U.S. patent application number 14/422093 was filed with the patent office on 2016-01-14 for vegfr3 inhibitors.
This patent application is currently assigned to CANCER THERAPEUTICS CRC PTY LIMITED. The applicant listed for this patent is CANCER THERAPEUTICS CRC PTY LIMITED. Invention is credited to Michelle Ang Camerino, Richard Charles Foitzik, Danny Ganame, Andrew John Harvey, Catherine Fae Hemley, Ian Peter Holmes, Wilhelmus Johannes Antonius Kersten, Romina Lessene, Gillian Elizabeth Lunniss, Benjamin Joseph Morrow, Paul Anthony Stupple.
Application Number | 20160009686 14/422093 |
Document ID | / |
Family ID | 50101126 |
Filed Date | 2016-01-14 |
United States Patent
Application |
20160009686 |
Kind Code |
A1 |
Foitzik; Richard Charles ;
et al. |
January 14, 2016 |
VEGFR3 INHIBITORS
Abstract
This invention relates to a compound of the formula (I): The
invention also relates to processes for the preparation of the
compound of the formula (I), pharmaceutical agents or compositions
containing the compound or a method of using the compound for the
treatment of proliferative diseases, such as cancer, as well as the
treatment of diseases ameliorated by the control and/or inhibition
of lymphangiogenesis. ##STR00001##
Inventors: |
Foitzik; Richard Charles;
(Parkville, AU) ; Morrow; Benjamin Joseph;
(Parkville, AU) ; Hemley; Catherine Fae;
(Parkville, AU) ; Lunniss; Gillian Elizabeth;
(Parkville, AU) ; Camerino; Michelle Ang;
(Parkville, AU) ; Ganame; Danny; (Bundoora,
AU) ; Stupple; Paul Anthony; (Bundoora, AU) ;
Lessene; Romina; (Bundoora, AU) ; Kersten; Wilhelmus
Johannes Antonius; (Bundoora, AU) ; Harvey; Andrew
John; (Thebarton, AU) ; Holmes; Ian Peter;
(Parkville, AU) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
CANCER THERAPEUTICS CRC PTY LIMITED |
Bundoora |
|
AU |
|
|
Assignee: |
CANCER THERAPEUTICS CRC PTY
LIMITED
Bundoora
AU
|
Family ID: |
50101126 |
Appl. No.: |
14/422093 |
Filed: |
August 16, 2013 |
PCT Filed: |
August 16, 2013 |
PCT NO: |
PCT/AU2013/000913 |
371 Date: |
February 17, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
61684202 |
Aug 17, 2012 |
|
|
|
Current U.S.
Class: |
514/210.2 ;
435/375; 514/235.8; 514/252.02; 514/275; 544/122; 544/238; 544/296;
544/331; 544/332 |
Current CPC
Class: |
A61P 35/04 20180101;
A61K 31/505 20130101; C07D 413/12 20130101; C07D 401/14 20130101;
C07D 403/12 20130101; C07D 239/42 20130101; C07D 403/14 20130101;
A61P 35/02 20180101; A61P 35/00 20180101; A61K 31/506 20130101;
C07D 401/12 20130101; A61P 43/00 20180101 |
International
Class: |
C07D 401/12 20060101
C07D401/12; C07D 239/42 20060101 C07D239/42; C07D 403/14 20060101
C07D403/14; C07D 403/12 20060101 C07D403/12; C07D 401/14 20060101
C07D401/14 |
Claims
1-78. (canceled)
79. A compound of the formula (I), or a stereoisomer, salt,
solvate, protected form or prodrug thereof: ##STR00204## wherein: A
is an optionally substituted phenyl or an optionally substituted
5-10 membered heteroaryl group linked to the NH group through an
aromatic ring carbon atom, in which the heteroaryl ring system
contains 1 to 4 heteroatoms selected from the group consisting of
N, O and S, and; when A is an optionally substituted phenyl, A may
bear a substituent R.sup.1A which is not alpha to the NH group and
may optionally further bear one or two substituents R.sup.1B which
are not alpha to the NH group, where R.sup.1A is selected from the
group consisting of: (i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where
R.sup.C1 is H or C.sub.1-2 alkyl, Z.sup.1 is selected from the
group consisting of H, C.sub.1-3 alkyl optionally substituted by
OH, C(.dbd.O)OC.sub.1-4 alkyl and C(.dbd.O)Me, and Z.sup.3 is H, or
Z.sup.1 and Z.sup.3 together with the N to which they are attached
form a 4-6 membered heterocycle containing at least one N and
optionally one O; (ii) XNHZ.sup.2, where X is selected from the
group consisting of CMe.sub.2, cyclopropylidene, cyclobutylidene,
cyclopentylidene and oxetanylidine, and Z.sup.2 is selected from
the group consisting of H, C.sub.1-3 alkyl optionally substituted
by OH, C(.dbd.O)OC.sub.1-3 alkyl and C(.dbd.O)Me; and (iii) a group
selected from the group consisting of: ##STR00205## ##STR00206##
wherein: R.sup.N1 is H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl, or
C(.dbd.O)Me; R.sup.N2 is H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl
or C(.dbd.O)Me; R.sup.N3 is H, C.sub.1-4 alkyl, C.sub.3-4
cycloalkyl or C(.dbd.O)Me; R.sup.N4 is H or CH.sub.3; R.sup.N5 is
H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl or C(.dbd.O)Me; R.sup.N6
is H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl or C(.dbd.O)Me;
R.sup.N7 and R.sup.N8 are each independently H or CH.sub.3;
R.sup.N9 is H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl or
C(.dbd.O)Me; R.sup.N10 is H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl
or C(.dbd.O)Me; R.sup.N11 is H, C.sub.1-4 alkyl, C.sub.3-4
cycloalkyl or C(.dbd.O)Me; and R.sup.N12 is H, C.sub.1-4 alkyl,
C.sub.3-4 cycloalkyl or C(.dbd.O)Me; and where each R.sup.1B is
independently selected from the group consisting of C.sub.1-3
alkyl, CF.sub.3, F, Cl, O--(C.sub.1-3 alkyl), and CN; when A is an
optionally substituted 5-10 membered heteroaryl group, A may bear a
single substituent R.sup.1A which is not alpha to the NH group,
where R.sup.1A is as defined above, and may optionally further bear
one, two or three substituents R.sup.1C, where each R.sup.1C is
independently selected from the group consisting of: (i) C.sub.1-3
alkyl optionally substituted with one to three substituents
selected from the group consisting of F, OH and O--(C.sub.1-3
alkyl); (ii) F; (iii) Cl; (iv) O--(C.sub.1-3 alkyl); (v) CN;
(vi)=O; and (vii) C(.dbd.O) C.sub.1-3 alkyl; R.sup.2 is selected
from the group consisting of H, halo, C.sub.1-4 alkyl, CF.sub.3,
CF.sub.2H, CN and O--(C.sub.1-3 alkyl); R.sup.3 is a substituted
phenyl or a substituted 6-membered heteroaryl group, where the
heteroaryl contains 1 or 2 N heteroatoms, and R.sup.3 bears a
substituent R.sup.4 either alpha or beta to the --C.sub.2H.sub.4--
group, and may additionally bear further substituents selected from
the group consisting of F, methyl and CF.sub.3; and R.sup.4 is
--Y--C(O)N(R.sup.N13)Z.sup.4, where Y is selected from the group
consisting of --CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--,
--C(CH.sub.3).sub.2-- and C.sub.3-5 cycloalkylidene; R.sup.N13 is H
or CH.sub.3; and Z.sup.4 is H, CH.sub.3 or OCH.sub.3.
80. A compound according to claim 79, with the proviso it is not:
##STR00207##
81. A compound according to claim 79, with the proviso that: when
R.sup.3 is selected from the group consisting of: ##STR00208##
R.sup.4 is --CH(CH.sub.3)C(O)N(R.sup.N13)Z.sup.4; A is either:
##STR00209## where R.sup.1A is selected from the group consisting
of ##STR00210## and R.sup.N1 is H, C.sub.1-3 alkyl, or C(.dbd.O)Me;
R.sup.N2 is H, C.sub.1-3 alkyl, or C(.dbd.O)Me; R.sup.N3 is
selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N4 is
selected from H and CH.sub.3; R.sup.N5 is selected from H,
C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N7 and R.sup.N8 are
independently selected from H and CH.sub.3; R.sup.N9 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N10 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; Or A is (b) ##STR00211##
where R.sup.1A is ##STR00212## and R.sup.N1 is H, C.sub.1-3 alkyl,
or C(.dbd.O)Me; R.sup.N2 is H, C.sub.1-3 alkyl, or C(.dbd.O)Me;
then R.sup.2 is not CF.sub.3, halo, CF.sub.2H or CN.
82. A compound according to claim 79, wherein when A is phenyl, Y
is --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2-- or C.sub.3-5
cycloalkylidene.
83. A compound according to claim 79, wherein A is optionally
substituted phenyl.
