2 mRNA is highly enriched in brain, lungs,
kidney, spleen and blood suggesting that functional impact of
increased LRRK2 activity is likely to be most relevant in
pathogenic and pathologic conditions associated with those regions.
Support for that notion can be found in studies showing an
increased risk of non-skin cancer in LRRK2 Gly2019Ser mutation
carriers and especially for renal and lung cancer [Mov. Disorder,
25, 2536-2541, 2010]. Over-expression of LRRK2 by chromosomal
amplification has also been identified in papillary renal and
thyroid carcinomas. Also, genetic association of LRRK2 has been
reported to diseases in where aberrant responses of the immune
system are involved. This is the case for inflammatory bowel
diseases such as Crohn's disease and ulcerative colitis as well as
for leprosy [Nat Genet.42, 1118-1125, 2010; Inflamm. Bowel. Dis.
16, 557-558, 2010; N Engl. J Med. 361, 2609-2618, 2009; Inflamm.
Bowel. Dis.doi: 10.1002/ibd.21651, 2011].
SUMMARY OF THE INVENTION
[0012] The present inventors have surprisingly found certain
aminopyridine derivatives which are LRRK2 inhibitors. Accordingly,
in one embodiment the invention provides compounds of formula (A),
below:
##STR00002##
wherein [0013] R1 represents triazolyl or oxadizolyl, said
triazolyl or oxadizolyl may optionally be substituted with 1 R5
group, [0014] X represents N or CH when R1 is triazolyl or when R1
is oxadiazole X represents CH, [0015] R5 represents C.sub.1-C.sub.6
alkyl, C.sub.3-C.sub.6 cycloalkyl or a 5-6 membered heterocyclic
ring with 1 or 2 heteroatom(s), said cycloalkyl or heterocyclic
ring are optionally substituted with 1 or 2 C.sub.1-C.sub.3 alkyl
or 1 or 2 C.sub.1-C.sub.3 alkoxy, [0016] R2, R3 and R4 each
independently represents hydrogen, halogen, C.sub.1-C.sub.6 alkyl,
C.sub.1-C.sub.6 alkoxy, O(CH.sub.2).sub.nCF.sub.3,
(CH.sub.2).sub.nR6, (C.dbd.O)R6 or (CH.sub.2).sub.n(C.dbd.O)R6,
n=0, 1, 2 or 3, or [0017] R2 and R3 or R3 and R4 may together with
the atom they are attached to form a 9-10 membered bicyclic
heterocyclic ring with 1 or 2 heteroatom(s), said bicyclic
heterocyclic ring may optionally be substituted 1 or 2
C.sub.1-C.sub.6 alkyl or 1 or 2 C.sub.1-C.sub.6 alkoxy, [0018] R6
represents a 5-6 membered heterocyclic ring with 1, 2 or 3
heteroatom(s), said heterocyclic ring may optionally be substituted
with 1 or 2 C.sub.1-C.sub.3 alkyl or 1 or 2 C.sub.1-C.sub.3 alkoxy,
and pharmaceutically acceptable salts thereof.
[0019] In one embodiment, the invention provides a pharmaceutical
composition comprising a compound of the above formula A and
pharmaceutically acceptable salts thereof together with a
pharmaceutically acceptable excipient.
[0020] In one embodiment, the invention provides compounds of the
above formula A and pharmaceutically acceptable salts thereof for
use in therapy.
[0021] In one embodiment, the invention provides compounds of the
above formula A and pharmaceutically acceptable salts thereof for
use in a method for the treatment of a disease associated with
LRRK2.
[0022] In one embodiment, the invention relates to the use of a
compound of the above formula A and pharmaceutically acceptable
salts thereof in the manufacture of a medicament for use in the
treatment of a disease associated with LRRK2.
[0023] In one embodiment, the invention relates to a method for the
treatment of a disease associated with LRRK2, the method comprising
the administration of a therapeutically effective amount of a
compound of the above formula A and pharmaceutically acceptable
salts thereof to a patient in need thereof.
DETAILED DESCRIPTION OF THE INVENTION
[0024] According to an embodiment of the invention the halogens in
formula A hereinabove are independently selected from the group
comprising fluoro, chloro, bromo or iodo and the heteroatoms may
independently be selected from N, O or S.
[0025] Furthermore, the C.sub.1-C.sub.6 alkyl group may be selected
from the group comprising methyl, ethyl, propyl, isopropyl, butyl,
isobutyl, t-butyl, pentyl or isopentyl and the C.sub.1-C.sub.3
alkyl group may be selected from the group comprising methyl,
ethyl, propyl, isopropyl.
[0026] The C.sub.1-C.sub.6 alkoxy group may be selected from the
group comprising methoxy, ethoxy, propoxy, butoxy, isobutoxy,
t-butoxy, pentoxy, isopropoxy and the C.sub.1-C.sub.3 alkoxy group
may be selected from the group comprising methoxy, ethoxy, propoxy
or isopropoxy.
[0027] In an embodiment R1 in formula A may be selected from the
group comprising
##STR00003##
wherein * denotes the attachment point.
[0028] R5 in Formula (A) may be selected from the group comprising
C.sub.1-C.sub.3 alkyl, C.sub.3-C.sub.6 cycloalkyl or a heterocyclic
ring selected from
##STR00004##
wherein * denotes the attachment point. [0029] R2, R3 and/or R4 in
formula A may be selected from the group comprising
##STR00005##
[0029] wherein * denotes the attachment point. [0030] R2 and R3 or
R3 and R4 may according to some embodiments form a bicyclic
heterocyclic ring together with the atom they are attached selected
from the group comprising
##STR00006##
[0030] wherein * denotes the attachment point.
[0031] In a specific embodiment the compound are selected from the
group comprising
5-(1H-Indol-5-yl)-3-(5-isopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine-
,
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1H-indol-5-yl)-pyridin-2-ylam-
ine,
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1-methyl-2,3-dihydro-1H-in-
dol-5-yl)-pyridin-2-ylamine,
3-[5-(1-Methyl-piperidin-4-yl)-[1,3,4]oxadiazol-2-yl]-5-p-tolyl-pyridin-2-
-ylamine,
5-(1-Methyl-2,3-dihydro-1H-indol-5-yl)-3-(5-piperidin-4-yl-[1,3,-
4]oxadiazol-2-yl)-pyridin-2-ylamine,
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(4-morpholin-4-ylmethyl-phenyl)-
-pyridin-2-ylamine,
5-(3-Methoxy-phenyl)-3-(5-piperidin-4-yl-[1,3,4]oxadiazol-2-yl)-pyridin-2-
-ylamine,
5-(3-Methoxy-phenyl)-3-[5-(1-methyl-piperidin-4-yl)-[1,3,4]oxadi-
azol-2-yl]-pyridin-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-1H-indol-5-yl)-pyridin-
-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indol-5-
-yl)-pyridin-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indol-5-
-yl)-pyrazin-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-ylmethyl-phenyl)--
pyridin-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-yl-phenyl)-pyridi-
n-2-ylamine,
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-[4-(2-morpholin-4-yl-ethyl)-phen-
yl]-pyridin-2-ylamine,
5-(3-fluoro-4-(morpholinomethyl)phenyl)-3-(1-isopropyl-1H-1,2,3-triazol-4-
-yl)pyridin-2-amine,
(4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-fluoroph-
enyl)(morpholino)methanone,
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methoxy-phenyl]-morp-
holino-methanone,
3-(1-isopropyltriazol-4-yl)-5-[3-methoxy-4-(morpholinomethyl)phenyl]pyrid-
in-2-amine,
5-[4-fluoro-3-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine,
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methyl-phe-
nyl]-morpholino-methanone,
3-(1-Isopropyltriazol-4-yl)-5-[3-methyl-4-(morpholinomethyl)phenyl]pyridi-
n-2-amine,
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-chloro-phe-
nyl]-morpholino-methanone,
5-[3-Chloro-4-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine,
3-(1-Isopropyltriazol-4-yl)-5-[3-(morpholinomethyl)phenyl]pyrid-
in-2-amine,
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morpholino-meth-
anone,
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-5-chloro-phenyl]-
-morpholino-methanone,
5-[3-Chloro-5-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridi-
n-2-amine,
[4-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morph-
olino-methanone,
3-(5-cyclopropyl-1,3,4-oxadiazol-2-yl)-5-[3-(morpholinomethyl)phenyl]pyri-
din-2-amine,
2-[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-1-morpholino--
ethanone,
or a pharmaceutical acceptable salt thereof.
[0032] The above mentioned compounds may be in a composition as the
sole active ingredient or in combination with other active
ingredients. Additionally, one or more pharmaceutically acceptable
carriers or diluents may be in the composition.
[0033] The compounds preferably have an IC50 value below 1000
nM.
[0034] 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.
[0035] In this context is understood that when specifying the
enantiomeric form, 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%.
[0036] Racemic forms can 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. 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 of
diastereomeric derivatives. 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.
[0037] Furthermore, when a double bond or a fully or partially
saturated ring system is present in the molecule geometric isomers
may be formed. It is intended that any geometric isomers as
separated, pure or partially purified geometric isomers or mixtures
thereof are included within the scope of the invention. Likewise,
molecules having a bond with restricted rotation may form geometric
isomers. These are also intended to be included within the scope of
the present invention.
[0038] Furthermore, some of the compounds of the present invention
may exist in different tautomeric forms and it is intended that any
tautomeric forms that the compounds are able to form are included
within the scope of the present invention.
[0039] In the present context, "pharmaceutically acceptable salts"
include pharmaceutical acceptable acid addition salts,
pharmaceutically acceptable metal salts, ammonium and alkylated
ammonium salts. Acid addition salts include salts of inorganic
acids as well as organic acids.
[0040] Examples of suitable inorganic acids include hydrochloric,
hydrobromic, hydroiodic, phosphoric, sulfuric, sulfamic, nitric
acids and the like.
[0041] 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
pharmaceutical acceptable inorganic or organic acid addition salts
include the pharmaceutically acceptable salts listed in J. Pharm.
Sci. 1977, 66,2, which is incorporated herein by reference.
[0042] Examples of metal salts include lithium, sodium, potassium,
magnesium salts and the like.
[0043] Examples of ammonium and alkylated ammonium salts include
ammonium, methyl-, dimethyl-, trimethyl-, ethyl-, hydroxyethyl-,
diethyl-, n-butyl-, sec-butyl-, tert-butyl-, tetramethylammonium
salts and the like.
[0044] The term "aromatic" refers to a cyclic or polycyclic moiety
having a conjugated unsaturated (4n+2) .pi. electron system (where
n is a positive integer), sometimes referred to as a delocalized
.pi. electron system. The term "heteroaromatic" intents to indicate
an aromatic ring structure with one or more heteroatoms. Examples
may include pyridinyl and pyrimidinyl.
[0045] In the present context, "alkyl" is intended to indicate a
cyclic, straight or branched saturated hydrocarbon. In particular,
C.sub.1-6-alkyl is intended to indicate such hydrocarbon having 1,
2, 3, 4, 5 or 6 carbon atoms and likewise C.sub.1-3-alkyl is
intended to indicate a hydrocarbon having 1, 2 or 3 carbon atoms.
Typical alkyl groups include, but are not limited to, methyl,
ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl,
isopentyl, hexyl and the like.
[0046] The term "alkoxy" as used herein refers to a group of
formula --O-- alkyl, wherein alkyl is defined as above. In
particular, C.sub.1-C.sub.6-akoxy is intended to indicate such
hydrocarbon having 1, 2, 3, 4, 5 or 6 carbon atoms and likewise
C.sub.1-C.sub.3-akoxy indicate is intended to indicate a
hydrocarbon having 1, 2 or 3 carbon atoms. Examples of alkoxy
groups include, but are not limited to, methoxy, ethoxy, propoxy,
butoxy, isobutoxy, t-butoxy, pentoxy, isopropoxy and the like.
[0047] In the present context, "halogen" is intended to indicate
members of the 7.sup.th main group of the periodic table of the
elements, such as fluoro, chloro, bromo and iodo.
[0048] "Heteroatom" is intended to mean sulfur, oxygen or
nitrogen.
[0049] The term "cyclic" as used herein refers to any cyclic
structure, including alicyclic, heterocyclic, aromatic,
heteroaromatic and polycyclic fused or non-fused ring systems. The
term "membered" is meant to denote the number of skeletal atoms
that constitute the ring. Thus, for example, pyridinyl, pyranyl,
and pyrimidinyl are six-membered rings and pyrrolyl,
tetrahydrofuranyl, and thiophenyl are five-membered rings.
[0050] Alkyl groups as described herein-above may according to some
embodiments of the present invention also be cyclic. Examples of
such types of alkylgroups include, but are not limited to
cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl,
cycloheptenyl, and cyclooctanyl. The term "cycloalkyl" as used
herein refers to a saturated monocyclic hydrocarbon radical. In
particular, C.sub.3-C.sub.6 cycloalkyl refers to a saturated
monocyclic hydrocarbon radical having 3, 4, 5 or 6 carbon
atoms.
[0051] The terms "heterocycle", "heterocyclic" and "heterocyclyl"
as used herein, alone or in combination, refers to saturated or
unsaturated nonaromatic rings containing from five to about 10 ring
atoms where one or more of the ring atoms are heteroatoms.
[0052] In some embodiments of the invention a heterocylic ring is
intended to mean a 5 or 6 membered cyclic ring structure with 1, 2
or 3 heteroatom(s).
[0053] The term "heterocyclic" may also include fused rings. A
fused heterocyclic ring may contain from two to four fused rings
where the attaching ring is a heterocyclic, and the other
individual rings within the fused heterocyclic ring may be
alicyclic, heterocyclic, aromatic, heteroaromatic or any
combination thereof. Example of heterocyclics include, without
limitation, morpholinyl, piperidinyl, tetrahydrofuranyl,
benzodiazepinyl, tetrahydroindazolyl, dihyroquinolinyl, and the
like.
[0054] In some embodiments the fused heterocyclic ring constitutes
2 fused rings which each may be saturated or unsaturated to form a
"bicyclic heterocyclic ring" constituting a bicyclic ring structure
of a total of 9-10 members. This bicyclic heterocyclic ring may
have 1 or 2 heteroatoms in one or both of the rings.