84. A compound according to claim 83, wherein A is selected from
the group consisting of: ##STR00213##
85. A compound according to claim 79, wherein A is an optionally
substituted 6-membered heteroaryl group.
86. A compound according to claim 85, wherein A is optionally
substituted pyridyl.
87. A compound according to claim 86, wherein A is selected from
the group consisting of: ##STR00214##
88. A compound according to claim 87, wherein A is ##STR00215##
89. A compound according to claim 79, wherein A is an optionally
substituted 5-membered heteroaryl group.
90. A compound according to claim 89, wherein A is optionally
substituted pyrazolyl.
91. A compound according to claim 90, wherein A is selected from
the group consisting of: ##STR00216##
92. A compound according to claim 91, wherein A is ##STR00217##
93. A compound according to claim 79, wherein A is selected from
the group consisting of: ##STR00218##
94. A compound according to claim 79, wherein R.sup.2 is H, F, Cl,
or C.sub.1-4 alkyl, CF.sub.3, CF.sub.2H, CN or methoxy.
95. A compound according to claim 94, wherein R.sup.2 is Cl, methyl
or CF.sub.3.
96. A compound according to claim 79, wherein R.sup.3 has the
structure: ##STR00219## where R.sup.6, R.sup.7 and R.sup.8 are each
independently H, F, methyl or CF.sub.3; and one of R.sup.4A and
R.sup.4B is R.sup.4, and the other of R.sup.4A and R.sup.4B is H,
F, methyl or CF.sub.3.
97. A compound according to claim 79, wherein R.sup.3 is a
substituted 6-membered heteroaryl group, where the heteroaryl
contains 1 or 2 N heteroatoms.
98. A compound according to claim 97, wherein R.sup.3 is selected
from the group consisting of: ##STR00220## where R.sup.6, R.sup.7
and R.sup.8 (if present) are each independently H, F, methyl or
CF.sub.3; one of R.sup.4A and R.sup.4B (if present) is R.sup.4, and
the other is H, F, methyl or CF.sub.3.
99. A compound according to claim 79, wherein R.sup.N13 is H or
Me.
100. A compound according to claim 79, wherein Z.sup.4 is H, Me or
OMe.
101. A compound according to claim 79, wherein Y is selected from
the group consisting of --CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--,
--C(CH.sub.3).sub.2-- and C.sub.3-5 cycloalkylidene.
102. A compound according to claim 79, wherein A is an optionally
substituted phenyl or an optionally substituted 5- or 6-membered
heteroaryl group linked to the NH group through an aromatic ring
carbon atom, in which the heteroaryl contains 1 or 2 heteroatoms
selected from N and O, and; when A is optionally substituted
phenyl, A may bear a substituent R.sup.1A which is not alpha to the
NH group, R.sup.1A is selected from the group consisting of: (i)
CH(R.sup.C1)NZ.sup.1Z.sup.3, where R.sup.C1 is H or C.sub.1-2
alkyl, Z.sup.1 is selected from the group consisting of H,
C.sub.1-3 alkyl substituted by C(.dbd.O)OC.sub.1-4 alkyl and
C(.dbd.O)Me, and Z.sup.3 is H, or Z.sup.1 and Z.sup.3 together with
the N to which they are attached form a 4-6 membered heterocycle
containing one N and optionally one O; and (ii) a group selected
from the group consisting of: ##STR00221## wherein: R.sup.N2 is H,
C.sub.1-3 alkyl or C(.dbd.O)Me; R.sup.N5 is H or C.sub.1-4 alkyl;
and R.sup.N6 is H; when A is an optionally substituted 5- or
6-membered heteroaryl group, A may bear a single substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is as
defined above, and may optionally further bear one or two
substituents R.sup.1C, where each R.sup.1C is independently
selected from the group consisting of: (i) C.sub.1-3 alkyl
optionally substituted with one to three substituents selected from
the group consisting of F, OH and O--(C.sub.1-3 alkyl); (ii)
O--(C.sub.1-3 alkyl); (iii) CN; and (iv) C(.dbd.O)Me; R.sup.2 is
halo, CH.sub.3 or CF.sub.3; R.sup.3 is substituted phenyl, where
R.sup.3 bears a substituent R.sup.4 either alpha or beta to the
--C.sub.2H.sub.4-- group, and may additionally bear a further
substituent F; and R.sup.4 is --Y--C(O)NH.sub.2, where Y is
selected from the group consisting of --CHCH.sub.3--,
--CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and C.sub.3-4
cycloalkylidene.
103. A compound according to claim 79, wherein A is selected from
optionally substituted phenyl and an optionally substituted 5- or
6-membered heteroaryl group linked to the NH group through an
aromatic ring carbon atom, in which the heteroaryl contains 1 or 2
heteroatoms selected from N and O, and; when A is optionally
substituted phenyl, A may bear a substituent R.sup.1A which is not
alpha to the NH group, where R.sup.1A is selected from the group
consisting of: (i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where R.sup.C1 is H
or C.sub.1-2 alkyl, Z.sup.1 is selected from the group consisting
of H, C.sub.1-3 alkyl substituted by C(.dbd.O)OC.sub.1-4 alkyl, and
C(.dbd.O)Me, and Z.sup.3 is H; (ii) a group selected from the group
consisting of: ##STR00222## wherein: R.sup.N2 is H, C.sub.1-3 alkyl
or C(.dbd.O)Me; R.sup.N5 is H or C.sub.1 alkyl; and R.sup.N6 is H;
when A is an optionally substituted 5- or 6-membered heteroaryl
group, A may bear a single substituent R.sup.1A which is not alpha
to the NH group, where R.sup.1A is as defined above, and may
optionally further bear one or two substituents R.sup.1C, where
each R.sup.1C is independently selected from the group consisting
of: (i) C.sub.1-3 alkyl optionally substituted with one to three
substituents independently selected from the group consisting of F,
OH and O--(C.sub.1-3 alkyl); (ii) O--(C.sub.1-3 alkyl); (iii) CN;
and (iv) C(.dbd.O)Me; R.sup.2 is halo, CH.sub.3 or CF.sub.3;
R.sup.3 is substituted phenyl, where R.sup.3 bears a substituent
R.sup.4 either alpha or beta to the --C.sub.2H.sub.4-- group, and
may additionally bear a further substituent F; and R.sup.4 is
--Y--C(O)NH.sub.2, where Y is selected from the group consisting of
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
and C.sub.3-4 cycloalkylidene.
104. A compound according to claim 79, wherein A is selected from
substituted phenyl and an optionally substituted 5- or 6-membered
heteroaryl group linked to the NH group through an aromatic ring
carbon atom, in which the heteroaryl contains 1 or 2 heteroatoms
selected from N and O, and; when A is optionally substituted
phenyl, A may bear a substituent R.sup.1A which is not alpha to the
NH group, where R.sup.1A is CH(Me)NH.sub.2, or a group selected
from the group consisting of: ##STR00223## wherein: R.sup.N2 is H
or methyl; R.sup.N5 is H or methyl; and R.sup.N6 is H; when A is an
optionally substituted 5- or 6-membered heteroaryl group, A may
bear a single substituent R.sup.1A which is not alpha to the NH
group, where R.sup.1A is as defined above, and may optionally
further bear one or two substituents R.sup.1C, where each R.sup.1C
is CH.sub.3 or CF.sub.3; R.sup.2 is halo or CF.sub.3; R.sup.3 is
substituted phenyl, where R.sup.3 bears a substituent R.sup.4
either alpha or beta to the --C.sub.2H.sub.4-- group, and may
additionally bear a further substituent F; and R.sup.4 is
--Y--C(O)NH.sub.2, where Y is selected from the group consisting of
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
and C.sub.3 cycloalkylidene.
105. A compound of claim 79, wherein A is selected from substituted
phenyl and an optionally substituted pyrazolyl or pyridyl group
linked to the NH group through an aromatic ring carbon atom; when A
is optionally substituted phenyl, A may bear a substituent R.sup.1A
which is not alpha to the NH group, where R.sup.1A is ##STR00224##
and R.sup.N2 is methyl; when A is an optionally substituted
pyrazolyl or pyridyl group, A may bear a single substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is as
defined above, and may optionally further bear one or two
substituents R.sup.1C, where each R.sup.1C is independently
CH.sub.3 or CF.sub.3; R.sup.2 is Cl or CF.sub.3; R.sup.3 is
substituted phenyl, where R.sup.3 bears a substituent R.sup.4 alpha
to the --C.sub.2H.sub.4-- group, and may additionally bear a
further substituent F; and R.sup.4 is --Y--C(O)NH.sub.2, where Y is
--CHCH.sub.3-- or C.sub.3 cycloalkylidene.