[0055] Examples of "unsaturated heterocyclic groups" includes for
example a furyl group, a thienyl group, a pyrrolyl group, an
azepinyl group, a pyrazolyl group, an imidazolyl group, an oxazolyl
group, an isoxazolyl group, a thiazolyl group, an isothiazolyl
group, a 1,2,3-oxadiazolyl group, a triazolyl group, a tetrazolyl
group, a thiadiazolyl group, a pyranyl group, a pyridyl group, a
pyridazinyl group, a pyrimidinyl group and a pyrazinyl group; and
"saturated heterocyclic group" such as a tetrahydropyranyl group, a
tetrahydrothienyl group, a morpholinyl group, a thiomorpholinyl
group, a pyrrolidinyl group, a pyrrolinyl group, an imidazolidinyl
group, a pyrazolidinyl group, a piperidinyl group, a piperazinyl
group, an oxazolidinyl group, an isoxazolidinyl group, a
thiazolidinyl group, a pyrazolidinyl group, a dioxolanyl group and
a dioxanyl group.
[0056] The terms "substituents" or "substituted" as used herein,
alone or in combination, refer to groups which may be used to
replace hydrogen. The substituted molecule may itself be further
substituted in some embodiments of the invention.
[0057] In the present context, the term "therapeutically effective
amount" of a compound is intended to indicate an amount sufficient
to cure, alleviate or partially arrest the clinical manifestations
of a given disease and its complications in a therapeutic
intervention comprising the administration of said compound. An
amount adequate to accomplish this is defined as "therapeutically
effective amount". Effective amounts for each purpose will depend
on the severity of the disease or injury as well as the weight and
general state of the subject. It will be understood that
determining an appropriate dosage may be achieved using routine
experimentation, e.g. by constructing a matrix of values and
testing different points in the matrix, which is all within the
ordinary skills of a trained physician.
[0058] In the present context, the term "treatment" and "treating"
means the management and care of a patient for the purpose of
combating a disease. The term is intended to include the full
spectrum of treatments for a given disease from which the patient
is suffering, such as administration of the active compound to
alleviate the symptoms or complications, to delay the progression
of the disease, to alleviate or relief the symptoms and
complications, and/or to cure or eliminate the disease. The patient
to be treated is preferably a mammal, in particular a human being.
In the present context, "disease" can be used synonymous with
disorder, condition, malfunction, dysfunction and the like.
[0059] As established above, LRRK2 inhibitors may by used in the
treatment of Parkinson's disease and particular mention is made of
Parkinson's disease associated with mutations in LRRK2, such as
Gly2019Ser. Moreover, LRRK2 inhibitors are also expected to be
useful in the treatment of other diseases which are associated with
LRRK2. LRRK2 has been identified as a core component in Lewy bodies
and is thus expected to be useful in the treatment of Lewy body
dementia [Neuropathol. Appl. Neurobiol., 34, 272-283, 2008].
Expression of LRRK2 mRNA is highly enriched in brain, lungs,
kidney, spleen and blood suggesting that functional impact of
increased LRRK2 activity is likely to be most relevant in
pathogenic and pathologic conditions associated with those regions.
Support for that notion can be found in studies showing an
increased risk of non-skin cancer in LRRK2 Gly2019Ser mutation
carriers and especially for renal and lung cancer [Mov. Disorder,
25, 2536-2541, 2010]. Over-expression of LRRK2 by chromosomal
amplification has also been identified in papillary renal and
thyroid carcinomas. Also, genetic association of LRRK2 has been
reported for diseases where aberrant responses of the immune system
are involved. This is the case for inflammatory bowel diseases such
as Crohn's disease and ulcerative colitis as well as for leprosy
[Nat Genet. 42, 1118-1125, 2010; Inflamm. Bowel. Dis. 16, 557-558,
2010; N Engl. J Med. 361, 2609-2618, 2009; Inflamm. Bowel. Dis.
doi: 10.1002/ibd.21651, 2011].
[0060] Thus, the compounds, as outlined in formula A hereinabove,
or composition comprising said compounds may be used in treatment
of a disease or disorder characterised by over-expression of LRRK2
or a mutated form of LRRK2 such as G2019S, I2020T, M1646T, G2385R
or A419V.
[0061] These disease or disorder may be a CNS disease selected from
Lewy body dementia or Parkinson's disease, such as idiopathic
Parkinson's disease or sporadic Parkinson's disease or in a
Parkinson disease patient carrying anyone of the above mentioned
LRRK2 mutations, in particular the G2019S mutation.
[0062] In a further embodiment, the compounds, as outlined in
formula A hereinabove, or composition comprising said compounds may
be used in the treatment of cancer or an immune related disorder
characterised by over-expression of LRRK2 or a mutated form of
LRRK2 such as G2019S, I2020T, M1646T, G2385R or A419V.
[0063] The cancer diseases may reside in the brain, lungs, kidney,
spleen or blood organs such as renal cancer, lung cancer, skin
cancer, and papillary renal and thyroid carcinomas.
[0064] The immune related disorder may in one embodiment be Crohn's
disease, ulcerative colitis or leprosy.
[0065] In one embodiment, the compound of the present invention is
administered in an amount from about 0.001 mg/kg body weight to
about 100 mg/kg body weight per day. In particular, daily dosages
may be in the range of 0.01 mg/kg body weight to about 50 mg/kg
body weight per day. The exact dosages will depend upon the
frequency and mode of administration, the sex, the age the weight,
and the general condition of the subject to be treated, the nature
and the severity of the condition to be treated, any concomitant
diseases to be treated, the desired effect of the treatment and
other factors known to those skilled in the art.
[0066] A typical oral dosage for adults will be in the range of
1-1000 mg/day of a compound of the present invention, such as 1-500
mg/day.
[0067] The compounds of the present invention may be administered
alone as a pure compound 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, 21 Edition, Mack
Publishing Co, 2005. In the present context, "excipient",
"carrier", "diluent", "adjuvant" and the like are used synonymously
and are intended to mean the same.
[0068] The pharmaceutical compositions may be specifically
formulated for administration by any suitable route such as the
oral, rectal, nasal, pulmonary, topical (including buccal and
sublingual), transdermal, intracisternal, intraperitoneal, vaginal
and parenteral (including subcutaneous, intramuscular, intrathecal,
intravenous and intradermal) route, the oral route being preferred.
It will be appreciated that the preferred route will depend on the
general condition and age of the subject to be treated, the nature
of the condition to be treated and the active ingredient
chosen.
[0069] Pharmaceutical compositions for oral administration include
solid dosage forms such as capsules, tablets, dragees, pills,
lozenges, powders and granules. Where appropriate, they can be
prepared with coatings.
[0070] Liquid dosage forms for oral administration include
solutions, emulsions, suspensions, syrups and elixirs.
[0071] 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.
[0072] Other suitable administration forms include suppositories,
sprays, ointments, cremes, gels, inhalants, dermal patches,
implants, etc.
[0073] Conveniently, the compounds of the invention are
administered in a unit dosage form containing said compounds in an
amount of about 0.1 to 500 mg, such as 10 mg, 50 mg 100 mg, 150 mg,
200 mg or 250 mg of a compound of the present invention.
[0074] For parenteral administration, solutions of the compound of
the invention in sterile aqueous solution, aqueous propylene
glycol, aqueous vitamin E or sesame or peanut oil may be employed.
Such aqueous solutions should be suitably buffered if necessary and
the liquid diluent first rendered isotonic with sufficient saline
or glucose. The aqueous solutions are particularly suitable for
intravenous, intramuscular, subcutaneous and intraperitoneal
administration. The sterile aqueous media employed are all readily
available by standard techniques known to those skilled in the
art.
[0075] Suitable pharmaceutical carriers include inert solid
diluents or fillers, sterile aqueous solution and various organic
solvents. Examples of solid carriers are lactose, terra alba,
sucrose, cyclodextrin, talc, gelatine, agar, pectin, acacia,
magnesium stearate, stearic acid and lower alkyl ethers of
cellulose. Examples of liquid carriers are syrup, peanut oil, olive
oil, phospho lipids, fatty acids, fatty acid amines,
polyoxyethylene and water. The pharmaceutical compositions formed
by combining the compound of the invention and the pharmaceutical
acceptable carriers are then readily administered in a variety of
dosage forms suitable for the disclosed routes of
administration.
[0076] Formulations of the present invention suitable for oral
administration may be presented as discrete units such as capsules
or tablets, each containing a predetermined amount of the active
ingredient, and which may include a suitable excipient.
Furthermore, the orally available formulations may be in the form
of a powder or granules, a solution or suspension in an aqueous or
non-aqueous liquid, or an oil-in-water or water-in-oil liquid
emulsion.
[0077] If a solid carrier is used for oral administration, the
preparation may be tablet, e.g. placed in a hard gelatine capsule
in powder or pellet form or in the form of a troche or lozenge. The
amount of solid carrier may vary but will usually be from about 25
mg to about 1 g.
[0078] If a liquid carrier is used, the preparation may be in the
form of a syrup, emulsion, soft gelatine capsule or sterile
injectable liquid such as an aqueous or non-aqueous liquid
suspension or solution.
[0079] Tablets may be prepared by mixing the active ingredient with
ordinary adjuvants and/or diluents followed by the compression of
the mixture in a conventional tabletting machine. Examples of
adjuvants or diluents comprise: Corn starch, potato starch, talcum,
magnesium stearate, gelatine, lactose, gums, and the like. Any
other adjuvants or additives usually used for such purposes such as
colourings, flavourings, preservatives etc. may be used provided
that they are compatible with the active ingredients.
[0080] All references, including publications, patent applications,
and patents, cited herein are hereby incorporated by reference in
their entirety and to the same extent as if each reference were
individually and specifically indicated to be incorporated by
reference and were set forth in its entirety herein (to the maximum
extent permitted by law), regardless of any separately provided
incorporation of particular documents made elsewhere herein.
[0081] The use of the terms "a" and "an" and "the" and similar
referents in the context of describing the invention are to be
construed to cover both the singular and the plural, unless
otherwise indicated herein or clearly contradicted by context. For
example, the phrase "the compound" is to be understood as referring
to various "compounds" of the invention or particular described
aspect, unless otherwise indicated.
[0082] The description herein of any aspect or aspect of the
invention using terms such as "comprising", "having," "including,"
or "containing" with reference to an element or elements is
intended to provide support for a similar aspect or aspect of the
invention that "consists of", "consists essentially of", or
"substantially comprises" that particular element or elements,
unless otherwise stated or clearly contradicted by context (e.g., a
composition described herein as comprising a particular element
should be understood as also describing a composition consisting of
that element, unless otherwise stated or clearly contradicted by
context).
[0083] Synthetic Routes
[0084] The compounds of the present invention of the general
formula A herein above, wherein R1 to R6 are as defined above can
be prepared by the methods outlined in the following reaction
schemes and 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.
[0085] Preparation of the Compounds of the Invention
[0086] The products of the invention can be prepared by the
following general methods: [0087] a) Reacting a compound of formula
II or salt thereof with a boronic acid of formula III or a
corresponding boronic acid ester in a suitable solvent such as a
mixture of dioxane and water in the presence of a suitable catalyst
such as tetrakis(triphenylphosphine)palladium and a suitable base
such as or dicesium carbonate or potassium carbonate at a suitable
temperature from 60.degree. C.-150.degree. C. The heating may be
performed in a microwave system.
[0087] ##STR00007## [0088] b) Where R2 and R3 together with the
atom they are attached to form a compound as shown in formula V,
the compounds of the invention can be prepared by reacting a
compound of formula IV with TFA and triethylsilane at a suitable
temperature such as 0.degree. C.
[0088] ##STR00008## [0089] c) Where the R1 group is as shown in
formula VI the compounds of the invention can be prepare by
removing a protecting group (PG) from a compound of formula VI.
This utilises standard chemical transformations known to a person
skilled in the art. This includes treating a compound of formula VI
with where PG is COO.sup.tBu with a suitable acid such as hydrogen
chloride in a suitable solvent such as ether and/or chloroform at a
suitable temperature such as 0.degree. C. to room temperature.
[0089] ##STR00009## [0090] d) Where the R1 group is as shown in
formula IX the compounds of the invention can be prepared by
alkylation of a compound of formula VIII using a suitable base such
as NaH and a suitable alkylating reagent such as MeI in a suitable
solvent such as THF at a suitable temperature such as 0.degree.
C.
[0090] ##STR00010## [0091] e) Where either the R2 or R3 group is
(CO)R6 as shown in formula XV, the compounds of the invention can
be prepared by reduction of the carbonyl. This can be done using a
suitable reducing agent such as BH.sub.3.DMS in a solvent such as
THF at temperatures between 0.degree. C. and 70.degree. C.
[0091] ##STR00011## [0092] f) Compounds of the invention where R2
or R3 is (CO)R6 can be prepared by reacting a compound of formula
XVII with R6H in the presence of an activating agent such as HATU
or EDC together with HOBT in a suitable solvent (e.g. DMF) at a
suitable temperature (such as room temperature (r.t)). The reaction
may take place in the presence of a base (e.g. triethylamine)
[0093] g)
##STR00012##
[0094] Compounds of formula II or salts thereof, where X.dbd.CH and
R1 is an [1,3,4]oxadiazole, can be prepared by reacting a compound
of formula XI with a hydrazide of formula XII in POCl.sub.3 at
suitable temperatures such as between room temperature and
100.degree. C.
[0095] Alternatively, the reaction can be performed by reacting a
compound of formula XI with a hydrazide of formula XII in a
suitable solvent such as DMF in the presence of an activating agent
such as HATU in the presence of a base such as DIPEA at a suitable
temperature such as room temperature. The product of this reaction
can ring close by treatment with PPh.sub.3, DBU and CCl.sub.4 in a
suitable solvent such as acetonitrile at a suitable temperature
such as room temperature.
##STR00013##
[0096] Compounds of formula II or salts thereof where C.dbd.CH and
R1 is an [1,2,3]triazole can be prepared by reaction of a compound
of formula XIII with sodium azide in a suitable solvent such as a
mixture of butanol and water at a suitable temperature such as
80.degree. C. followed by addition of a compound of formula XV and
CuI.
##STR00014##
[0097] Synthesis of compounds of formula IV, VI and XV can be
carried out as described above for a).
[0098] Compounds of formula III where R2 or R3 are (CO)R6 can be
prepared by reacting can be prepared by reacting a compound of
formula XIX with R6H in the presence of an activating agent such as
HATU or EDC together with HOBT in a suitable solvent (e.g. DMF) at
a suitable temperature (such as room temperature (r.t)). The
reaction may take place in the presence of a base (e.g.
triethylamine)
##STR00015##
[0099] Compounds of formula XXI can be prepared by reacting a
compound of formula II or salt hereof by methods described above in
a). Compounds of formula XXI can be hydrolysed using a suitable
base such as LiOH in a solvent such as a mixture of dioxane and
water at a temperature between room temperature and 100.degree. C.
to give compounds of formula XVII.
##STR00016##
[0100] Experimental:
[0101] General Methods:
[0102] .sup.1H NMR spectra were recorded at 400 MHz on a Varian
400MR instrument at T=298.15 K or at 400 MHz on a Varian vnmrs
instrument. Deuterated solvents (CDCl.sub.3, DMSO-d.sub.6,
CD.sub.3OD) were used for preparation. Tetramethylsilane was used
as internal reference standard. Chemical shift values are expressed
in ppm-values relative to tetramethylsilane unless noted
otherwise.
[0103] Method A
[0104] Waters Acquity UPLC-MS [0105] (Bin. Solv. manager, col.
manager, PDA, ELSD, Sample manager+organizer, SQD) [0106] Duration
1.15 min [0107] Column type: Acquity UPLC BEH C18 1.7 .mu.m;
2.1.times.50 mm [0108] Column temperature: 60.degree. C. [0109]
PDA: 254 nm [0110] Desolvation temp.: 350.degree. C. [0111] Ion
source: ESCI+/- [0112] Source temp.: 150.degree. C. [0113] Make-up
flow: 0.5 ml/min. EtOH [0114] Flow: 1.2 ml/min [0115] Solvents:
[0116] A: Water containing 0.1% Formic acid
[0117] B: Acetonitrile containing 5% Water and 0.1% Formic acid
TABLE-US-00001 Gradient: Time, min. % B Curve 0.00 10.0 6 1.00 99.9
6 1.01 10.0 6 1.15 10.0 6
[0118] Method B
[0119] Waters Acquity UPLC-MS [0120] Bin. Solv. manager, col.
manager, PDA, ELSD, Sample manager, TQD) [0121] Duration 1.15 min
[0122] Column type: Acquity UPLC BEH C18 1.7 .mu.m; 2.1.times.50 mm
[0123] Column temperature: 60.degree. C. [0124] PDA: 254 nm [0125]
Probe temp.: 450.degree. C. [0126] Source temp.: 150.degree. C.
[0127] Ion source: APPI pos. [0128] Dopant flow: 0.04 ml/min
Toluene [0129] Flow: 1.2 ml/min [0130] Solvents:
[0131] A: Water containing 0.05% TFA
[0132] B: Acetonitrile containing 5% Water and 0.035% TFA
TABLE-US-00002 Gradient: Time, min. % B Curve 0.00 10.0 6 1.00 100
6 1.01 10.0 6 1.15 10.0 6
[0133] Abbreviations
[0134] RT: room temperature
[0135] BINAP: 2,2'-bis(diphenylphosphino)-1,1'-binaphthyl
[0136] DMF: Dimethylformamide
[0137] DCM: Dichloromethane
[0138] EtOAc: Ethylacetate
[0139] HATU:
N,N,N',N'-tetramethyl-O-(7-azabenzotriazol-1-yl)uronium
hexafluorophosphate
[0140] DIPEA: N,N-Diisopropylethylamine
[0141] DBU: 1,8-Diazabicycloundec-7-ene
[0142] THF: Tetrahydrofuran
[0143] TFA: Trifluoroacetic acid
[0144] ACN: Acetonitrile
[0145] TLC: thin layer chromatography
2-Amino-5-bromo-nicotinic acid
##STR00017##
[0147] To a solution of pyridine-2-amino-3-carboxylic acid (2.05 g,
81.1 mmol) in AcOH (250 mL) was added bromine (11.0 mL, 199.2 mmol)
at RT. The reaction mixture was stirred for 18 h at RT. On
completion of the reaction, the solvent was evaporated completely
under reduced pressure, filtered, washed with diethyl ether
(3.times.75 mL) to afford the title compound as a yellow solid.
[0148] Yield: 50.0 g (92.6%)
[0149] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 11.7 (3H,
br, s), 8.33 (1H, s), 8.20 (1H, s).
5-Bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
##STR00018##
[0151] A solution of 2-amino-5-bromo-nicotinic acid (300 mg, 1.38
mmol) and cyclopropanecarboxylic acid hydrazide (300 mg, 3.0 mmol)
in POCl.sub.3 (30.0 mL,) was stirred for 18 h at 65.degree. C. The
reaction mixture was distilled under reduced pressure to obtain a
residue, which was cooled to 0.degree. C. by adding iced water and
neutralizing with saturated NaHCO.sub.3 solution (100 mL). The
aqueous layer was extracted with ethyl acetate. The ethyl acetate
layer was separated, dried over anhydrous Na.sub.2SO.sub.4 and
evaporated to afford 370 mg of crude
5-bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine.
Crude compound was purified by column chromatography using silica
gel (100-200 mesh). The column was eluted with 15% EtOAc in
petroleum ether to afford the title compound as a pale yellow solid
used without further purification.
[0152] Yield: 300 mg (Yield: 81%)
5-Bromo-3-(5-isopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
##STR00019##
[0154] To a solution of 2-amino-5-bromo-nicotinic acid (1.0 g, 4.60
mmol) in POCl.sub.3 (20 mL) was added isobutyric acid hydrazide
(705 mg, 6.912 mmol) at RT. The reaction temperature was slowly
heated to 100.degree. C. and stirred for 18 h. The solvent was
evaporated under reduced pressure, the residue was basified with
saturated cold sodium bicarbonate solution (50 mL), extracted with
ethyl acetate (3.times.50 mL), organic layer was washed with brine,
dried over anhydrous sodium sulphate and concentrated under reduced
pressure. Crude compound was purified by column chromatography
using 100-200 mesh silica gel. The column was eluted with 40% EtOAc
in petroleum ether to afford the title compound as a yellow
solid.
[0155] Yield: 650 mg (50%)
[0156] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.28-8.27
(1H, d), 8.18-8.17 (1H, d), 7.45 (2H, br, s), 3.32-3.24 (1H, m),
1.38-1.37 (6H, d).
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1-methyl-1H-indol-5-yl)-pyridin-
-2-ylamine
##STR00020##
[0158] To a solution of
5-bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
(500 mg, 1.77 mmol) in dioxane (15.0 mL) and water (5.0 mL) were
added K.sub.2CO.sub.3 (730 mg, 5.31 mmol) and
N-methylindole-5-boronic acid (140 mg, 1.95 mmol) in a sealed tube.
The reaction mixture was degassed with argon for 20 min, then
Pd(PPh.sub.3).sub.4 (100 mg, 0.088 mmol) was added and stirred for
18 h at 100.degree. C. The reaction mixture was diluted with water
(20 mL) and extracted with ethyl acetate (3.times.20 mL). The
organic layer was washed with brine, dried over anhydrous sodium
sulphate and concentrated under reduced pressure to get crude title
compound. Crude product was purified by column chromatography using
100-200 mesh silica gel and compound was eluted with 40% ethyl
acetate in petroleum ether to afford the title compound.
[0159] Yield: 350 mg (61%)
[0160] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.54-8.53
(1H, d), 8.22-8.21 (1H, d), 7.82-7.81 (1H, d), 7.53-7.51 (1H, d),
7.46-7.44 (1H, d), 7.36-7.35(1H, d), 7.25 (2H, broad s, NH.sub.2),
6.48-6.47 (1H, d), 3.82 (3H, s), 2.36-2.33 (m, 1H), 1.20-1.18 (4H,
d).
4-[N'-(2-Amino-5-bromo-pyridine-3-carbonyl)-hydrazinocarbonyl]-piperidine--
1-carboxylic acid tert-butyl ester
##STR00021##
[0162] To a solution of 2-amino-5-bromo-nicotinic acid (1.2 g, 5.53
mmol) in DMF (30 mL) were added
4-hydrazinocarbonyl-piperidine-1-carboxylic acid tert-butyl ester
(1.5 g, 6.173 mmol), HATU (2.35 g, 6.184 mmol) and DIPEA (4 mL,
23.56 mmol) at RT. The reaction mixture was stirred for 18 h at RT.
The reaction mixture was poured into ice-water (30 mL), extracted
with ethyl acetate (3.times.50 mL), organic layer was washed with
brine dried over anhydrous sodium sulphate concentrated under
reduced pressure. Crude compound was purified by column
chromatography using 100-200 mesh silica gel. The column was eluted
with 60% EtOAc in petroleum ether to afford the title compound as
an off-white solid.
[0163] Yield: 1.3 g (53.19%)
[0164] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 10.32 (1H,
s), 9.92 (1H, s), 8.20 (1H, s), 8.12-8.11 (1H, d), 7.2 (2H, s),
3.96-3.94 (2H, m), 2.77-2.69 (2H, m), 2.50-2.41 (1H, m), 1.73-1.70
(2H, m), 1.49-1.44 (2H, m), 1.40 (9H, s).
4-[5-(2-Amino-5-bromo-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1-ca-
rboxylic acid tert-butyl ester
##STR00022##
[0166] To a solution of
4-[N'-(2-amino-5-bromo-pyridine-3-carbonyl)-hydrazinocarbonyl]-piperidine-
-1-carboxylic acid tert-butyl ester (1.3 g, 2.939 mmol) in ACN (50
mL) were added DBU (1.68 g, 11.035 mmol), PPh.sub.3 (1.95 g, 7.434
mmol) and CCl.sub.4 (4 mL, 41.558 mmol) at RT. The reaction mixture
was stirred for 18 h at RT. The reaction mixture was poured into
ice-water (30 mL), extracted with ethyl acetate (3.times.50 mL),
organic layer was washed with brine dried over anhydrous sodium
sulphate concentrated under reduced pressure. Crude compound was
purified by column chromatography using 100-200 mesh silica gel.
The column was eluted with 40% EtOAc in petroleum ether to afford
the title compound as a white solid.
[0167] Yield: 700 mg (56.4%)
[0168] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.28-8.27
(1H, d), 8.21-8.20 (1H, d), 7.44 (2H, br, s), 7.2 (2H, s),
3.95-3.91 (2H, m), 3.31-3.23 (1H, m), 2.99 (2H, br, s), 2.08-2.04
(2H, m), 1.74-1.64 (2H, m), 1.41 (9H, s).
4-{5-[2-Amino-5-(1-methyl-1H-indol-5-yl)-pyridin-3-yl]-[1,3,4]oxadiazol-2--
yl}-piperidine-1-carboxylic acid tert-butyl ester
##STR00023##
[0170] To a solution of
4-[5-(2-amino-5-bromo-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1-c-
arboxylic acid tert-butyl ester (1.1 g, 2.59 mmol) in dioxane (20
mL) and water (10 mL) were added K.sub.2CO.sub.3 (1.07 g, 7.77
mmol) and N-methylindole-5-boronic acid (0.50 g, 2.85 mmol) in a
sealed tube. The reaction mixture was degassed with argon for 20
min, then Pd(PPh.sub.3).sub.4 (0.15 g, 0.13 mmol) was added to the
reaction mixture. The reaction mixture was stirred for 18 h at
100.degree. C. The reaction was diluted with water (100 mL),
extracted with ethyl acetate (2.times.150 mL) and organic layer was
washed with brine, dried over anhydrous sodium sulphate and
concentrated under reduced pressure to get crude compound. Crude
compound was purified by column chromatography using 100-200 mesh
silica gel compound eluted with 60% ethyl acetate in petroleum
ether to afford 900 mg of the title compound which is further
purified by washing with 20% chloroform in hexane to afford the
title compound as a pale yellow solid.
[0171] Yield: 700 mg (56.9%)
[0172] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.56-8.55
(1H, d), 8.28-8.27 (1H, d), 7.82 (1H, s), 7.53-7.51 (1H, d),
7.47-7.44 (1H, d), 7.36-7.35 (1H, d), 7.30-7.26 (2H, broad s),
6.48-6.47 (1H, d), 3.97-3.94 (2H, d), 3.82 (3H, s), 3.30-3.27 (1H,
m), 3.05-2.92 (2H, m), 2.12-2.08 (2H, d), 1.76-1.66 (2H, m), 1.41
(9H, s).
4-[5-(2-Amino-5-p-tolyl-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1--
carboxylic acid tert-butyl ester
##STR00024##
[0174] To a solution of
4-[5-(2-amino-5-bromo-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1-c-
arboxylic acid tert-butyl ester (500 mg, 1.179 mmol) in dioxane (20
mL) and water (10 mL) were added K.sub.2CO.sub.3 (488 mg, 3.537
mmol) and 4-tolyl boronic acid (179 mg, 1.297 mmol) in a sealed
tube. The reaction mixture was degassed with N.sub.2 for 20 min,
then Pd(PPh.sub.3).sub.4 (136 mg, 0.118 mmol) was added to the
reaction mixture at RT. The reaction mixture temperature was raised
to 100.degree. C., stirred for 18 h at 100.degree. C. The reaction
mixture was diluted with water (30 mL) extracted with ethyl acetate
(3.times.30 mL) and organic layer was washed with brine, dried the
organic layer over anhydrous sodium sulphate and concentrated under
reduced pressure to affords the crude product. Crude was purified
by column chromatography using 100-200 mesh silica gel compound
eluted with 35% ethyl acetate in petroleum ether to afford the
title compound as a white solid.