106. A compound according to claim 79, having a structure of
formula (Ie) or a stereoisomer, salt, solvate, protected form or
prodrug thereof: ##STR00225## wherein: A is optionally substituted
phenyl or optionally substituted pyridyl, wherein A may bear one
substituent R.sup.1A which is not alpha to the NH group, where
R.sup.1A is ##STR00226## wherein: R.sup.N2 is H or methyl; R.sup.N6
is H or methyl; R.sup.2 is Cl, methyl or CF.sub.3; and R.sup.4 is
--Y--C(O)NH.sub.2, where Y is selected from the group consisting of
--CHCH.sub.3--, --C(CH.sub.3).sub.2--, cyclopropylidene and
cyclobutylidene; with the proviso that: when A is either:
##STR00227## and R.sup.2 is Cl or CF.sub.3, then R.sup.4 is not
--CH(CH.sub.3)C(O)NH.sub.2.
107. A compound according to claim 79, having a structure of
formula (If) or a stereoisomer, salt, solvate, protected form, or
prodrug thereof: ##STR00228## wherein: A is optionally substituted
phenyl or optionally substituted pyridyl, A may bear one
substituent R.sup.1A which is not alpha to the NH group, R.sup.1A
is ##STR00229## wherein: R.sup.N2 is H or methyl; R.sup.N6 is H or
methyl; R.sup.2 is Cl, methyl or CF.sub.3; and R.sup.4 is
--Y--C(O)NH.sub.2, where Y is --C(CH.sub.3).sub.2--,
cyclopropylidene or cyclobutylidene.
108. A compound according to claim 79, having a structure of
formula (Ig) or a stereoisomer, salt, solvate, protected form, or
prodrug thereof: ##STR00230## wherein A is an optionally
substituted 5- or 6-membered heteroaryl group which contains 1 or 2
heteroatoms selected from N and O, A may bear one substituent
R.sup.1A which is not alpha to the NH group, R.sup.1A is selected
from the group consisting of R.sup.1A2, R.sup.1A3 and R.sup.1A6:
##STR00231## wherein: R.sup.N2 is H or C.sub.1-4 alkyl; R.sup.N3 is
H, C.sub.1-3 alkyl, or C(.dbd.O)Me; and R.sup.N6 is H or
C.sub.1-4alkyl; and A may optionally further bear one substituent
R.sup.1C selected from the group consisting of CF.sub.3,
C.sub.1-3alkyl, CH.sub.2CF.sub.3, CN, C(.dbd.O)(C.sub.1-3alkyl),
CH(CH.sub.3)NH.sub.2, CHCF.sub.2, OCH.sub.3,
CH(CH.sub.3)NHCH.sub.3, C.sub.1-3alkylOH and C.sub.1-3alkylOMe;
R.sup.2 is halo, C.sub.1-4alkyl or CF.sub.3; and R.sup.4 is
--Y--C(.dbd.O)NH.sub.2, where Y is selected from the group
consisting of --CHCH.sub.3--, --C(CH.sub.3).sub.2--,
cyclopropylidene and cyclobutylidene.
109. A compound selected from the group consisting of
2-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1);
1-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (2);
1-(2-(2-(2-((4-(1-methylpiperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)p-
yrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (3);
1-(2-(2-(2-((4-(azetidin-3-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (4);
1-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (5);
1-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclop-
ropanecarboxamide (6);
1-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclop-
ropanecarboxamide (7);
1-(2-(2-(5-chloro-2-((4-(1-methylpiperidin-4-yl)phenyl)amino)pyrimidin-4--
yl)ethyl)phenyl)cyclopropanecarboxamide (8);
1-(2-(2-(5-chloro-2-((6-(trifluoromethyl)pyridin-3-yl)amino)pyrimidin-4-y-
l)ethyl)phenyl)cyclopropanecarboxamide (9);
1-(2-(2-(5-(trifluoromethyl)-2-((6-(trifluoromethyl)pyridin-3-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (10);
1-(2-(2-(5-chloro-2-(oxazol-2-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclopr-
opanecarboxamide (11);
1-(2-(2-(5-chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)phenyl)cycl-
opropanecarboxamide (12);
1-(2-(2-(5-chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (13);
1-(2-(2-(2-((1-(azetidin-3-yl)-1H-pyrazol-4-yl)amino)-5-chloropyrimidin-4-
-yl)ethyl)phenyl)cyclopropanecarboxamide (14);
1-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)cyclopropanecarboxamide (15);
1-(2-(2-(5-chloro-2-(5-(1-methylpiperidin-3-yl)pyridin-3-ylamino)pyrimidi-
n-4-yl)ethyl)phenyl)cyclopropanecarboxamide (16);
1-(2-(2-(5-chloro-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (17);
1-(2-(2-(5-chloro-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (18);
1-(2-(2-(2-(1H-pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phenyl)cyc-
lopropanecarboxamide (19);
1-(2-(2-(5-chloro-2-(1-(pyrrolidin-3-yl)-1H-pyrazol-4-ylamino)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (20);
1-(2-(2-(5-chloro-2-(1-(1-methylpyrrolidin-3-yl)-1H-pyrazol-4-ylamino)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (21);
1-(2-(2-(5-chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)phenyl)cycl-
opropanecarboxamide (22);
1-(2-(2-(5-chloro-2-(phenylamino)pyrimidin-4-yl)ethyl)phenyl)cyclopropane-
carboxamide (23); tert-butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate (24);
1-(2-(2-(5-chloro-2-((3-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (25);
1-(2-(2-(5-chloro-2-((1,3-dimethyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)e-
thyl)phenyl)cyclopropanecarboxamide (26);
1-(2-(2-(5-chloro-2-((5-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (27);
1-(2-(2-(5-chloro-2-((1-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (28);
1-(2-(2-(5-chloro-2-((1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-yl)amino)pyri-
midin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (29);
1-(2-(2-(5-chloro-2-((1-isopropyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (30);
1-(2-(2-(5-chloro-2-((6-cyanopyridin-3-yl)amino)pyrimidin-4-yl)ethyl)phen-
yl)cyclopropanecarboxamide (31);
1-(2-(2-(2-((6-acetylpyridin-3-yl)amino)-5-chloropyrimidin-4-yl)ethyl)phe-
nyl)cyclopropanecarboxamide (32);
1-(2-(2-(2-((6-(1-aminoethyl)pyridin-3-yl)amino)-5-chloropyrimidin-4-yl)e-
thyl)phenyl)cyclopropanecarboxamide (33);
1-(2-(2-(5-chloro-2-((1-(difluoromethyl)-1H-pyrazol-4-yl)amino)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (34);
2-(2-(2-(2-(1H-pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phenyl)
propanamide (35);
2-(2-(2-(5-chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl) propanamide (36);
2-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)propanamide (37);
2-(2-(2-(5-chloro-2-(6-(1-methylpiperidin-4-yl)pyridin-3-ylamino)pyrimidi-
n-4-yl)ethyl)phenyl)propanamide (38);
2-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (39);
2-(2-(2-(5-chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (40);
2-(2-(2-(5-chloro-2-(6-methylpyridin-3-ylamino)pyrimidin-4-yl)ethyl)pheny-
l) propanamide (41);
2-(2-(2-(5-chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (42);
2-(2-(2-(5-chloro-2-(1-(piperidin-4-yl)-1H-pyrazol-4-ylamino)pyrimidin-4--
yl)ethyl)phenyl)propanamide (43);
2-(2-(2-(5-chloro-2-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-ylamino)pyri-
midin-4-yl)ethyl)phenyl)propanamide (44);
2-(2-(2-(5-chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)-2-methylpropanamide (45);
1-(2-(2-(2-((1H-pyrazol-4-yl)amino)-5-methylpyrimidin-4-yl)ethyl)phenyl)c-
yclopropanecarboxamide (46);
1-(2-(2-(5-methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (47);
1-(2-(2-(5-methyl-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (48);
1-(2-(2-(5-methyl-2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)pyrimi-
din-4-yl)ethyl)phenyl)cyclopropanecarboxamide (49);
1-(2-(2-(2-((4-(1-aminoethyl)phenyl)amino)-5-methylpyrimidin-4-yl)ethyl)p-
henyl)cyclopropanecarboxamide (50);
2-(2-(2-(5-methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)propanamide (51);
1-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclobutanecarboxamide (52);
1-(3-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (53);
2-(2-(2-(2-((1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)eth-
yl)phenyl)propanamide (54);
2-(2-(2-(2-((1-methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)propanamide (55);
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide (56);
2-(2-(2-(2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57);
2-(2-(2-(2-((1-(1-acetylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (58);
2-(2-(2-(2-((1-(1-ethylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluor-
omethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (59);
2-(2-(2-(2-((1-(1-isopropylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trif-
luoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (60);
2-(2-(2-(2-((6-methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-4--
yl)ethyl)phenyl)propanamide (61);
2-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)propanamide (62);
2-(2-(2-(2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (63);
1-(2-(2-(2-((6-methoxypyridin-3-yl)amino)-5-(trifluoromethyl)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (64);
1-(2-(2-(2-((1-methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (65);
1-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (66);
1-(2-(2-(2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (67);
1-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (68);
1-(2-(2-(2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (69);
tert-butyl
1-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)
pyrimidin-2-ylamino)phenyl)ethylcarbamate (70);
1-(2-(2-(2-(4-(1-acetamidoethyl)phenylamino)-5-(trifluoromethyl)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (71);
1-(2-(2-(2-((6-(1-aminoethyl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyrim-
idin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (72);
1-(2-(2-(2-((6-(1-(methylamino)ethyl)pyridin-3-yl)amino)-5-(trifluorometh-
yl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (73);
1-(2-(2-(2-((6-(1-(azetidin-1-yl)ethyl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (74);
1-(2-(2-(2-((6-(1-morpholinoethyl)pyridin-3-yl)amino)-5-(trifluoromethyl)-
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (75);
1-{2-[2-(5-chloro-2-{[1-(2-methoxyethyl)-1H-pyrazol-4-yl]amino}pyrimidin--
4-yl)ethyl]phenyl}cyclopropanecarboxamide (76); and
1-{2-[2-(5-chloro-2-{[1-(2-hydroxyethyl)-1H-pyrazol-4-yl]amino}pyrimidin--
4-yl)ethyl]phenyl}cyclopropanecarboxamide (77), or a stereoisomer,
salt, solvate, protected form or prodrug thereof.
110. A process for the preparation of a compound according to claim
79 or a stereoisomer, salt, solvate, protected form or prodrug
thereof, comprising reacting a compound of formula F1 ##STR00232##
with a compound of formula A-NH.sub.2 to displace the group L1 and
with a compound of the formula HC.ident.R3 to displace the group
L2; or with a compound of formula HC.ident.R3 to displace the group
L2 and with a compound of formula A-NH.sub.2 to displace the group
L2, wherein R2, A and R3 are as defined in formula (I) in claim 79
and L1 and L2 are each leaving groups.
111. A pharmaceutical agent comprising a compound according to
claim 79 or a stereoisomer, salt, solvate, protected form or
prodrug thereof.
112. A composition comprising a compound according to claim 79 or a
stereoisomer, salt, solvate, protected form or prodrug thereof, and
a pharmaceutically acceptable carrier or diluent.
113. A method for treating a disease ameliorated by the inhibition
of VEGFR3 comprising administering an effective amount of a
compound according to claim 79, or a stereoisomer, salt, solvate,
protected form or prodrug thereof, to a subject in need
thereof.
114. A method for treating cancer comprising administering an
effective amount of a compound according to claim 79, or a
stereoisomer, salt, solvate, protected form or prodrug thereof, to
a subject in need thereof.
115. A method of inhibiting VEGFR3 in vitro or in vivo, comprising
contacting a cell with an effective amount of a compound according
to claim 79, or a stereoisomer, salt, solvate, protected form or
prodrug thereof.
116. The method of claim 114, wherein the cancer is melanoma,
breast cancer or head and neck cancer.
117. An anti-cancer treatment comprising a compound according to
claim 79, or a stereoisomer, salt, solvate, protected form or
prodrug thereof, and an anti-tumor agent.
Description
[0001] This invention relates to 2,4,5-substituted pyrimidines that
inhibit vascular endothelial growth factor receptor 3 (VEGFR3),
also known as Fms related tyrosine kinase 4 (FLT4), processes for
their preparation and pharmaceutical agents or compositions
containing such compounds. This invention also relates to a method
of using such compounds for the treatment of proliferative
diseases, such as cancer, as well as the treatment of diseases
ameliorated by the control and/or inhibition of
lymphangiogenesis.
BACKGROUND
[0002] Cancer remains a major cause of death in the 21st century.
Consequently, considerable drug research and development effort is
currently placed on the discovery of therapeutics that may provide
life extending or curative options to cancer sufferers.
[0003] While there are many different varieties of cancer, each
exhibiting a different array of genetic and growth properties, a
common denominator among many solid cancer types is the ability to
metastasise. Until the occurrence of metastasis, tumors are
confined to one area of the body and may be controlled through
surgical intervention and/or radiotherapy. However, metastasis
causes cancer cells to spread to disparate parts of the body and
while surgical intervention may remove the primary tumor lesion,
removal of all metastatic lesions is very difficult to manage.
[0004] Tumor metastasis is a multistage process, involving the
breakdown of extracellular matrix, invasion of local tissue
parenchyma, intravasation into regional blood vessels and
lymphatics, survival in the circulation and finally extravasation,
survival and growth in secondary tissue sites (Front. Biosci.
(Elite Ed). 2012; 4: 1888-1897).
[0005] Metastasis may occur through blood vessels or lymphatic
vessels. Lymphatic vessels differ from blood vessels in several
ways. Large collecting lymphatic vessels contain vascular smooth
muscle cells in their wall, as well as valves, which prevent the
backflow of lymph. However, lymphatic capillaries, unlike typical
blood capillaries, lack pericytes and continuous basal lamina and
contain large inter-endothelial valve-like openings (J. Theor. Med.
2003; 5: 59-66). Due to their greater permeability, lymphatic
capillaries are more effective than blood capillaries in allowing
tumor cells to pass. Experimental evidence demonstrates that
lymphangiogenesis (the formation of new lymphatic vessels) within a
growing tumor lesion promotes metastasis through lymphatic vessels.
The control of lymphangiogenesis presents an attractive therapeutic
strategy for preventing lymph node metastasis (J. Clin. Onc. 2007;
25: 4298-4307).
[0006] The lymphatic system is comprised of capillaries and larger
collecting vessels continuously lined by endothelial cells which
return extravasated fluid and macromolecules from the interstitial
space back to the blood circulation. Metastasis to regional lymph
nodes via lymphatic vessels is a tumor progression process that is
common to many cancer types. The extent of lymph node involvement
is a major determinant for the staging of many types of cancer and
is an important prognostic factor that is used as the basis for
surgical and radiation treatment intervention of the affected lymph
nodes.
[0007] Molecular signalling through binding of the growth factors
VEGFC or VEGFD to their membrane receptor VEGFR3 has been shown to
play a central role in the process of lymphangiogenesis (Brit. J.
Cancer 2006; 94: 1355-1360). Stimulation of the VEGFR3 receptor
occurs through the phosphorylation of its intracellular region and
triggers a downstream signalling cascade that drives lymphatic
endothelial cell proliferation, migration and differentiation
leading to formation of lymphatic vessels (Exp. Cell Res. 2006;
312: 575-583). Increased expression of VEGFC or VEGFD has been
shown to promote tumor associated lymphangiogenesis enabling
lymphatic-mediated metastasis to regional lymph nodes. These
observations have been reported for several different tumor types,
including colorectal (Oncol. Rep. 2009; 22: 1093-1100) lung (Ann.
Oncol. 2010; 21: 223-231), gastric (Surgery 2009; 146: 896-905),
kidney (Oncol. Rep. 2008; 20: 721-725) prostate (Clin. Cancer Res.
2004; 10: 5137-5144) and ovarian (Cancer 2004; 101: 1364-1374).
Blockade of VEGFC, VEGFD/VEGFR3 mediated signalling has been shown
to inhibit lymphangiogenesis and suppress lymph node metastasis in
several tumor experimental models in rodents (Ann. N.Y. Acad. Sci.
2008; 113: 225-234; Int. J. Cancer 2009; 125: 2747-2756).
[0008] VEGFR3 is a transmembrane tyrosine kinase receptor that is
broadly expressed in endothelial cells during embryogenesis
(Biochem. J. 2011; 437: 169-183). In the latter stages of
development VEGFR3 expression becomes restricted to developing
lymphatic vessels. In adults, VEGFR3 expression is primarily
restricted to lymphatic endothelium and a subset of CD34+
hematopoietic cells. In addition, fenestrated capillaries and veins
in certain endocrine organs, as well as monocytes, macrophages and
some dendritic cells (DCs), continue to express VEGFR3 in adults.
Disruption of the VEGFR3 gene in mouse embryos results in the
failure of vascular network formation and death after embryonic day
9.5 (Biochem. J. 2011; 437: 169-183). This observation demonstrates
that VEGFR3 plays an essential role in the development of embryonic
vasculature. In cancer, VEGFR3 is overexpressed in lymphatic
sinuses in metastatic lymph nodes and in lymphangiomas.
Furthermore, in many instances cancer cells themselves express
VEGFR3. VEGFR3 expressing cancer cells have been shown to be
dependent on VEGFR3/VEGFC signalling for their proliferation (Eur.
J. Canc. 2011; 47: 2353-2363).
[0009] Based on the foregoing, it is apparent that inhibition of
VEGFR3 signalling has strong potential as therapeutic strategy for
mammalian subjects that have been diagnosed with a disease
characterised by proliferation of endothelial cells that express
this receptor. In the case of cancer, targeting VEGFR3 is likely to
result in therapeutic benefit through suppression of lymphatic
metastasis and suppression of growth in cancer cells that express
VEGFR3.