[0175] Yield: 250 mg, (48.8%)
[0176] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.54 (1H,
s), 8.25 (1H, s), 7.59-7.57 (2H, m), 7.36 (2H, br, s), 7.28-7.26
(2H, m), 3.96-3.93 (2H, m), 3.32 (1H, m), 2.998 (2H, br, s), 2.34
(3H, s), 2.10-2.08 (2H, m), 1.75-1.67 (2H, m), 1.41 (9H, s).
3-(5-Piperidin-4-yl-[1,3,4]oxadiazol-2-yl)-5-p-tolyl-pyridin-2-ylamine
##STR00025##
[0178] To a solution of
4-[5-(2-amino-5-p-tolyl-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1-
-carboxylic acid tert-butyl ester (250 mg, 0.574 mmol) in DCM (20
mL) was added 2M HCl in diethyl ether (20 mL) at 0.degree. C. The
reaction mixture was allowed to warm to ambient temperature, and
was stirred for 2 h. After completion of the reaction, reaction
mixture was filtered and washed with diethyl ether (2.times.30 mL),
which afforded the title compound as a yellow solid.
[0179] Yield: 200 mg (93.8%)
[0180] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.80 (1H,
s), 8.57-8.56 (2H, d), 8.28-8.27 (1H, s), 7.59-7.57 (2H, m), 7.49
(1H, br, s), 7.30-7.28 (2H, m), 3.51 (1H, m), 3.37-3.34 (2H, m),
3.17-3.08 (2H, m), 2.35 (3H, s), 2.32-2.28 (2H, m), 2.07-1.98 (2H,
m).
4-{5-[2-Amino-5-(3-methoxy-phenyl)-pyridin-3-yl]-[1,3,4]oxadiazol-2-yl}-pi-
peridine-1-carboxylic acid tert-butyl ester
##STR00026##
[0182] To a solution of
4-[5-(2-amino-5-bromo-pyridin-3-yl)-[1,3,4]oxadiazol-2-yl]-piperidine-1-c-
arboxylic acid tert-butyl ester (1.0 g, 2.35 mmol) in dioxane (20
mL) and water (20 mL) were added K.sub.2CO.sub.3 (0.95 g, 7.05
mmol), 3-methoxyphenyl boronic acid (0.39 g, 2.59 mmol) in a sealed
tube. The reaction mixture was degassed with argon for 20 min, then
Pd(PPh.sub.3).sub.4 (0.13 g, 0.11 mmol) was added to the reaction
mixture. The reaction mixture was stirred for 18 h at 100.degree.
C. The reaction mixture was diluted with water (100 mL) extracted
with ethyl acetate (2.times.150 mL) and organic layer was washed
with brine, dried over anhydrous sodium sulphate and concentrated
under reduced pressure to get crude product. The crude compound was
purified by column chromatography using 100-200 mesh silica gel and
compound was eluted with 40% ethyl acetate in petroleum ether to
afford 500 mg, which was further purified by washing with diethyl
ether and hexane to afford the title compound as a pale yellow
solid.
[0183] Yield: 400 mg (37.7%)
[0184] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.57-8.56
(1H, d), 8.28-8.27 (1H, d), 7.42-7.35 (3H, m), 7.25-7.21 (2H, m),
6.93-6.91 (1H, dd), 3.96-3.93 (2H, d), 3.83 (3H, s), 3.30-3.26 (1H,
m), 3.03-2.95 (2H, m), 2.11-2.08 (2H, d), 1.76-1.65 (2H, m), 1.41
(9H, s).
5-Bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
##STR00027##
[0186] A mixture of isopropyl bromide (5.0 g, 40.65 mmol) and
sodium azide (2.6 g, 40.65 mmol) in t-BuOH/water (1:2) (50 mL) was
stirred for 2 h at 80.degree. C. Then the reaction mixture was
cooled to room temperature, to which was added
5-bromo-3-ethynyl-pyridin-2-ylamine (0.8 g, 4.065 mmol) and CuI
(catalytic amount). The reaction mixture was stirred for 16 h at
80.degree. C. The reaction mixture was cooled to RT, diluted with
EtOAc (100 mL) and washed with water (50 mL). The organic layer was
washed with brine solution (50 mL), dried over anhydrous
Na.sub.2SO.sub.4 and solvent was evaporated under reduced pressure
to afford crude compound. Crude compound was washed with diethyl
ether to afford the title compound as ash colour solid.
[0187] Yield: 450 mg (40.9%)
[0188] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.87 (1H,
s), 8.09-8.08(1H, d), 8.04 (1H, s), 7.15 (2H, br), 4.90-4.83 (1H,
m), 1.55 (3H, s), 1.53 (3H, s).
5-Bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyrazin-2-ylamine
##STR00028##
[0190] A mixture of isopropyl bromide (2.5 g, 20.3 mmol) and sodium
azide (1.3 g, 20.3 mmol) in t-BuOH/water (1:2) (25 mL) was stirred
for 2 h at 80.degree. C. Then the reaction mixture was cooled to
room temperature, was added 5-bromo-3-ethynyl-pyrazin-2-ylamine
(0.40 g, 2.03 mmol) and CuI (catalytic amount) and stirred for 16 h
at 80.degree. C. The reaction mixture was cooled to RT, diluted
with EtOAc (100 mL) and washed with water (50 mL). The organic
layer was washed with brine solution (50 mL), dried over anhydrous
Na.sub.2SO.sub.4 and solvent was evaporated under reduced pressure
to afford crude product. The crude product was washed with diethyl
ether to afford the title compound as a solid.
[0191] Yield: 370 mg (64.7%)
[0192] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.82 (1H,
s), 8.11 (1H, s), 7.55 (2H, s, br), 4.97-4.90 (1H, m), 1.57-1.55
(6H, d).
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-1H-indol-5-yl)-pyrazin--
2-ylamine
##STR00029##
[0194] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyrazin-2-ylamine
(750 mg, 2.65 mmol) and N-methylindole-5-boronic acid (556 mg, 3.18
mmol) in 1,4-dioxane (15.0 mL)/water (5.0 mL) was added
Cs.sub.2CO.sub.3 (1.29 g, 3.97 mmol) at room temperature. The
reaction mixture was purged with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (153 mg, 0.13 mmol) was added and allowed to
stir at 100.degree. C. for 16 h. The reaction mixture was cooled to
RT, diluted with EtOAc (20 mL) and washed with water (20 mL). The
organic layer was washed with brine solution (20 mL), dried over
anhydrous Na.sub.2SO.sub.4 and solvent was evaporated under reduced
pressure to afford crude product. The crude compound was purified
by column using 100-200 mesh silica gel. The column was eluted with
70-80% EtOAc in Hexane to afford the title compound as a yellow
solid.
[0195] Yield: 610 mg (69.1%)
[0196] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.95 (1H,
s), 8.60 (1H, s), 8.27 (1H, d), 7.92-7.90 (1H, dd), 7.50-7.48 (1H,
d), 7.35 (1H, d), 7.30 (2H, br), 6.49-6.48 (1H, d), 5.02-4.95 (1H,
m), 3.82 (3H, s), 1.62-1.60 (6H, d).
2-(4-Iodo-phenyl)-1-morpholin-4-yl-ethanone
##STR00030##
[0198] Triethylamine (5.30 mL, 38.0 mmol), HOBt (3.96 g, 28.5 mmol)
and EDC.HCl (5.47 g, 28.5 mmol) were added to a solution of
(4-iodo-phenyl)-acetic acid (5.0 g, 19.0 mmol) in DMF (100 mL) and
stirred. After 5 min morpholine (2.0 mL, 22.9 mmol) was added at
RT. The reaction mixture was stirred at RT for 16 h. The reaction
mixture was poured into ice-cold water (50 mL) and obtained solid
was filtered, dried to get 3.92 g (Yield: 62.1%) of the title
compound as a white solid.
[0199] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 7.66-7.64
(2H, d), 7.04-7.02 (2H, d), 3.68 (2H, s), 3.54-3.42 (8H, m).
4-[2-(4-Iodo-phenyl)-ethyl]-morpholine
##STR00031##
[0201] BF.sub.3.Et.sub.2O (6.43 g, 45.3 mmol) was added to a
solution of 2-(4-iodo-phenyl)-1-morpholin-4-yl-ethanone (3.0 g,
9.06 mmol) in THF (75 mL) at 0.degree. C., stirred for 30 min, then
NaBH.sub.4 (1.72 g, 45.3 mmol) was added at 0.degree. C. The
reaction mixture was slowly warmed to RT. After 16 h the reaction
mixture was cooled to 0.degree. C., quenched with ice-cold water,
extracted with EtOAc (2.times.100 mL). The organic layer was washed
with brine solution (50 mL), dried over anhydrous Na.sub.2SO.sub.4
and solvent was evaporated under reduced pressure. The crude
compound was purified by column chromatography using 100-200 mesh
silica gel, eluted with 20-30% EtOAc in pet-ether to afford 1.56 g
(Yield: 54.3%) of the title compound as a yellow solid.
[0202] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 7.64-7.62
(2H, d), 7.07-7.05 (2H, d), 4.11-4.07 (2H, q), 3.58 (4H, br, s),
2.67 (2H, br, s), 2.46 (4H, br, s).
4-{2-[4-(4,4,5,5-Tetramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-ethyl}-morp-
holine
##STR00032##
[0204] To a solution of 4-[2-(4-iodo-phenyl)-ethyl]-morpholine (250
mg, 0.78 mmol) and
4,4,5,5,4',4',5',5'-Octamethyl-[2,2']bi[[1,3,2]dioxaborolanyl] (240
mg, 0.94 mmol) in DMSO (15.0 mL) was added KOAc (191 mg, 1.95 mmol)
at RT. N.sub.2 was purged through the reaction mixture for 10 min.
Then Pd(dppf)Cl.sub.2.DCM (31.8 mg, 0.039 mmol) was added and
through the reaction mixture N.sub.2 was purged for 10 min and
stirred at 100.degree. C. for 16 h. Reaction mixture was cooled to
RT, diluted with EtOAc (50 mL) and washed with water (50 mL).
Organic layer was washed with brine solution (50 mL), dried over
anhydrous Na.sub.2SO.sub.4 and solvent was evaporated under reduced
pressure. Crude compound was purified by column chromatography
using 100-200 mesh silica gel and eluted with 40-50% EtOAc in
Pet-ether to afford 110 mg (Yield: 44.0%) of the title compound as
brown semi-solid.
[0205] LCMS Conditions:
[0206] Column: SYMMETRY C18 (4.6.times.75 mm); 3.5 u
[0207] M-Phase A: 5 mM Ammonium Acetate in H.sub.2O
[0208] M-Phase B: ACN
[0209] T/% B: 0/05,2.5/98,8.0/98,8.1/5
[0210] Flow: 0.8 ml/min
[0211] Diluent: ACN+H.sub.2O
[0212] Purity: 93.65%
[0213] r.t.=4.49 min, m/z=318.3[M+H].sup.+
1-Morpholin-4-yl-2-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-pheny-
l]-ethanone
##STR00033##
[0215] To a solution of 2-(4-Iodo-phenyl)-1-morpholin-4-yl-ethanone
(1.0 g, 3.02 mmol) and
4,4,5,5,4',4',5',5'-Octamethyl-[2,2']bi[[1,3,2]dioxaborolanyl]
(0.92 g, 3.62 mmol) in DMSO (20.0 mL) was added KOAc (0.74 mg, 7.55
mmol) at room temperature. N.sub.2 was purged through the reaction
mixture for 10 min. Pd(dppf)Cl.sub.2.DCM (0.12 g, 0.15 mmol) was
added and through the reaction mixture N.sub.2 was purged for 10
min and stirred at 100.degree. C. for 16 h. The reaction mixture
was cooled to RT, diluted with EtOAc (50 mL) and washed with water
(50 mL). The organic layer was washed with brine solution (50 mL),
dried over anhydrous Na.sub.2SO.sub.4 and solvent was evaporated
under reduced pressure. Crude compound was purified by column
chromatography using 100-200 mesh silica gel and eluted with 40-50%
EtOAc in pet-ether to afford 720 mg (yield: 72%) of the title
compound as brown solid.
[0216] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 7.62-7.60
(2H, d), 7.24-7.22 (2H, d), 3.52 (2H, br, s), 3.47-3.45 (8H, br,
m).
(3-Fluoro-4-(morpholine-4-carbonyl)phenyl)boronic acid
##STR00034##
[0218] To a solution of 4-borono-2-fluorobenzoic acid (1.0 g, 5.43
mmol) and HATU (2.0 g, 5.43 mmol) in DMF (20.0 mL) was added
morpholine (0.9 g, 10.87 mmol) at room temperature.
[0219] The reaction mixture was stirred at room temperature for 18
h. The reaction mixture was poured into ice cold water, acidified
with 1N HCl, extracted with EtOAc (2.times.50 mL). The organic
layer was washed with brine solution (20 mL), dried over anhydrous
Na.sub.2SO.sub.4 and solvent was evaporated under reduced pressure
to afford 1.2 g of crude compound as a colourless gum. Crude
compound was directly used for the next step without any further
purification.
4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-methoxyben-
zoic acid
##STR00035##
[0221] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(300 mg, 1.06 mmol) and
(3-methoxy-4-(methoxycarbonyl)phenyl)boronic acid (245 mg, 1.16
mmol) in 1,4-dioxane (10.0 mL)/water (5.0 mL) was added
Cs.sub.2CO.sub.3 (1.0 g, 3.18 mmol) at room temperature. The
reaction mixture was degassed with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (61 mg, 0.05 mmol) was added and allowed to
stir at 120.degree. C. for 30 min in CEM micro wave. The reaction
mixture was cooled to RT, diluted with EtOAc (50 mL) and washed
with water (50 mL). The aqueous layer was neutralized with 1N HCl,
extracted with DCM (2.times.50 mL), washed with brine solution (20
mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent was
evaporated under reduced pressure. The crude compound was washed
with diethyl ether to afford 260 mg (yield: 69.33%) of the title
compound as pale yellow solid.
[0222] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 12.50 (1H,
s), 8.93 (1H, s), 8.44-8.43 (1H, d), 8.26-8.25 (1H, d), 7.74-7.72
(1H, d), 7.36-7.29 (2H, m), 7.24 (2H, s), 4.94-4.87 (1H, m), 3.93
(3H, s), 1.58 (3H, s), 1.57 (3H, s).