[0010] Interestingly, and perhaps importantly from the view point
of target selection within the VEGFR3 axis, in mice in which both
the VEGFC and the VEGFD genes have been homozygously deleted, the
blood vasculature develops normally, unlike the embryonic
cardiovascular phenotype of VEGFR3 homozygous knockout mice: i.e.
deletion of these two ligands is not the same as deletion of the
receptor (Mol. Cell. Biol. 2008; 28: 4843-4850). These data raise
the possibility that another ligand for VEGFR3 exists or that
VEGFR3 may be able to act by an as-yet-unknown manner independent
of its ligands VEGFC and VEGFD. The foregoing suggest that
targeting VEGFR3 is more advantageous to blocking VEGFC/D-VEGFR3
signalling compared to targeting either VEGFC or VEGFD alone.
[0011] Whilst there are a number of studies reported involving
tyrosine kinase inhibitors with various levels of VEGFR3 activity
and selectivity (Nat. Rev. Drug Discov. 2006; 5: 835-844; Mol.
Cancer Ther. 2007; 6: 2012-2021; Cancer Res. 2009; 69: 8009-8016;
Mol. Cancer Ther. 2012; 11: 1637-1649) these studies have some
limitations, resulting in part at least from inhibition at other
tyrosine kinases.
[0012] Nonetheless, collectively these studies strengthen the
conclusion that inhibition of VEGFR3 suppresses or reduces
lymphangiogenesis and/or lymphogenic metastasis.
[0013] Accordingly, compounds that selectively inhibit VEGFR3 would
be useful for the treatment of proliferative diseases, such as
cancer.
[0014] As described above, VEGFR3 plays an important role in the
control of lymphangiogenesis. Accordingly, inhibitors of VEGFR3 may
have utility in the treatment of diseases other than cancer where
control/inhibition of lymphangiogenesis has a therapeutic benefit.
The lymphatic system plays a major role in chronic inflammatory
diseases and in transplant rejection. Inhibition of
lymphangiogenesis through suppression of VEGFR3 function may
provide a viable therapeutic strategy in these conditions.
[0015] For example, preclinical studies have demonstrated that the
expression of VEGFR3 in the cornea and ocular surface is modified
during corneal neovascularisation and that VEGFR3 mediates corneal
dendritic cell migration to lymph nodes and induction of immunity
to corneal transplant. High-risk corneal transplantation, where
grafting is performed on inflamed and highly vascularized host
beds, has a very poor success rate, with rejection rates as high as
90% (J. Leukoc Biol. 2003; 74: 172-178). In preclinical models,
treatment with a VEGFR3 antibody leads to significant suppression
of corneal graft rejection (Nat. Med. 2004; 10: 813-815).
[0016] Choroidal neovascularization (CNV), the creation of new
blood vessels in the choroid layer of the eye, leads to chronic
inflammation which is implicated in the pathogenesis of age related
macular degeneration (AMD) and is driven by factors which include
uncontrolled expression of the vascular endothelial growth factor
(VEGF) family members VEGFA and VEGFC (J. Cell. Physiol. 2012;
227(1): 116-26). Treatments for AMD have been developed that target
VEGFA, for example the anti-VEGFA antibodies ranibizumab and
bevacizumab and the anti-VEGF aptamer pegaptanib, but to date no
treatments have been clinically evaluated that mediate effects
through modulation of VEGFC and its cognate receptor VEGFR3.
[0017] Accordingly, compounds that inhibit VEGFR3 may be useful for
the prevention and/or treatment of eye diseases, for example
corneal graft rejection and age related macular degeneration.
[0018] Furthermore, there is increasing evidence that lymphatic
vessels have an active role in chronic inflammation of the skin.
Lymphatic endothelial cell proliferation and lymphatic hyperplasia
have been described in chronic skin inflammation in mice and have
been reported for skin lesions in psoriasis patients (Blood 2004;
104: 1048-1057).
[0019] Accordingly, compounds that inhibit VEGFR3 may be useful for
the prevention and/or treatment of skin inflammations, such as skin
lesions in patients with psoriasis.
[0020] Lymphangiogenesis has also been found to be associated with
kidney transplant rejection. VEGFC producing macrophages induce
formation of new lymphatics which induce and support the
maintenance of an alloreactive immune response in renal transplants
(Nat. Med. 2006; 12: 230-234).
[0021] Accordingly, compounds that inhibit VEGFR3 may be useful for
the prevention and/or treatment of rejection in renal
transplantation.
[0022] Co-pending application WO2012/110773 discloses compounds
which inhibit FAK and VEGFR3.
SUMMARY
[0023] The present inventors have discovered a particular class of
compounds which are effective as VEGFR3 inhibitors. These compounds
may exhibit selectivity for VEGFR3 over kinases such as FAK and/or
VEGFR2.
[0024] In a first aspect, the present invention provides compounds
of the following formula (I), isomers, salts, solvates, protected
forms or prodrugs thereof:
##STR00002##
wherein: A is selected from optionally substituted phenyl and an
optionally substituted 5-10 membered heteroaryl group linked to the
NH group through an aromatic ring carbon atom, in which the
heteroaryl ring system contains 1 to 4 heteroatoms selected from N,
O and S, and; when A is optionally substituted phenyl, A may bear a
substituent R.sup.1A which is not alpha to the NH group and may
optionally further bear one or two substituents R.sup.1B which are
not alpha to the NH group, where R.sup.1A is selected from: [0025]
(i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where R.sup.C1 is selected from H,
C.sub.1-2 alkyl, Z.sup.1 is selected from H, C.sub.1-3 alkyl
optionally substituted by OH, C(.dbd.O)OC.sub.1-4 alkyl and
C(.dbd.O)Me, and Z.sup.3 is H, or Z.sup.1 and Z.sup.3 together with
the N to which they are attached form a 4-6 membered heterocycle
containing at least one N and optionally one O; [0026] (ii)
XNHZ.sup.2, where X is selected from CMe.sub.2, cyclopropylidene,
cyclobutylidene, cyclopentylidene and oxetanylidine and Z.sup.2 is
selected from H, C.sub.1-3 alkyl optionally substituted by OH,
C(.dbd.O)OC.sub.1-3 alkyl and C(.dbd.O)Me; [0027] (iii) a group
selected from R.sup.1A1 to R.sup.1A13.
##STR00003## ##STR00004##
[0027] wherein: [0028] R.sup.N1 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0029] R.sup.N2 is
selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me; [0030] R.sup.N3 is selected from H, C.sub.1-4 alkyl,
C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0031] R.sup.N4 is selected
from H and CH.sub.3; [0032] R.sup.N5 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0033] R.sup.N6 is
selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me; [0034] R.sup.N7 and R.sup.N8 are independently
selected from H and CH.sub.3; [0035] R.sup.N9 is selected from H,
C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0036]
R.sup.N10 is selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl
and C(.dbd.O)Me; [0037] R.sup.N11 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; and [0038] R.sup.N12
is selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me. and where each R.sup.1B is independently selected
from: [0039] (i) C.sub.1-3 alkyl; [0040] (ii) CF.sub.3; [0041]
(iii) F; [0042] (iv) Cl; [0043] (v) O--(C.sub.1-3 alkyl); and
[0044] (vi) CN; when A is an optionally substituted 5-10 membered
heteroaryl group, A may bear a single substituent R.sup.1A which is
not alpha to the NH group, where R.sup.1A is as defined above, and
may optionally further bear one, two or three substituents
R.sup.1C, where each R.sup.1C is independently selected from: (i)
C.sub.1-3 alkyl optionally substituted with one to three
substituents independently selected from F, OH and
O--(C.sub.1-3alkyl);
(ii) F;
[0045] (iii) Cl; (iv) O--(C.sub.1-3 alkyl);
(v) CN;
[0046] (vi) =o; and (vii) C(.dbd.O)(C.sub.1-3 alkyl); R.sup.2 is
selected from H, halo, C.sub.1-4 alkyl, CF.sub.3, CF.sub.2H, CN and
O--(C.sub.1-3alkyl); R.sup.3 is selected from substituted phenyl
and a substituted 6 membered heteroaryl group, where the heteroaryl
ring system contains 1 or 2 N heteroatoms, where R.sup.3 bears a
substituent R.sup.4 either alpha or beta to the --C.sub.2H.sub.4--
group, and may additionally bear further substituents selected from
F, methyl and CF.sub.3; and R.sup.4 is
--Y--C(O)N(R.sup.N13)Z.sup.4, where Y is selected from
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
C.sub.35 cycloalkylidene; R.sup.N13 is selected from H and
CH.sub.3; and Z.sup.4 is selected from H, CH.sub.3 and
OCH.sub.3.
[0047] In some embodiments, the compounds of the first aspect of
the present invention are of formula (I) as defined above with the
proviso that the compound is not:
##STR00005##
[0048] In some embodiments, the compounds of the first aspect are
of formula (I) as defined above with the proviso that:
when R.sup.3 is selected from:
##STR00006##
R.sup.4 is --CH(CH.sub.3)C(O)N(R.sup.N13)Z.sup.4;
[0049] A is either:
##STR00007##
where R.sup.1A is selected from
##STR00008## ##STR00009##
and R.sup.N1 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N2 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N3 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N4 is selected from H and CH.sub.3; R.sup.N5 is selected from
H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N7 and R.sup.N8 are
independently selected from H and CH.sub.3; R.sup.N9 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N10 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; or
##STR00010##
where R.sup.1A is
##STR00011##
and R.sup.N1 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N2 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; then
R.sup.2 is not selected from CF.sub.3, halo, CF.sub.2H and CN.
[0050] In some embodiments the compounds of the first aspect are of
formula (I) as defined above, wherein when A is phenyl, Y is
selected from --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and
C.sub.3-5 cycloalkylidene.
[0051] A second aspect of the invention provides a process for the
preparation of a compound of formula (I) or an isomer, salt,
solvate, protected form or prodrug thereof of the first aspect,
comprising:
reacting a compound of formula F1
##STR00012##
with a compound of formula A-NH.sub.2 to displace the group L.sup.1
and with a compound of the formula HC.ident.R.sup.3 to displace the
group L.sup.2; or with a compound of formula HC.ident.R.sup.3 to
displace the group L.sup.2 and with a compound of formula
A-NH.sub.2 to displace the group L.sup.2, wherein R.sup.2, A and
R.sup.3 areas defined in formula (I) above and L.sup.1 and L.sup.2
are leaving groups.
[0052] A third aspect of the invention provides a pharmaceutical
agent comprising a compound of the formula (I) or isomers, salts,
solvates, protected forms or prodrugs thereof of the first
aspect.
[0053] There is also provided use of a compound of formula (I) or
isomers, salts, solvates, protected forms or prodrugs thereof of
the first aspect as a pharmaceutical agent.
[0054] There is further provided a compound of formula (I) or
isomers, salts, solvates, protected forms or prodrugs thereof of
the first aspect, for use as a pharmaceutical agent.
[0055] The pharmaceutical agent may be an anticancer agent, a
lymphangiogenesis inhibitor, an antimetastasis agent or a VEGFR3
inhibitor.
[0056] A fourth aspect of the invention provides a composition
comprising a compound or an isomer, salt, solvate, protected form
or prodrug thereof of the first aspect and a pharmaceutically
acceptable carrier or diluent.
[0057] A fifth aspect of the invention provides a compound or an
isomer, salt, solvate, protected form or prodrug thereof of the
first aspect, an agent of the third aspect or a composition of the
fourth aspect for use in a method of therapy.
[0058] A sixth aspect of the invention provides for the use of a
compound or an isomer, salt, solvate, protected form or prodrug
thereof of the first aspect, an agent of the third aspect or a
composition of the fourth aspect in the preparation of a medicament
for treating a disease or condition ameliorated by the inhibition
of VEGFR3. The sixth aspect of the invention also provides a
compound or an isomer, salt, solvate, protected form or prodrug
thereof of the first aspect, an agent of the third aspect or a
composition of the fourth aspect for use in a method of treatment
of a disease or condition ameliorated by the inhibition of
VEGFR3.
[0059] A seventh aspect of the invention provides for the use of a
compound or an isomer, salt, solvate, protected form or prodrug
thereof of the first aspect, an agent of the third aspect or a
composition of the fourth aspect in the preparation of a medicament
for the treatment of cancer. The seventh aspect of the invention
also provides a compound or an isomer, salt, solvate, protected
form or prodrug thereof, an agent of the third aspect or a
composition of the fourth aspect of the first aspect for use in a
method for the treatment of cancer.
[0060] A further aspect of the invention provides a compound or an
isomer, salt, solvate, protected form or prodrug thereof of the
first aspect, an agent of the third aspect or a composition of the
fourth aspect for use in a method of treatment of the human or
animal body, preferably in the form of a pharmaceutical agent or
composition.
[0061] Another aspect of the invention provides a method of
inhibiting VEGFR3 in vitro or in vivo, comprising contacting a cell
or cell lysates with an effective amount of a compound or an
isomer, salt, solvate, protected form or prodrug thereof of the
first aspect, an agent of the third aspect or a composition of the
fourth aspect.
[0062] A still further aspect of the invention provides an
anti-cancer treatment comprising a compound or an isomer, salt,
solvate, protected form or prodrug thereof of the first aspect, an
agent of the third aspect or a composition of the fourth aspect and
an anti-tumour agent.
[0063] Each of the groups A, and R.sup.1 to R.sup.4 will be
discussed in more detail below.
A
[0064] A is selected from optionally substituted phenyl and an
optionally substituted 5-10 membered heteroaryl group linked to the
NH group through an aromatic ring carbon atom, in which the
heteroaryl ring system contains 1 to 4 heteroatoms selected from N,
O and S.
[0065] If A is unsubstituted phenyl, it has the structure:
##STR00013##
[0066] If A is substituted phenyl, the R.sup.1A group can either be
meta or para, and so A can have the structures:
##STR00014##
where the R.sup.1B group cannot be alpha to the connection point to
the rest of the compound (i.e., it cannot be in the asterixed
positions).
[0067] When A is a 5-10 membered heteroaryl group linked to the NH
group through an aromatic ring carbon atom, in which the heteroaryl
ring system contains 1 to 4 heteroatoms selected from N, O and S,
it is a monovalent moiety obtained by removing a hydrogen atom from
an aromatic ring atom of an heteroaromatic compound (i.e. a
compound having at least one heteroaromatic ring), which moiety has
from 5 to 10 ring atoms. Preferably, each ring has from 5 to 7 ring
atoms.
[0068] Examples of monocyclic heteroaryl groups include, but are
not limited to, those derived from:
N.sub.1: pyrrole (azole) (5-membered), pyridine (azine)
(6-membered); O.sub.1: furan (oxole) (5-membered); S.sub.1:
thiophene (thiole) (5-membered); N.sub.1O.sub.1: oxazole
(5-membered), isoxazole (5-membered), isoxazine (6-membered);
N.sub.2O.sub.1: oxadiazole (furazan) (5-membered); N.sub.3O.sub.1:
oxatriazole (5-membered); N.sub.1S.sub.1: thiazole (5-membered),
isothiazole (5-membered); N.sub.2: imidazole (1,3-diazole)
(5-membered), pyrazole (1,2-diazole) (5-membered), pyridazine
(1,2-diazine) (6-membered), pyrimidine (1,3-diazine) (6-membered)
(e.g., cytosine, thymine, uracil), pyrazine (1,4-diazine)
(6-membered); N.sub.3: triazole (5-membered), triazine
(6-membered); and, N.sub.4: tetrazole (5-membered).
[0069] Examples of heteroaryl groups which comprise fused rings,
include, but are not limited to, those derived from:
9-membered (with 2 fused rings) derived from benzofuran (O.sub.1),
isobenzofuran (O.sub.1), indole (N.sub.1), isoindole (N.sub.1),
indolizine (N.sub.1), indoline (N.sub.1), isoindoline (N.sub.1),
purine (N.sub.4) (e.g., adenine, guanine), benzimidazole (N.sub.2),
indazole (N.sub.2), benzoxazole (N.sub.1O.sub.1), benzisoxazole
(N.sub.1O.sub.1), benzodioxole (O.sub.2), benzofurazan
(N.sub.2O.sub.1), benzotriazole (N.sub.3), benzothiofuran
(S.sub.1), benzothiazole benzothiadiazole (N.sub.2S); 10-membered
(with 2 fused rings) derived from chromene (O.sub.1), isochromene
(O.sub.1), chroman (O.sub.1), isochroman (O.sub.1), benzodioxan
(O.sub.2), quinoline (N.sub.1), isoquinoline (N.sub.1), quinolizine
(N.sub.1), benzoxazine (N.sub.1O.sub.1), benzodiazine (N.sub.2),
pyridopyridine (N.sub.2), quinoxaline (N.sub.2), quinazoline
(N.sub.2), cinnoline (N.sub.2), phthalazine (N.sub.2),
naphthyridine (N.sub.2), pteridine (N.sub.4).
[0070] Thus, when A is a 5 to 10 membered heteroaryl group, it may
be selected from any of the groups listed above.
[0071] If A is 6-membered heteroaryl, the R.sup.1A group can either
be meta or para to the NH group. If A is 5-membered heteroaryl or a
7 to 10 membered heteroaryl, the R.sup.1A group is not alpha to the
--NH-- group. Thus, when A is 5-membered heteroaryl, the R.sup.1A
group is beta to the --NH-- group.