[0223] LCMS Conditions:
[0224] Column: BEH C18 (2.1.times.50 mm) 1.7 u
[0225] M-Phase A: 0.05% TFA in Water
[0226] M-Phase B: ACN
[0227] T/% B: 0/3, 0.3/3, 1/35, 2/98, 3.8/98, 3.9/3, 4/3
[0228] Flow: 0.6 ml/min
[0229] Diluent: ACN+H.sub.2O
[0230] Purity: 87.26%
[0231] tR=1.24 min, m/z=354.48 [M+H].sup.+
[5-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-fluoro-phenyl]-morpho-
lino-methanone
##STR00036##
[0233] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.70 mmol) and
(4-fluoro-3-(morpholine-4-carbonyl)phenyl)boronic acid (197 mg,
0.77 mmol) in 1,4-dioxane (10.0 mL)/water (5.0 mL) was added
Cs.sub.2CO.sub.3 (690 mg, 2.12 mmol) at room temperature. The
reaction mixture was degassed with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (40 mg, 0.035 mmol) was added and allowed to
stir at 110.degree. C. for 30 min in CEM micro wave. The reaction
mixture was cooled to RT, diluted with EtOAc (50 mL) and washed
with water (50 mL). The organic layer was washed with brine
solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure. Crude compound was purified
by column using 100-200 mesh silica gel. The column was eluted with
5% MeOH in DCM to afford 210 mg (yield: 72.41%) of title compound
as pale yellow solid.
[0234] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.93 (1H,
s), 8.34-8.33 (1H, d), 8.21-8.20 (1H, d), 7.84-7.80 (1H, m),
7.73-7.70 (1H, m), 7.18 (2H, s), 7.39-7.35 (1H, t), 7.16 (2H, s),
4.91-4.88 (1H, m), 3.67 (4H, br), 3.56 (2H, br), 3.28 (2H, br),
1.58 (3H, s), 1.56 (3H, s).
[0235] LCMS Conditions:
[0236] Column: XBRIDGE-C18(4.6.times.75 mm) 3.5 u
[0237] M-Phase A: 0.1% HCOOH(Aq)
[0238] M-Phase B: 0.1% HCOOH in(ACN)
[0239] T/% B 0/5, 0.5/5, 2/98, 8/98, 8.1/5
[0240] Flow: 1.0 ml/min
[0241] Diluent: ACN+H.sub.2O
[0242] Purity: 98.61%
[0243] tR=1.67 min, m/z=411.2 [M+H].sup.+
4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-methylbenz-
oic acid
##STR00037##
[0245] Cs.sub.2CO.sub.3 (1.03 g, 3.18 mmol) was added to a solution
of 5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(300 mg, 1.06 mmol) and (4-(methoxycarbonyl)-3-methylphenyl)boronic
acid (247 mg, 1.27 mmol) in dioxane:water (10 mL:5 mL), degassed
the reaction mixture for 10 min with nitrogen. Then added
Pd(PPh.sub.3).sub.4 (61 mg, 0.053 mmol), degassed the reaction
mixture for additional 10 min with nitrogen, the reaction mixture
was heated to 120.degree. C. in microwave for 1 h. LiOH (222 mg,
5.30 mmol) was added to the reaction mixture and, heated to
100.degree. C. for 3 h. Then diluted with water (50 mL) and
extracted with ethyl acetate (2.times.30 mL). Aqueous layer was
acidified with 2N HCl, pH was adjusted to 6 and extracted with
ethyl acetate (2.times.30 mL). Dried over Na.sub.2SO.sub.4 and
concentrated under reduced pressure to afford 210 mg of (Yield:
58.65%) the title compound as a white solid.
[0246] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.95 (1H,
s), 8.40-8.39 (1H, d), 8.26 (1H, d), 7.89-7.87 (1H, d), 7.63-7.60
(2H, m), 7.22 (2H, br, s), 4.94-4.87 (1H, m), 2.60 (3H, s),
1.58-1.56 (6H, d, J=6.8 Hz).
[0247] LCMS Conditions:
[0248] Column: BEH C18 (2.1.times.50 mm) 1.7 u)
[0249] M-Phase A: 0.1% HCOOH IN H.sub.2O
[0250] M-Phase B: ACN
[0251] T/% B: 0.0/3, 0.2/3, 1/35, 2/98, 3.85/98, 3.9/3, 4/3
[0252] Flow: 0.6 ml/min
[0253] Diluents: ACN+H.sub.2O
[0254] Purity: 95.76%
[0255] tR=1.17 min, m/z=338.2 [M+H].sup.+
[0256] Compounds of the Invention:
1:
5-(1H-Indol-5-yl)-3-(5-isopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylami-
ne
##STR00038##
[0258] To a solution of
5-bromo-3-(5-isopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
(250 mg, 0.883 mmol) in dioxane (15 mL) and water (7 mL) were added
K.sub.2CO.sub.3 (365 mg, 2.649 mmol), indole-5-boronic acid (156
mg, 0.971 mmol) in a sealed tube. The reaction mixture was degassed
by N.sub.2 for 20 min, then Pd(PPh.sub.3).sub.4 (102 mg, 0.088
mmol) was added and stirred for 18 h at 100.degree. C. Water (20
mL) was added and extracted with ethyl acetate (3.times.20 mL) and
the organic layer was washed with brine, dried over anhydrous
sodium sulphate, and concentrated under reduced pressure. Crude
product was purified by column chromatography using 100-200 mesh
silica gel compound eluted with 40% ethyl acetate in petroleum
ether to afford the title compound as a yellow solid.
[0259] Yield: 200 mg, (71.1%)
[0260] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 11.14 (1H,
br s), 8.55-8.54 (1H, d), 8.25-8.24 (1H, d), 7.81 (1H, s),
7.49-7.47 (1H, d), 7.39-7.27 (4H, m), 6.49 (1H, s), 3.35-3.28 (1H,
m), 1.41-1.39 (6H, d).
[0261] LC-MS: m/z=320.0 (MH.sup.+), t.sub.R=0.53, method B.
2:
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1H-indol-5-yl)-pyridin-2-yla-
mine
##STR00039##
[0263]
5-Bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
(200 mg, 0.71 mmol), 5-indolyl boronic acid (112 mg, 0.78 mmol),
K.sub.2CO.sub.3 (290 mg, 2.13 mmol), 1,4-dioxane (10.0 mL) and
water (4.0 mL) were taken in a sealed tube and degassed with
nitrogen for 15 min Then Pd(PPh.sub.3).sub.4 (41 mg, 0.03 mmol) was
added and stirred for 18 h at 100.degree. C. The reaction mixture
was diluted with ice-cold water and extracted with EtOAc. The
combined extracts were dried over Na.sub.2SO.sub.4 and evaporated
under reduced pressure to get crude
3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1H-indol-5-yl)-pyridin-2-ylami-
ne. Crude compound was purified by column chromatography using
silica gel (100-200 mesh). The column was eluted with 30% EtOAc in
petroleum ether to afford the title compound as a pale yellow
solid.
[0264] Yield: 28 mg (21%)
[0265] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 11.13 (s,
1H), 8.53-8.52 (d), 8.21-8.20 (d, 1H), 7.80 (s, 1H), 7.48-7.46 (d,
1H), 7.38-7.36 (m, 2H), 7.24-7.23 (broad s, 2H, NH.sub.2), 6.48 (s,
1H), 2.38-2.31 (m, 1H), 1.20-1.18(m, 4H).
[0266] LC-MS: m/z=318.2 (MH.sup.+), t.sub.R=0.49, method B.
3:
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1-methyl-2,3-dihydro-1H-indo-
l-5-yl)-pyridin-2-ylamine
##STR00040##
[0268] To a solution of
3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(1-methyl-1H-indol-5-yl)-pyridi-
n-2-ylamine (160 mg, 0.48 mmol) in TFA at 0.degree. C. was added
triethylsilane (0.2 mL, 1.20 mmol) drop-wise over a period of 5
min, and stirred for 2 h at 0.degree. C. The reaction mixture was
diluted with water (20 mL) and neutralized with saturated aqueous
NaHCO.sub.3 solution and extracted with ethyl acetate (3.times.20
mL). The organic layer was washed with brine, dried over anhydrous
sodium sulphate and concentrated under reduced pressure to get
crude product. The crude product was purified by column
chromatography using 100-200 mesh silica gel and compound was
eluted with 40% ethyl acetate in petroleum ether to afford the
title compound.
[0269] Yield: 80 mg (Yield: 50%)
[0270] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.42-8.41
(1H, d), 8.09-8.08 (1H, d), 7.35 (1H, s), 7.32-7.30 (1H, d),
7.20-7.18 (2H, broad s, NH.sub.2), 6.58-6.56 (1H, d), 3.31-3.26
(2H, t), 2.95-2.91 (2H, t), 2.73 (3H, s), 2.36-2.30 (1H, m),
1.19-1.17 (4H, m).
4:
3-[5-(1-Methyl-piperidin-4-yl)-[1,3,4]oxadiazol-2-yl]-5-p-tolyl-pyridin-
-2-ylamine
##STR00041##
[0272] To a solution of
3-(5-piperidin-4-yl-[1,3,4]oxadiazol-2-yl)-5-p-tolyl-pyridin-2-ylamine
(250 mg, 0.746 mmol) in THF (25 mL) was added NaH (55%) (48.8 mg,
1.119 mmol) at 0.degree. C. The reaction mixture was stirred at
0.degree. C. for 1 h. Then MeI (95 mg, 0.671 mmol) was added at
0.degree. C., and the reaction mixture was stirred at 0.degree. C.
for a further 3 h. The reaction mixture was then poured into
ice-water (30 mL) and extracted with ethyl acetate (3.times.20 mL).
The organic layer was washed with brine, dried over anhydrous
sodium sulphate and concentrated under reduced pressure to afford
crude compound. Crude compound was purified by preparative HPLC to
afford the title compound as a pale yellow solid.
[0273] Yield: 15 mg (5.7%).
[0274] Preparative HPLC Conditions:
[0275] Column: X-BRIDGEC8 (250.times.20 mm) 5.0 .mu.m
[0276] Mobile phase A: 0.01M Ammonium Acetate
[0277] Mobile phase B: ACN:MeOH (1:1)
[0278] Time (min)/% B: 0/30, 5/50, 10/50, 15/100
[0279] Flow: 15 mL/min
[0280] Solubility: MeOH+THF
[0281] Loading: 20 mg/injection
[0282] Column Temp .degree. C.: Ambient
[0283] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.54-8.53
(1H, d), 8.25-8.24 (1H, d), 7.59-7.57 (2H, d), 7.37 (2H, br, s),
7.28-7.26 (2H, d), 3.03 (1H, m), 2.83-2.67 (2H, m), 2.49 (3H, s),
2.34 (3H, br, s), 2.23-2.08 (4H, m), 1.86-1.83 (2H, m).
[0284] LC-MS: m/z=350.2 (MH.sup.+), t.sub.R=0.4, method B.
5:
5-(1-Methyl-2,3-dihydro-1H-indol-5-yl)-3-(5-piperidin-4-yl-[1,3,4]oxadi-
azol-2-yl)-pyridin-2-ylamine
##STR00042##
[0286] To a solution of
4-{5-[2-amino-5-(1-methyl-1H-indol-5-yl)-pyridin-3-yl]-[1,3,4]oxadiazol-2-
-yl}-piperidine-1-carboxylic acid tert-butyl ester (300 mg, 0.63
mmol) in TFA (2.0 mL) at 0.degree. C. was added triethylsilane (146
mg, 1.26 mmol) drop-wise over a period of 5 min, and stirred for 2
h at 0.degree. C. The reaction was diluted with water (20 mL),
neutralized with saturated aqueous NaHCO.sub.3 solution and
extracted with ethyl acetate (3.times.20 mL). The organic layer was
washed with brine, dried over anhydrous sodium sulphate and
concentrated under reduced pressure to obtain crude product. Crude
product was purified by washing with diethyl ether and hexane to
afford the title compound as a yellow solid:
[0287] Yield: 150 mg (63.2%)
[0288] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.44-8.43
(1H, d), 8.12-8.11 (1H, d), 7.35 (1H, s), 7.32-7.30 (1H, d),
7.24-7.22 (2H, broad s), 6.58-6.56 (1H, d), 3.31-3.26 (2H, m),
3.17-3.11 (1H, m), 3.04-3.01 (2H, d), 2.96-2.91 (2H, t), 2.73 (3H,
s), 2.67-2.61 (2H, t), 2.03-2.00 (2H, d), 1.75-1.65 (2H, m).
6:
3-(5-Cyclopropyl-[1,3,4]oxadiazol-2-yl)-5-(4-morpholin-4-ylmethyl-pheny-
l)-pyridin-2-ylamine
##STR00043##
[0290] To a solution of
5-bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
(300 mg, 1.06 mmol) in dioxane (10 mL) and water (6 mL) were added
K.sub.2CO.sub.3 (438 mg, 3.18 mmol), 4-(morpholinomethyl)-boronic
acid (259 g, 1.17 mmol) in a sealed tube. The reaction mixture was
degassed with argon for 20 min and then Pd(PPh.sub.3).sub.4 (61.2
mg, 0.05 mmol) was added to the reaction mixture. The reaction
mixture was stirred for 18 h at 100.degree. C. The reaction was
diluted with water (50 mL) extracted with ethyl acetate (2.times.50
mL) and organic layer was washed with brine, dried over anhydrous
sodium sulphate and concentrated under reduced pressure to get
crude product, which was purified by column chromatography using
100-200 mesh silica gel compound eluted with ethyl acetate to
afford 120 mg of the crude product. This was further purified by
washing with 20% chloroform in hexane to afford the title compound
as a brown solid.
[0291] Yield: 28 mg (6.9%)
[0292] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.53-8.52
(1H, s), 8.22-8.21 (1H, s), 7.64-7.62 (2H, d), 7.40-7.35 (4H, m),
3.60-3.56 (4H, m), 3.50-3.49 (2H, m), 2.37-2.30 (5H, m), 1.20-1.18
(4H, d).
[0293] LC-MS: m/z=378.2 (MH.sup.+), t.sub.R=0.32, method A.
7:
5-(3-Methoxy-phenyl)-3-(5-piperidin-4-yl-[1,3,4]oxadiazol-2-yl)-pyridin-
-2-ylamine
##STR00044##
[0295] To a solution of
4-{5-[2-amino-5-(3-methoxy-phenyl)-pyridin-3-yl]-[1,3,4]oxadiazol-2-yl}-p-
iperidine-1-carboxylic acid tert-butyl ester (200 mg, 0.44 mmol) in
chloroform (10 mL) was added 2M HCl in diethyl ether (10 mL) at
0.degree. C., and the reaction mixture was stirred for 4 h at
0.degree. C. After completion of the reaction, solvents in reaction
mixture were evaporated under vacuum to get crude product. Crude
product was purified by washing with ethyl acetate and hexane to
afford the title compound as a pale yellow powder.
[0296] Yield: 130 mg (75.0%)
[0297] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 9.02-8.98
(1H, broad s), 8.78-8.74 (1H, broad s), 8.62-8.61 (1H, d),
8.36-8.35 (1H, d), 7.42-7.38 (1H, t), 7.26-7.23 (2H, m), 6.96-6.94
(1H, dd), 3.83 (3H, s), 3.49-3.43 (1H, m), 3.43-3.36 (2H, d),
3.13-3.04 (2H, m), 2.33-2.28 (2H, dd), 2.09-1.99 (2H, m).
[0298] LC-MS: m/z=352.15 (MH.sup.+), t.sub.R=0.34, method A.
8:
5-(3-Methoxy-phenyl)-3-[5-(1-methyl-piperidin-4-yl)-[1,3,4]oxadiazol-2--
yl]-pyridin-2-ylamine
##STR00045##
[0300] To a solution of
5-(3-methoxyphenyl)-3-[5-(4-piperidyl)-1,3,4-oxadiazol-2-yl]pyridin-2-ami-
ne (150 mg, 0.42 mmol) in THF (20 mL) was added 60% NaH (25.0 mg,
064 mmol) at 0.degree. C. and the reaction mixture was stirred at
0.degree. C. for 10 min. Then MeI (54.5 mg, 0.38 mmol) was added at
0.degree. C. and stirred the reaction mixture at 0.degree. C. for 2
h. The reaction mixture was poured into ice-water (30 mL),
extracted with ethyl acetate (3.times.20 mL), organic layer was
washed with brine, dried over anhydrous sodium sulphate and
concentrated under reduced pressure. Crude product was purified by
preparative TLC to afford the title compound as a yellow solid.
[0301] Yield: 30 mg (Yield: 19.3%)
[0302] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.59-8.58
(1H, d), 8.27-8.26 (1H, d), 7.42-7.36 (3H, m), 7.24-7.20 (2H, m),
6.94-6.91 (1H, dd), 3.83 (3H, s), 3.45-3.36 (3H, m), 3.11-2.96 (2H,
m), 2.70 (3H, s), 2.35-2.28 (2H, m), 2.16-1.98 (2H, m).
[0303] LC-MS: m/z=366.1 (MH.sup.+), t.sub.R=0.35, method A.
9:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-1H-indol-5-yl)-pyrid-
in-2-ylamine
##STR00046##
[0305] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.708 mmol) and N-methylindole-5-boronic acid (124 mg,
0.708 mmol) in 1,4-dioxane (6.0 mL)/water (3.0 mL) was added
K.sub.2CO.sub.3 (293 mg, 2.126 mmol) at room temperature. The
reaction mixture was purged with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (41 mg, 0.035 mmol) was added and the mixture
was allowed to stir at 100.degree. C. for 16 h. The reaction
mixture was cooled to RT, diluted with EtOAc (20 mL) and washed
with water (20 mL). The organic layer was washed with brine
solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure to afford crude compound.
Crude compound was purified by column using 100-200 mesh silica
gel. The column was eluted with 80% EtOAc in Pet. Ether to afford
the title compound as pale yellow solid.
[0306] Yield: 160 mg (69.5%)
[0307] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.96 (1H,
s), 8.31-8.30 (1H, d), 8.21-8.20 (1H, d), 7.82 (1H, s), 7.51-7.45
(2H, m), 7.34-7.33 (1H, d), 6.96 (2H, br), 6.46-6.45 (1H, d),
4.94-4.85 (1H, m), 1.58 (3H, s), 1.56 (3H, s).
[0308] LC-MS: m/z=333.15 (MH.sup.+), t.sub.R=0.57, method A.
10:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indo-
l-5-yl)-pyridin-2-ylamine
##STR00047##
[0310] To a solution of
3-(1-isopropyltriazol-4-yl)-5-(1-methylindol-5-yl)pyridin-2-amine
(170 mg, 0.511 mmol) in TFA (5.0 mL) was added triethylsilane (0.16
mL, 1.022 mmol) at 0.degree. C. The reaction temperature was
stirred for 1 h at 0.degree. C. The reaction mixture was poured
into ice-cold water (20 mL), basified with saturated NaHCO.sub.3,
extracted with EtOAc (30 mL), dried over anhydrous Na.sub.2SO.sub.4
and evaporated the solvent under reduced pressure to get crude
compound. Crude compound was washed with diethyl ether to afford
title compound as a yellow solid.
[0311] Yield: 80 mg (yield: 46.7%)
[0312] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.91 (1H,
s), 8.19 (1H, s), 8.08 (1H, s), 7.37 (1H, s), 7.33-7.31 (1H, d),
6.91 (2H, br), 6.57-6.55 (1H, d), 4.92-4.85 (1H, m), 3.31-3.26 (2H,
m), 2.94-2.91 (2H, t), 2.72 (3H, s), 1.57 (3H, s), 1.56 (3H,
s).
[0313] LC-MS: m/z=335 (MH.sup.+), t.sub.R=0.48, method A.
11:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-2,3-dihydro-1H-indo-
l-5-yl)-pyrazin-2-ylamine
##STR00048##
[0315] To a solution of
3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-5-(1-methyl-1H-indol-5-yl)-pyrazin-
-2-ylamine (200 mg, 0.60 mmol) in TFA (10.0 mL) was added
triethylsilane (0.19 mL, 1.20 mmol) at 0.degree. C. The reaction
temperature was stirred for 2 h at 0.degree. C. The reaction
mixture was poured into ice-cold water (20 mL), basified with
saturated NaHCO.sub.3, extracted with EtOAc (30 mL), dried over
anhydrous Na.sub.2SO.sub.4 and evaporated the solvent under reduced
pressure to get crude compound. Crude compound was washed with
diethyl ether to afford the title compound as a brown solid.
[0316] Yield: 125 mg (62.1%)
[0317] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.88 (1H,
s), 8.45 (1H, s), 7.78-7.45 (2H, m), 7.22 (2H, br), 6.56-6.54 (1H,
d), 5.00-4.93 (1H, m), 3.32-3.29 (2H, m), 2.97-2.93 (2H, t), 2.75
(3H, s), 1.60-1.58 (1H, d).
12:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-ylmethyl-pheny-
l)-pyridin-2-ylamine
##STR00049##
[0319] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(500 mg, 1.77 mmol) and
4-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-benzyl]-morpholine
(537 mg, 1.77 mmol) in ethylene glycol dimethyl ether (6.0
mL)/water (4.0 mL) was added Cs.sub.2CO.sub.3 (1.15 g, 3.54 mmol)
at room temperature. The reaction mixture was purged with argon for
30 min. Then Pd(dppf)Cl.sub.2.DCM (72.2 mg, 0.08 mmol) was added
and allowed to stir at 140.degree. C. for 6 h in microwave. After 6
h, the reaction mixture was cooled to RT, diluted with EtOAc (50
mL) and washed with water (50 mL). The organic layer was washed
with brine solution (50 mL), dried over anhydrous Na.sub.2SO.sub.4
and solvent was evaporated under reduced pressure to afford crude
product. The crude compound was purified by column using 100-200
mesh silica gel. The column was eluted with 5-95% MeOH in DCM to
afford the title compound as a brown solid.
[0320] Yield: 72 mg (10.7%)
[0321] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.94 (1H,
s), 8.31 (1H, s), 8.20 (1H, d), 7.65-7.63 (2H, d), 7.38-7.36 (2H,
d), 7.10 (2H, br s,), 4.93-4.86 (1H, m), 3.58 (4H, s), 3.49 (2H,
s), 2.37 (4H, m), 1.57-1.56 (6H, d).
[0322] LC-MS: m/z=379.1 (MH.sup.+), t.sub.R=0.3, method A.
13:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-(4-morpholin-4-yl-phenyl)-pyr-
idin-2-ylamine
##STR00050##
[0324] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(100 mg, 0.35 mmol) and
4-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-morpholine
(88 mg, 0.42 mmol) in 1,4-dioxane (7.0 mL)/water (3.0 mL) was added
Cs.sub.2CO.sub.3 (227 mg, 0.70 mmol) at room temperature. The
reaction mixture was purged with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (20.2 mg, 0.017 mmol) was added and allowed to
stir at 100.degree. C. for 16 h. The reaction mixture was cooled to
RT, diluted with EtOAc (50 mL) and washed with water (50 mL). The
organic layer was washed with brine solution (50 mL), dried over
anhydrous Na.sub.2SO.sub.4 and solvent was evaporated under reduced
pressure to afford crude compound. Crude compound was purified by
column using 100-200 mesh silica gel. The column was eluted with
3-97% MeOH in DCM to afford the title compound as a yellow
solid.
[0325] Yield: 45 mg (34.8%)
[0326] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.92 (1H,
s), 8.25 (1H, d), 8.13 (1H, d), 7.57-7.54 (2H, d), 7.02-7.00 (2H,
d), 6.97 (2H, br s,), 4.92-4.85 (1H, m), 3.77-3.74 (4H, t),
3.15-3.12 (4H, t), 1.57-1.56 (6H, d).
[0327] LC-MS: m/z=365.2 (MH.sup.+), t.sub.R=0.47, method A.
14:
3-(1-Isopropyl-1H-[1,2,3]triazol-4-yl)-5-[4-(2-morpholin-4-yl-ethyl)-p-
henyl]-pyridin-2-ylamine
##STR00051##
[0329] To a solution of
4-{2-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-ethyl}-mor-
pholine (can be prepared as described in US2002026052) (101 mg,
0.31 mmol) and
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(90 mg, 0.31 mmol) in 1,4-dioxane (7.0 mL)/water (3.0 mL) was added
Cs.sub.2CO.sub.3 (251 mg, 0.70 mmol) at RT. N.sub.2 was purged
through the reaction mixture for 10 min. Pd(PPh.sub.3).sub.4 (18
mg, 0.015 mmol) was added and through the reaction mixture N.sub.2
was purged for 10 min and stirred at 100.degree. C. for 16 h. The
reaction mixture was cooled to RT, diluted with EtOAc (50 mL) and
washed with water (50 mL). The organic layer was washed with brine
solution (50 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure to afford crude compound
(GVK-B1319-120A1). Crude compound was purified by column using
100-200 mesh silica gel and eluted with 2-3% MeOH in DCM to afford
65 mg of a brown semisolid. Further purification of Prep-HPLC
furnished 14 mg of the title compound as an off-white solid.
[0330] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.93 (1H,
s), 8.29 (1H, d), 8.18 (1H, d), 7.60-7.58 (2H, d), 7.31-7.29 (2H,
d), 7.06 (2H, br s), 4.92-4.86 (1H, m), 3.58 (4H, s), 2.78-2.75
(2H, t), 2.44 (6H, br, s), 1.57-1.56 (6H, d).
[0331] LC-MS: m/z=393.2 (MH.sup.+), t.sub.R=0.32, method A.
15:
5-(3-fluoro-4-(morpholinomethyl)phenyl)-3-(1-isopropyl-1H-1,2,3-triazo-
l-4-yl)pyridin-2-amine
##STR00052##
[0333] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.70 mmol) and
4-(2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)morphol-
ine (250 mg, 0.77 mmol) in 1,4-dioxane (10.0 mL)/water (5.0 mL) was
added Cs.sub.2CO.sub.3 (690 mg, 2.12 mmol) at room temperature. The
reaction mixture was degassed with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (40 mg, 0.03 mmol) was added and allowed to
stir at 110.degree. C. for 30 min in CEM micro wave. The reaction
mixture was cooled to RT, diluted with EtOAc (50 mL) and washed
with water (50 mL). The organic layer was washed with brine
solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure. The crude compound was
purified by column using 100-200 mesh silica gel. The column was
eluted with 80% EtOAc in hexanes to afford 100 mg (yield: 35.71%)
of the title compound as pale yellow solid.
[0334] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.95 (1H,
s), 8.37-8.36 (1H, d), 8.24-8.23 (1H, d), 7.55-7.52 (2H, m),
7.46-7.42 (1H, t), 7.19 (2H, s), 4.93-4.86 (1H, m), 3.58-3.56 (4H,
m), 3.54 (2H, s), 2.40 (4H, br), 1.58 (3H, s), 1.56 (3H, s).
[0335] LC-MS: m/z=397.5 (MH.sup.+), t.sub.R=0.33, method A.
16:
(4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-fluor-
ophenyl)(morpholino)methanone
##STR00053##
[0337] To a solution of
(3-fluoro-4-(morpholine-4-carbonyl)phenyl)boronic acid (269 mg,
1.06 mmol) and of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(250 mg, 0.886 mmol) in 1,4-dioxane (10.0 mL)/water (5.0 mL) was
added Cs.sub.2CO.sub.3 (863 mg, 2.65 mmol) at room temperature. The
reaction mixture was degassed with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (51 mg, 0.04 mmol) was added and allowed to
stir at 120.degree. C. for 30 min in CEM micro wave. The reaction
mixture was cooled to RT, diluted with EtOAc (50 mL) and washed
with water (50 mL). The organic layer was washed with brine
solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure to afford crude compound.
Crude compound was purified by column using 100-200 mesh silica
gel. The column was eluted with 5% MeOH in DCM to afford 125 mg
(yield: 34.43%) of the title compound as pale yellow solid.