R.sup.1A
[0072] R.sup.1A may have one of the following structures:
CH.sub.2NZ.sup.1Z.sup.3;
CH(CH.sub.3)NZ.sup.1Z.sup.3;
CH(C.sub.2H.sub.5)NZ.sup.1Z.sup.3;
##STR00015## ##STR00016## ##STR00017##
[0073] wherein: [0074] R.sup.N1 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0075] R.sup.N2 is
selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me; [0076] R.sup.N3 is selected from H, C.sub.1-4 alkyl,
C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0077] R.sup.N4 is selected
from H and CH.sub.3; [0078] R.sup.N5 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0079] R.sup.N6 is
selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me; [0080] R.sup.N7 and R.sup.N8 are independently
selected from H and CH.sub.3; [0081] R.sup.N9 is selected from H,
C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; [0082]
R.sup.N10 is selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl
and C(.dbd.O)Me; [0083] R.sup.N11 is selected from H, C.sub.1-4
alkyl, C.sub.3-4 cycloalkyl and C(.dbd.O)Me; and [0084] R.sup.N12
is selected from H, C.sub.1-4 alkyl, C.sub.3-4 cycloalkyl and
C(.dbd.O)Me.
[0085] Each of R.sup.N1, R.sup.N2, R.sup.N3, R.sup.N5, R.sup.N6,
R.sup.N9, R.sup.N10, R.sup.N11 and R.sup.N12 is independently
selected from H, C.sub.1-4 alkyl (i.e. methyl, ethyl, prop-1-yl,
prop-2-yl, n-butyl, iso-butyl, sec-butyl, tert-butyl), C.sub.3-4
cycloalkyl (i.e. cyclopropyl, methylcyclopropyl, cyclobutyl) and
C(.dbd.O)Me. Each of R.sup.N4, R.sup.N7 and R.sup.N8 is
independently selected from either H or methyl.
[0086] Z.sup.1 is independently selected from H, C.sub.1-3 alkyl
(i.e. methyl, ethyl, prop-1-yl and prop-2-yl), optionally
substituted by OH, C(.dbd.O)OC.sub.1-4 alkyl (i.e.
C(.dbd.O)O-methyl, C(.dbd.O)O-ethyl, C(.dbd.O)O-prop-1-yl,
C(.dbd.O)O-prop-2-yl), C(.dbd.O)O-n-butyl, C(.dbd.O)O-iso-butyl,
C(.dbd.O)O-sec-butyl, C(.dbd.O)O-tert-butyl) and C(.dbd.O)Me.
[0087] Z.sup.2 is independently selected from H, C.sub.1-3 alkyl
(i.e. methyl, ethyl, prop-1-yl and prop-2-yl), optionally
substituted by OH, C(.dbd.O)OC.sub.1-3 alkyl (i.e.
C(.dbd.O)O-methyl, C(.dbd.O)O-ethyl, C(.dbd.O)O-prop-1-yl and
C(.dbd.O)O-prop-2-yl) and C(.dbd.O)Me.
[0088] Z.sup.3 is H, or Z.sup.1 and Z.sup.3 together with N form a
4-6 membered heterocycle containing at least one N and optionally
one O. Examples of suitable 4-6 membered heterocycle containing at
least one N and optionally one O include azetidinyl, pyrrolidinyl,
piperidinyl, piperazinyl, and morpholinyl.
R.sup.1B
[0089] Each R.sup.1B group may be C.sub.1-3 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl), CF.sub.3, F, Cl, O--C.sub.1-3
alkyl (i.e. methoxy, ethoxy, prop-1-oxy and prop-2-oxy) or CN.
These groups may be any available ring position on A, except that
which is alpha to the NH group. There may be up to 2 R.sup.1B
groups (i.e. 1 or 2).
R.sup.1C
[0090] Each R.sup.1C group may be C.sub.1-3 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl) optionally substituted with one to
three substituents selected from F, --OH, Cl, O--C.sub.1-3 alkyl
(i.e. methoxy, ethoxy, prop-1-oxy and prop-2-oxy), CN, .dbd.O or
C(.dbd.O)Me. Examples of suitable optionally substituted C.sub.1-3
alkyl groups include --CFH.sub.2, --CF.sub.2H, --CF.sub.3,
--CH.sub.2CFH.sub.2, --CH.sub.2CF.sub.2H, --CH.sub.2CF.sub.3,
--CFHCH.sub.3, --CF.sub.2CH.sub.3, --CH.sub.2CH.sub.2CFH.sub.2,
--CH.sub.2CH.sub.2CF.sub.2H, --CH.sub.2CH.sub.2CF.sub.3,
--CH.sub.2CFHCH.sub.3, --CH.sub.2CF.sub.2CH.sub.3,
--CFHCH.sub.2CH.sub.3, --CF.sub.2CH.sub.2CH.sub.3, --CH.sub.2OH,
--CH.sub.2OCH3, --CH(OH)CH.sub.3, CH(OCH.sub.3)CH.sub.3,
--CH.sub.2CH.sub.2OH, --CH.sub.2CH.sub.2OCH.sub.3,
--CH(OH)CH.sub.2CH.sub.3, CH(OCH.sub.3)CH.sub.2CH.sub.3,
--CH.sub.2CH(OH)CH.sub.3, --CH.sub.2CH(OCH.sub.3)CH.sub.3,
--CH.sub.2CH.sub.2CH.sub.2OH and
--CH.sub.2CH.sub.2CH.sub.2OCH.sub.3. These groups may be
substituted at any available ring position on A. There may be up to
3 R.sup.1C groups (i.e. 1, 2 or 3) depending on the nature of A,
and in particular on the number of ring atoms and ring heteroatoms,
as well as whether R.sup.1A is present.
R.sup.2
[0091] R.sup.2 is selected from H, halo (i.e. F, Cl, Br, I),
C.sub.1-4 alkyl (i.e. methyl, ethyl, prop-1-yl and prop-2-yl,
n-butyl, iso-butyl, sec-butyl, tert-butyl), CF.sub.3, CF.sub.2H, CN
and methoxy.
[0092] In some embodiments, the halo group is either F or Cl.
R.sup.3
[0093] R.sup.3 is selected from substituted phenyl and a
substituted 6 membered heteroaryl group, where the heteroaryl ring
system contains 1 or 2 N heteroatoms.
[0094] When R.sup.3 is substituted phenyl, it has the
structure:
##STR00018##
where R.sup.6, R.sup.7 and R.sup.8 are independently selected from
H, F, methyl and CF.sub.3. One of R.sup.4A and R.sup.4B is R.sup.4,
and the other is selected from H, F, methyl and CF.sub.3.
[0095] When R.sup.3 is a substituted 6 membered heteroaryl group,
where the heteroaryl ring system contains 1 or 2 N heteroatoms, it
may be selected from the any of the groups: pyridyl; pyridazinyl
(1,2-diazinyl); pyrimidinyl (1,3-diazinyl); and pyrazinyl
(1,4-diazinyl).
[0096] When R.sup.3 is a substituted 6 membered heteroaryl group,
it may have one of the following structures:
##STR00019## ##STR00020##
where R.sup.6, R.sup.7 and R.sup.8 (if present) are independently
selected from H, F, methyl and CF.sub.3. One of R.sup.4A and
R.sup.4B (if present) is R.sup.4, and the other is selected from H,
F, methyl and CF.sub.3.
[0097] When R.sup.4 is alpha to the --C.sub.2H.sub.4-- group, it
may also be described as being ortho. When R.sup.4 is beta to the
--C.sub.2H.sub.4-- group, it may also be described as being
meta.
[0098] The further optional substituents on R.sup.3 are
independently selected from F, methyl and CF.sub.3. These further
groups may be at any available ring position on R.sup.3, except
that occupied by R.sup.4. There may be up to 4 further optional
substituents groups (i.e. 1, 2, 3 or 4) depending on the nature of
R.sup.3, and in particular on the number of ring heteroatoms.
R.sup.4
[0099] R.sup.4 is selected from groups of the following
formulae:
##STR00021##
[0100] R.sup.N13 is selected from H and CH.sub.3, and Z.sup.4 is
selected from H, CH.sub.3 or OCH.sub.3. Thus, R.sup.4 can be of the
following formulae:
(i) --Y--C(O)NH.sub.2;
(ii) --Y--C(O)NHMe;
[0101] (iii) --Y--C(O)NMe.sub.2;
(iv) --Y--C(O)N(OMe)H; and
(v) --Y--C(O)N(OMe)Me,
[0102] where Y is selected from --CHCH.sub.3--,
--CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and C.sub.3-5
cycloalkylidene.
Proviso
[0103] In some embodiments, compounds 12 and 155 of WO2012/110773
are disclaimed from the present application:
##STR00022##
[0104] In other embodiments, the compounds of the present invention
do not include those disclosed in WO2012/110773, which is
incorporated herein by reference. In particular, when R.sup.3 is
selected from:
##STR00023##
R.sup.4 is --CH(CH.sub.3)C(O)N(R.sup.N13)Z.sup.4;
[0105] A is either:
##STR00024##
where R.sup.1A is selected from
##STR00025##
and R.sup.N1 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N2 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N3 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N4 is selected from H and CH.sub.3; R.sup.N5 is selected from
H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N7 and R.sup.N8 are
independently selected from H and CH.sub.3; R.sup.N9 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; R.sup.N10 is selected
from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; or
##STR00026##
where R.sup.1A is
##STR00027##
and R.sup.N1 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me;
R.sup.N2 is selected from H, C.sub.1-3 alkyl, and C(.dbd.O)Me; then
R.sup.2 is not selected from CF.sub.3, halo, CF.sub.2H and CN.