[0338] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.96 (1H,
s), 8.43-8.42 (1H, d), 8.296-8.29 (1H, d), 7.70-7.64 (2H, m),
7.49-7.45 (1H, t), 7.28 (2H, s), 4.92-4.89 (1H, m), 3.66 (4H, br),
3.55 (2H, br), 3.29 (2H, br), 1.58 (3H, s), 1.57 (3H, s).
[0339] LC-MS: m/z=411.1 (MH.sup.+), t.sub.R=0.41, method A.
17:
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methoxy-phenyl]-m-
orpholino-methanone
##STR00054##
[0341] To a solution of
4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-methoxybe-
nzoic acid (260 mg, 0.735 mmol) and HATU (418 mg, 1.10 mmol) in DMF
(10.0 mL) were added morpholine (77 mg, 0.88 mmol) and DIPEA (0.4
mL, 2.205 mmol) at room temperature. The reaction mixture was
stirred at room temperature for 18 h. The reaction mixture was
poured into ice cold water, extracted with EtOAc (100 mL). The
organic layer was washed with brine solution (50 mL), dried over
anhydrous Na.sub.2SO.sub.4 and solvent was evaporated under reduced
pressure. Crude compound was purified by column using 100-200 mesh
silica gel. The column was eluted with 100% EtOAc to afford 250 mg
(yield: 80.64%) of the title compound as pale yellow solid.
[0342] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.91 (1H,
s), 8.39-8.38 (1H, d, J=2.4 Hz), 8.226-8.22 (1H, d, J=2.4 Hz),
7.32-7.30 (2H, m), 7.26-7.24 (1H, m), 7.12 (2H, s), 4.93-4.87 (1H,
m), 3.91 (3H, s), 3.63 (4H, br), 3.52 (2H, br), 3.17-3.16 (2H, br),
1.58 (3H, s), 1.56 (3H, s).
[0343] LC-MS: m/z=423.1 (MH.sup.+), t.sub.R=0.41, method A.
18:
3-(1-isopropyltriazol-4-yl)-5-[3-methoxy-4-(morpholinomethyl)phenyl]py-
ridin-2-amine
##STR00055##
[0345] To a solution of
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methoxy-phenyl]-morp-
holino-methanone (200 mg, 0.473 mmol) in THF (10.0 mL) was added
BF.sub.3.Et.sub.2O (0.3 mL, 2.36 mmol) at 0.degree. C. and stirred
for 30 min. After 30 min, NaBH.sub.4 (89 mg, 2.36 mmol) was added
at 0.degree. C. and stirred at room temperature for 18 h. After
cooling to 0.degree. C., quenched with MeOH (2 mL) and heated to
reflux for 6 h. The reaction mixture was diluted with EtOAc (50
mL), washed with water (30 mL), dried over anhydrous
Na.sub.2SO.sub.4 and solvent was evaporated under reduced pressure.
Crude compound was purified by column using 100-200 mesh silica
gel. The column was eluted with 100% EtOAc to afford 100 mg (yield:
51.81%) of the title compound as white solid.
[0346] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.90 (1H,
s), 8.34-8.33 (1H, d), 8.18-8.17 (1H, d), 7.36-7.34 (1H, d),
7.22-7.21 (2H, m), 7.08 (2H, s), 4.91-4.88 (1H, m), 3.88 (3H, s),
3.59-3.57 (4H, m), 3.48 (2H, s), 2.39 (4H, br), 1.58 (3H, s), 1.56
(3H, s).
[0347] LC-MS: m/z=409.1 (MH.sup.+), t.sub.R=0.33, method A.
19: 5-[4-fluoro
-3-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyridin-2-amine
##STR00056##
[0349] To a solution of
[5-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-fluoro-phenyl]-morph-
olino-methanone (150 mg, 0.365 mmol) in THF (10.0 mL) was added
BF.sub.3.Et.sub.2O (0.25 mL, 1.827 mmol) at 0.degree. C. and
stirred for 30 min. After 30 min, NaBH.sub.4 (69 mg, 1.827 mmol)
was added at 0.degree. C. and stirred at room temperature for 18 h.
After cooling to 0.degree. C., quenched with MeOH (2 mL) and heated
to reflux for 6 h. The reaction mixture was diluted with EtOAc (20
mL), washed with water (20 mL), dried over anhydrous
Na.sub.2SO.sub.4 and solvent was evaporated under reduced pressure.
Crude compound was purified by column using 100-200 mesh silica
gel. The column was eluted with 5% MeOH in DCM to afford 60 mg
(yield: 41.66%) of the title compound as white solid.
[0350] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.90 (1H,
s), 8.278-8.273 (1H, d), 8.16-8.15 (1H, d), 7.66-7.62 (2H, m),
7.28-7.23 (1H, t), 7.09 (2H, s), 4.91-4.88 (1H, m), 3.57 (6H, br),
2.42 (4H, br), 1.58 (3H, s), 1.56 (3H, s).
[0351] LC-MS: m/z=397.1 (MH.sup.+), t.sub.R=0.33, method A.
20:
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methyl-phenyl]-mo-
rpholino-methanone
##STR00057##
[0353] HATU (336 mg, 0.88 mmol) and DIPEA (0.30 mL, 1.77 mmol) were
added to a solution of
4-(6-amino-5-(1-isopropyl-1H-1,2,3-triazol-4-yl)pyridin-3-yl)-2-methylben-
zoic acid (200 mg, 0.59 mmol) in DMF (10.0 mL) at room temperature,
after 5 min, morpholine (0.06 mL, 0.71 mmol) was added, stirred at
RT for 18 h. The reaction mixture was poured into ice cold water,
extracted with EtOAc (2.times.50 mL). The organic layer was washed
with brine solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4
and solvent was evaporated under reduced pressure. Crude product
was purified by washing with ether to afford 148 mg of (Yield:
61.4%) of the title compound as a white solid.
[0354] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.94 (1H,
s), 8.34-8.33 (1H, d), 8.22-8.21 (1H, d), 7.60 (1H, s), 7.57-7.55
(1H, d), 7.25-7.23 (1H, d, J=8.0 Hz), 7.14 (2H, br, s), 4.93-4.86
(1H, m), 3.66 (4H, s), 3.51 (2H, br, s), 3.18 (2H, br, s), 2.30
(3H, s), 1.58-1.56 (6H, d, J=6.8 Hz).
[0355] LC-MS: m/z=407.1 (MH.sup.+), t.sub.R=0.42, method A.
21:
3-(1-Isopropyltriazol-4-yl)-5-[3-methyl-4-(morpholinomethyl)phenyl]pyr-
idin-2-amine
##STR00058##
[0357] BF.sub.3.Et.sub.2O in THF (47-49%) (0.87 mL, 2.46 mmol) was
added to a solution of
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-methyl-phenyl]-morph-
olino-methanone (200 mg, 0.46 mmol) in THF (10.0 mL) at 0.degree.
C. The reaction mixture was stirred for 30 min, NaBH.sub.4 (93.5
mg, 2.46 mmol) was added at 0.degree. C., reaction mixture was
slowly warmed to RT, stirred for 16 h. Reaction mixture was again
cooled to 0.degree. C., added MeOH (5.0 mL) and heated to
70.degree. C. for 2 h. The reaction mixture was diluted with water
(50 mL) and extracted with ethyl acetate (2.times.30 mL). Combined
organic layers was dried over Na.sub.2SO.sub.4 and concentrated
under reduced pressure. Crude product was purified by silica
(100-200 mesh) column eluted with 2% MeOH in DCM. Product was
further purified by Prep.HPLC furnished 30 mg (Yield: 15.5%) of the
title compound as a white solid.
[0358] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.91 (1H,
s), 8.29-8.28 (1H, d), 8.17-8.16 (1H, d), 7.47 (1H, s), 8.44-8.42
(1H, d), 7.29-7.27 (1H, d), 7.04 (2H, br, s), 4.91-4.87 (1H, m),
3.57-3.55 (4H, t), 3.45 (2H, s), 2.40 (3H, s), 2.38-2.37 (4H, t),
1.58-1.56 (6H, d).
[0359] LC-MS: m/z=393.1 (MH.sup.+), t.sub.R=0.33, method A.
22:
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-chloro-phenyl]-mo-
rpholino-methanone
##STR00059##
[0361] Cs.sub.2CO.sub.3 (682 mg, 2.10 mmol) was added to a solution
of 5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.70 mmol) and
(3-chloro-4-(morpholine-4-carbonyl)phenyl)boronic acid (229 mg,
0.85 mmol) in dioxane:water (7 mL:3 mL) degassed the reaction
mixture for 10 min with nitrogen. Then added Pd(PPh.sub.3).sub.4
(40 mg, 0.035 mmol), degassed the reaction mixture for additional
10 min with nitrogen, the reaction mixture was heated to
120.degree. C. in microwave for 1 h. The reaction mixture was
diluted with water (50 mL) and extracted with ethyl acetate
(3.times.30 mL). Combined organic layers was dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. Crude
product was purified by silica (100-200 mesh) column eluted with
70-80% ethyl acetate in pet-ether to afford 68 mg (Yield: 22.5%) of
the title compound as a pale yellow solid. Product was converted to
its HCl salt by using HCl in ether (2M) to afford 60 mg of the HCl
salt of the title compound as a pale yellow solid.
[0362] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 9.17 (1H,
s), 8.70 (1H, s), 8.50 (1H, d), 8.42-8.41 (2H, br), 7.99 (1H, d),
7.85-7.83 (1H, dd), 7.55-7.53 (1H, dd), 4.99-4.92 (1H, m),
3.71-3.64 (4H, br, s), 3.57-3.55 (2H, t), 3.18-3.16 (2H, t),
1.60-1.58 (6H, d).
[0363] LC-MS: m/z=427.4 (MH.sup.+), t.sub.R=0.44, method A.
23:
5-[3-Chloro-4-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyr-
idin-2-amine
##STR00060##
[0365] BH.sub.3.DMS in THF (2M) (1.17 mL, 2.34 mmol) was added to a
solution of
[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-2-chloro-phenyl]-morph-
olino-methanone (200 mg, 0.46 mmol) in THF (10.0 mL) at 0.degree.
C., then reaction mixture was heated to 70.degree. C. for 4 h.
Reaction mixture was cooled to 0.degree. C., added MeOH (5.0 mL) to
the reaction mixture, heated to 70.degree. C. for 6 h. The reaction
mixture was diluted with water (50 mL) and extracted with ethyl
acetate (2.times.30 mL). Combined organic layers was dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. Crude
product was purified by silica (100-200 mesh) column eluted with 2%
MeOH in DCM to afford 131 mg (Yield: 67.7%) of the title compound
as a white solid.
[0366] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.94 (1H,
s), 8.35-8.34 (1H, d), 8.22 (1H, d), 7.76 (1H, d), 7.66-7.64 (1H,
dd), 7.54-7.52 (1H, d), 7.18 (2H, br, s), 4.93-4.86 (1H, m), 3.59
(6H, s), 2.44 (4H, s), 1.58-1.56 (6H, d).
[0367] LCMS Conditions:
[0368] Column: BEH C18 (2.1.times.50 mm) 1.7 u
[0369] M-Phase A: 5 mM Ammonium Acetate in H.sub.2O
[0370] M-Phase B: ACN
[0371] T/% B: 0/03, 1.5/45, 2.5/45, 3.2/95, 4.7/95, 5/03
[0372] Flow: 0.4 ml/min
[0373] Diluent: MeOH
[0374] Purity: 95.01%
[0375] tR=2.90 min, m/z=413.2 [M+H].sup.+
24:
3-(1-Isopropyltriazol-4-yl)-5-[3-(morpholinomethyl)phenyl]pyridin-2-am-
ine
##STR00061##
[0377] Cs.sub.2CO.sub.3 (858 mg, 2.64 mmol) was added to a solution
of (3-(morpholinomethyl)phenyl)boronic acid (250 mg, 0.88 mmol) and
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(215 mg, 0.97 mmol) in dioxane:water (8 mL:4 mL) degassed the
reaction mixture for 10 min with nitrogen. Then added
Pd(PPh.sub.3).sub.4 (50 mg, 0.044 mmol), degassed the reaction
mixture for additional 10 min with nitrogen, the reaction mixture
was heated to 120.degree. C. in microwave for 1 h. The reaction
mixture was diluted with water (50 mL) and extracted with ethyl
acetate (2.times.30 mL). Combined organic layers was dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. Crude
product was purified by silica (100-200 mesh) column eluted with
80-90% ethyl acetate in pet-ether to afford 195 mg (Yield: 58.13%)
of the title compound as a pale yellow solid. Product was converted
to its HCl salt by using HCl in ether (2M) to afford 202 mg as a
pale yellow solid.
[0378] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 11.60 (1H,
br, s), 9.51 (1H, s), 9.04 (1H, s), 8.54 (3H, br, s), 8.37 (1H, s),
7.89-7.88 (1H, d), 7.60-7.52 (2H, m), 4.99-4.92 (1H, m), 4.44 (2H,
s), 3.93 (4H, s), 3.23 (2H, s), 3.16 (2H, br, s), 1.60-1.58 (6H,
d).
[0379] LC-MS: m/z=379.5 (MH.sup.+), t.sub.R=0.36, method A.
25:
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morpholino-m-
ethanone
##STR00062##
[0381] Cs.sub.2CO.sub.3 (858 mg, 2.64 mmol) was added to a solution
of 5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(250 mg, 0.88 mmol) and (3-(morpholine-4-carbonyl)phenyl)boronic
acid (228 mg, 0.97 mmol) in dioxane:water (8 mL:4 mL) degassed the
reaction mixture for 10 min with nitrogen. Then added
Pd(PPh.sub.3).sub.4 (50 mg, 0.044 mmol), degassed the reaction
mixture for additional 10 min with nitrogen, the reaction mixture
was heated to 120.degree. C. in microwave for 1 h. After completion
of reaction, the reaction mixture was diluted with water (50 mL)
and extracted with ethyl acetate (2.times.30 mL). Combined organic
layers was dried over Na.sub.2SO.sub.4 and concentrated under
reduced pressure. Crude product was purified by silica (100-200
mesh) column eluted with 80-90% ethyl acetate in pet-ether to
afford 210 mg (Yield: 60.34%) of the title compound as a pale
yellow solid. Product was converted to its HCl salt by using HCl in
ether (2M) to afford 208 mg as an off-white solid.
[0382] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 9.21 (1H,
s), 8.73 (1H, s), 8.48 (3H, br, s), 7.89-7.87 (1H, d), 7.82 (1H,
s), 7.61-7.57 (1H, t), 7.46-7.44 (1H, d), 4.99-4.92 (1H, m),
3.66-3.59 (8H, br), 1.59-1.58 (6H, d).
[0383] LC-MS: m/z=393.5 (MH.sup.+), t.sub.R=0.41, method A.
26:
[3-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-5-chloro-phenyl]-mo-
rpholino-methanone
##STR00063##
[0385] Cs.sub.2CO.sub.3 (682 mg, 2.10 mmol) was added to a solution
of 5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.70 mmol) and
(3-chloro-5-(morpholine-4-carbonyl)phenyl)boronic acid (210 mg,
0.78 mmol) in dioxane:water (8 mL:4 mL) degassed the reaction
mixture for 10 min with nitrogen. Then added Pd(PPh.sub.3).sub.4
(40 mg, 0.035 mmol), degassed the reaction mixture for additional
10 min with nitrogen, the reaction mixture was heated to
120.degree. C. in microwave for 1 h. The reaction mixture was
diluted with water (50 mL) and extracted with ethyl acetate
(2.times.30 mL). Combined organic layers was dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. Crude
product was purified by silica (100-200 mesh) column eluted with
80-90% ethyl acetate in pet-ether to afford 46 mg (Yield: 47.0%) of
the title compound as a pale yellow solid.
[0386] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.96 (1H,
s), 8.41-8.40 (1H, d), 8.26-8.25 (1H, d), 7.87 (1H, s), 7.68 (1H,
s), 7.38 (1H, s), 7.28 (2H, s), 4.94-4.87 (1H, m), 3.64-3.57 (6H,
br), 3.36 (2H, br), 1.58-1.57 (6H, d).
[0387] LC-MS: m/z=427.4 (MH.sup.+), t.sub.R=0.48, method A.
27:
5-[3-Chloro-5-(morpholinomethyl)phenyl]-3-(1-isopropyltriazol-4-yl)pyr-
idin-2-amine
##STR00064##
[0389] BF.sub.3.Et.sub.2O in THF (47-49%) (0.70 mL, 2.34 mmol) was
added to a solution of
[3-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]-5-chloro-phenyl]-morph-
olino-methanone (200 mg, 0.46 mmol) in THF (10.0 mL) at 0.degree.
C., then reaction mixture was stirred for 30 min, added NaBH.sub.4
(89 mg, 2.36 mmol) at 0.degree. C., reaction mixture was slowly
warmed to RT, stirred for 16 h. Reaction mixture was cooled to
0.degree. C., added MeOH (5.0 mL) to the reaction mixture, heated
to 70.degree. C. for 2 h. The reaction mixture was diluted with
water (50 mL) and extracted with ethyl acetate (2.times.30 mL).
Combined organic layers was dried over Na.sub.2SO.sub.4 and
concentrated under reduced pressure. Crude product was purified by
silica (100-200 mesh) column eluted with 2% MeOH in DCM to afford
39 mg (Yield: 20.2%) of the title compound. This was converted to
its HCl salt by using HCl in ether (2M) to afford 45 mg as a pale
gray solid.
[0390] .sup.1HNMR (D.sub.2O, 400 MHz, TMS) .delta.: 8.63 (1H, s),
8.46-8.45 (1H, d), 8.18 (1H, d), 7.83 (1H, s), 7.68 (1H, s), 7.63
(1H, s), 5.03-4.96 (1H, m), 4.45 (2H, s), 4.15-3.82 (4H, br), 3.41
(4H, br), 1.65-1.64 (6H, d).
[0391] LC-MS: m/z=413.1 (MH.sup.+), t.sub.R=0.4, method A.
28:
[4-[6-Amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-morpholino-m-
ethanone
##STR00065##
[0393] To a solution of
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(200 mg, 0.70 mmol) and (4-(morpholine-4-carbonyl)phenyl)boronic
acid (183 mg, 0.77 mmol) in 1,4-dioxane (10.0 mL)/water (5.0 mL)
was added Cs.sub.2CO.sub.3 (690 mg, 2.12 mmol) at room temperature.
The reaction mixture was degassed with argon for 30 min. Then
Pd(PPh.sub.3).sub.4 (40 mg, 0.03 mmol) was added and allowed to
stir at 110.degree. C. for 30 min in CEM micro wave. The reaction
mixture was cooled to RT, diluted with EtOAc (50 mL) and washed
with water (50 mL). The organic layer was washed with brine
solution (20 mL), dried over anhydrous Na.sub.2SO.sub.4 and solvent
was evaporated under reduced pressure to afford crude compound.
Crude compound was purified by column using 100-200 mesh silica
gel. The column was eluted with 5% MeOH in DCM to afford 100 mg
(yield: 35.97%) of the title compound as pale yellow solid.
[0394] .sup.1H NMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.95 (1H,
s), 8.378-8.373 (1H, d), 8.266-8.261 (1H, d), 7.78-7.76 (2H, d),
7.49-7.47 (2H, d), 7.18 (2H, s), 4.93-4.87 (1H, m), 3.61 (8H, br),
1.58 (3H, s), 1.56 (3H, s).
[0395] LC-MS: m/z=393.5 (MH.sup.+), t.sub.R=0.4, method A.
29:
3-(5-cyclopropyl-1,3,4-oxadiazol-2-yl)-5-[3-(morpholinomethyl)phenyl]p-
yridin-2-amine
##STR00066##
[0397] To a solution of
5-bromo-3-(5-cyclopropyl-[1,3,4]oxadiazol-2-yl)-pyridin-2-ylamine
(400 mg; 1.42 mmol) in dioxane (10 mL) and water (6 mL) were added
K.sub.2CO.sub.3 (587 mg; 4.26 mmol),
(3-(morpholinomethyl)phenyl)boronic acid (348 mg; 1.56 mmol) in a
sealed tube. The reaction mixture was degassed with argon for 30
min, then added Pd(PPh.sub.3).sub.4 (82.0 mg; 0.07 mmol) to the
reaction mixture. The reaction mixture was stirred for 18 h at
100.degree. C. The reaction mixture was diluted with water (50 mL)
extracted with ethyl acetate (2.times.50 mL) and organic layer was
washed with brine, dried over anhydrous sodium sulphate and
concentrated under reduced pressure. Crude compound was purified by
column chromatography using 100-200 mesh silica gel and eluted with
100% ethyl acetate, which is further purified by washing with 30%
chloroform in hexane to afford 90 mg (Yield: 16.7%) of the title
compound, as a yellow colour solid.
[0398] .sup.1HNMR (DMSO-d.sub.6, 400 MHz, TMS) .delta.: 8.52-8.51
(1H, d), 8.20-8.19 (1H, d), 7.58-7.54 (2H, m), 7.43-7.39 (1H, t),
7.35 (2H, s), 7.31-7.28 (1H, d), 3.59-3.54 (6H, m), 2.40-2.30 (5H,
m), 1.20-1.15 (4H, m).
[0399] LCMS Conditions:
[0400] Column: BEH C 18 (2.1.times.50 mm) 1.7 u
[0401] M-Phase A: 5 mM NH.sub.4OAC in H.sub.20
[0402] M-Phase B: ACN
[0403] T/% B: 0/03, 1.5/45, 2.5/45, 3.2/95, 4.7/95, 5/03
[0404] Flow: 0.4 ml/min
[0405] Diluent: MeOH
[0406] Drift Tube Temp: 55.degree. C.
[0407] Gas Pressure: 30 psi
[0408] Nebulizer Temp: 65%
[0409] Gain: 500
[0410] Purity: 97.21%
[0411] tR=2.10 min, m/z=378.2[M+H].sup.+
30:
2-[4-[6-amino-5-(1-isopropyltriazol-4-yl)-3-pyridyl]phenyl]-1-morpholi-
no-ethanone
##STR00067##
[0413] To a solution of
1-Morpholin-4-yl-2-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-phen-
yl]-ethanone (129 mg, 0.39 mmol) and
5-bromo-3-(1-isopropyl-1H-[1,2,3]triazol-4-yl)-pyridin-2-ylamine
(100 mg, 0.35 mmol) in 1,4-dioxane (6.0 mL)/water (4.0 mL) was
added Cs.sub.2CO.sub.3 (284 mg, 0.87 mmol) at room temperature.
N.sub.2 was purged through the reaction mixture for 10 min
Pd(PPh.sub.3).sub.4 (20 mg, 0.017 mmol) was added and through the
reaction mixture N.sub.2 was purged for 10 mM and stirred at
100.degree. C. for 16 h. The reaction mixture was cooled to RT,
diluted with EtOAc (50 mL) and washed with water (50 mL). The
organic layer was washed with brine solution (50 mL), dried over
anhydrous Na.sub.2SO.sub.4 and solvent was evaporated under reduced
pressure. Crude compound was purified by column chromatography
using 100-200 mesh silica gel and eluted with 2-3% MeOH in DCM to
afford 40 mg (yield: 27.7%) of the title compound as brown
semisolid. Product was converted to its HCl salt by using 2M HCl in
ether to afford 42 mg of an off-white solid.
[0414] .sup.1H NMR (DMSO-d6, 400 MHz, TMS) .delta.: 9.20 (1H, s),
8.71-8.70 (1H, d), 8.40-8.39 (1H, d), 7.73-7.71 (2H, d), 7.38-7.36
(2H, d), 4.99-4.92 (1H, m), 3.79 (2H, s), 3.56-3.45 (8H, m),
1.59-1.58 (6H, d).
[0415] LC-MS: m/z=407.1 (MH.sup.+), t.sub.R=0.42, method A.
[0416] LRRK2 wild-type and G2019S kinase activity assay.
[0417] LRRK2 kinase activity is measured using a LanthaScreen
kinase activity assay available from Invitrogen (Life Technologies
Corporation). The assay is a homogeneous time resolved-fluorescence
resonance energy transfer (TR-FRET) assay that measures
phosphorylation of a fluorescein-labelled peptide substrate
(flouorescein-LRRKtide, Fluorescein-GAGRLGRDKYKTLRQIRQ) (SEQ ID
NO:1) as a result of LRRK2 kinase activity. The phosphorylated
peptide is recognized by a terbium-labelled phospho-specific
anti-LRRKtide antibody and, subsequently, the phosphorylated
LRRKtide can be quantified by the extent of TR-FRET between the
terbium donor and fluorescein acceptor.
[0418] The LRRK2 kinase is obtained from Invitrogen (Life
Technologies Corporation) and comprises residue 970 to 2527 of the
full length human wildtype LRRK2 kinase or a similar sequence with
the G2019S mutation. As discussed above, this mutation increases
the kinase activity relative to the wild type. The kinase reactions
are performed in a 20 .mu.L volume in 384-well plates. The kinase
reaction buffer consists of 50 mM Tris pH 8.5, 0.01% BRIJ-35, 10 mM
MgCl2, 1 mM EGTA, and 2 mM DTT.
[0419] In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is
incubated with the test compound (typically at 0 to 30 .mu.M) for
30 minutes before the kinase reaction is initiated by addition of
1.3 mM ATP and 0.4 .mu.M fluorescein-LRRKtide. The reaction mixture
(20 .mu.l total volume) is incubated for 2 hours at 30.degree. C.
before the reaction is terminated by addition of 10 mM EDTA and 1
nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20
.mu.l). The mixture is further incubated for 30 minutes at RT.
TR-FRET is measured by excitation of the terbium-donor with 340 nm
light and subsequent (delay time 100 .mu.s) measurement of terbium
and fluorescein emission at 495 nm and 520 nm, respectively, over a
time window of 1000 .mu.s. The measurement is repeated 10 times for
fluorescein and 10 times for terbium emission with a 2000 .mu.s
time window between repeats. TR-FRET measurements are performed on
a Biomek Synergy plate. The TR-FRET signal is calculated as the
emission-ratio at 520 nm over 495 nm
[0420] The TR-FRET ratio readout for test compounds is normalized
to 0% inhibition corresponding to TR-FRET ratio measured in control
wells with no inhibition of the kinase activity and 100% inhibition
corresponding to TR-FRET ratio measured in control wells with no
kinase. Test compound potency (IC50) was estimated by nonlinear
regression using the sigmoidal dose-response (variable slope) using
Xlfit 4 (IDBS, Guildford, Surrey, UK, model 205).
y=(A+((B-A)/(1+((C/x) D))))
where y is the normalized TR-TRET ratio measurement for a given
concentration of test compound, x is the concentration of test
compound, A is the estimated efficacy (% inhibition) at infinite
compound dilution, and B is the maximal efficacy (% inhibition). C
is the IC50 value and D is the Hill slope coefficient. IC.sub.50
estimates were obtained from independent experiment and the
logarithmic average was calculated.
[0421] The table below shows the IC.sub.50 values in nM obtained as
described above for the exemplified compounds
TABLE-US-00003 LRRK2 G2019S LRRK2 WT Example no: IC50 (nM) IC50
(nM) 1 910 2 1000 570 3 710 970 4 860 3600 5 380 2200 6 390 490 7
1000 3300 8 880 2200 9 540 550 10 350 620 11 340 570 12 160 300 13
280 340 14 340 600 15 370 200 16 510 580 17 95 91 18 86 93 19 880
1000 20 110 99 21 240 260 22 290 290 23 220 320 24 500 370 25 950
650 26 690 490 27 760 610 28 430 370 29 760 900 30 520 670
Sequence CWU 1
1
1118PRTArtificial SequenceFluorescein-Labelled Peptide Substrate
(Flouorescein-LRRKtide) 1Gly Ala Gly Arg Leu Gly Arg Asp Lys Tyr
Lys Thr Leu Arg Gln Ile 1 5 10 15 Arg Gln
* * * * *