[0106] Thus, this proviso only applies when R.sup.4 is
--CH(CH.sub.3)C(O)N(R.sup.N13)Z.sup.4, i.e. when Y is
--CH(CH.sub.3)--.
[0107] In another embodiment the compounds of the first aspect are
of formula (I), wherein when A is phenyl, Y is selected from
--CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and C.sub.3-5
cycloalkylidene.
Includes Other Forms
[0108] Included in the above are the well known ionic, salt,
solvate, and protected forms of these substituents. For example, a
reference to carboxylic acid (--COOH) also includes the anionic
(carboxylate) form (--COO), a salt or solvate thereof, as well as
conventional protected forms. Similarly, a reference to an amino
group includes the protonated form (--N.sup.+HR.sup.1R.sup.2), a
salt or solvate of the amino group, for example, a hydrochloride
salt, as well as conventional protected forms of an amino group.
Similarly, a reference to a hydroxyl group also includes the
anionic form (--O.sup.-), a salt or solvate thereof, as well as
conventional protected forms of a hydroxyl group.
Alpha/Beta
[0109] The terms alpha and beta are used herein to indicate the
relative position of substituent groups on rings. For the avoidance
of doubt, their meaning is illustrated with the structure
below:
##STR00028##
wherein the bromo group is alpha to the chloro group, and the iodo
group is beta to the chloro group.
Isomers, Salts, Solvates, Protected Forms, and Prodrugs
[0110] Certain compounds may exist in one or more particular
geometric, optical, enantiomeric, diasteriomeric, epimeric,
stereoisomeric, tautomeric, conformational, or anomeric forms,
including but not limited to, cis- and trans-forms; E- and Z-forms;
c-, t-, and r-forms; endo- and exo-forms; R-, S-, and meso-forms;
D- and L-forms; B- and I-forms; (+) and (-) forms; keto-, enol-,
and enolate-forms; syn- and anti-forms; synclinal- and
anticlinal-forms; .alpha.- and .beta.-forms; axial and equatorial
forms; boat-, chair-, twist-, envelope-, and halfchair-forms; and
combinations thereof, hereinafter collectively referred to as
"isomers" (or "isomeric forms").
[0111] Note that, except as discussed below for tautomeric forms,
specifically excluded from the term "isomers", as used herein, are
structural (or constitutional) isomers (i.e. isomers which differ
in the connections between atoms rather than merely by the position
of atoms in space). For example, a reference to a methoxy group,
--OCH.sub.3, is not to be construed as a reference to its
structural isomer, a hydroxymethyl group, --CH.sub.2OH. Similarly,
a reference to ortho-chlorophenyl is not to be construed as a
reference to its structural isomer, meta-chlorophenyl. However, a
reference to a class of structures may well include structurally
isomeric forms falling within that class (e.g., C.sub.1-7 alkyl
includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-,
and tert-butyl; methoxyphenyl includes ortho-, meta-, and
para-methoxyphenyl).
[0112] The above exclusion does not pertain to tautomeric forms,
for example, keto-, enol-, and enolate-forms, as in, for example,
the following tautomeric pairs: keto/enol (illustrated below),
imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime,
thioketone/enethiol, N-nitroso/hyroxyazo, and nitro/aci-nitro.
##STR00029##
[0113] Note that specifically included in the term "isomer" are
compounds with one or more isotopic substitutions. For example, H
may be in any isotopic form, including .sup.1H, .sup.2H (D), and
.sup.3H (T); C may be in any isotopic form, including .sup.12C,
.sup.13C and .sup.14C; O may be in any isotopic form, including
.sup.16O and .sup.18O; and the like.
[0114] Unless otherwise specified, a reference to a particular
compound includes all such isomeric forms, including (wholly or
partially) racemic and other mixtures thereof. Methods for the
preparation (e.g. asymmetric synthesis) and separation (e.g.,
fractional crystallisation and chromatographic means) of such
isomeric forms are either known in the art or are readily obtained
by adapting the methods taught herein, or known methods, in a known
manner.
[0115] Unless otherwise specified, a reference to a particular
compound also includes ionic, salt, solvate, and protected forms of
thereof, for example, as discussed below.
[0116] It may be convenient or desirable to prepare, purify, and/or
handle a corresponding salt of the active compound, for example, a
pharmaceutically-acceptable salt. Examples of pharmaceutically
acceptable salts are discussed in Berge et al. J. Pharm. Sci., 66,
1-19 (1977).
[0117] For example, if the compound is anionic, or has a functional
group which may be anionic (e.g., --COOH may be --COO.sup.-), then
a salt may be formed with a suitable cation. Examples of suitable
inorganic cations include, but are not limited to, alkali metal
ions such as Na.sup.+ and K.sup.+, alkaline earth cations such as
Ca.sup.2+ and Mg.sup.2+, and other cations such as Al.sup.3+.
Examples of suitable organic cations include, but are not limited
to, ammonium ion (i.e., NH.sub.4.sup.+) and substituted ammonium
ions (e.g., NH.sub.3R.sup.+, NH.sub.2R.sup.2+, NHR.sub.3.sup.+,
NR.sub.4.sup.+). Examples of some suitable substituted ammonium
ions are those derived from: ethylamine, diethylamine,
dicyclohexylamine, triethylamine, butylamine, ethylenediamine,
ethanolamine, diethanolamine, piperazine, benzylamine,
phenylbenzylamine, choline, meglumine, and tromethamine, as well as
amino acids, such as lysine and arginine. An example of a common
quaternary ammonium ion is N(CH.sub.3).sub.4.sup.+.
[0118] If the compound is cationic, or has a functional group which
may be cationic (e.g., --NH.sub.2 may be --NH.sub.3.sup.+), then a
salt may be formed with a suitable anion. Examples of suitable
inorganic anions include, but are not limited to, those derived
from the following inorganic acids: hydrochloric, hydrobromic,
hydroiodic, sulphuric, sulphurous, nitric, nitrous, phosphoric, and
phosphorous. Examples of suitable organic anions include, but are
not limited to, those derived from the following organic acids:
acetic, propionic, succinic, glycolic, stearic, palmitic, lactic,
malic, pamoic, tartaric, citric, gluconic, ascorbic, maleic,
hydroxymaleic, phenylacetic, glutamic, aspartic, benzoic, cinnamic,
pyruvic, salicyclic, sulfanilic, 2-acetyoxybenzoic, fumaric,
phenylsulfonic, toluenesulfonic, methanesulfonic, ethanesulfonic,
ethane disulfonic, oxalic, pantothenic, isethionic, valeric,
lactobionic, and gluconic. Examples of suitable polymeric anions
include, but are not limited to, those derived from the following
polymeric acids: tannic acid, carboxymethyl cellulose.
[0119] It may be convenient or desirable to prepare, purify, and/or
handle a corresponding solvate of the active compound. The term
"solvate" is used herein in the conventional sense to refer to a
complex of solute (e.g. active compound, salt of active compound)
and solvent. If the solvent is water, the solvate may be
conveniently referred to as a hydrate, for example, a mono-hydrate,
a di-hydrate, a tri-hydrate, etc. It may be convenient or desirable
to prepare, purify, and/or handle the active compound in a
chemically protected form. The term "chemically protected form", as
used herein, pertains to a compound in which one or more reactive
functional groups are protected from undesirable chemical
reactions, that is, are in the form of a protected or protecting
group (also known as a masked or masking group or a blocked or
blocking group). By protecting a reactive functional group,
reactions involving other unprotected reactive functional groups
can be performed, without affecting the protected group; the
protecting group may be removed, usually in a subsequent step,
without substantially affecting the remainder of the molecule. See,
for example, Protective Groups in Organic Synthesis (T. Green and
P. Wuts, Wiley, 1999).
[0120] For example, a hydroxy group may be protected as an ether
(--OR) or an ester (--OC(.dbd.O)R), for example, as: a t-butyl
ether; a benzyl, benzhydryl (diphenylmethyl), or trityl
(triphenylmethyl) ether; a trimethylsilyl or t-butyldimethylsilyl
ether; or an acetyl ester (--OC(.dbd.O)CH.sub.3, --OAc).
[0121] For example, an aldehyde or ketone group may be protected as
an acetal or ketal, respectively, in which the carbonyl group
(>C=O) is converted to a diether (>C(OR).sub.2), by reaction
with, for example, a primary alcohol. The aldehyde or ketone group
is readily regenerated by hydrolysis using a large excess of water
in the presence of acid.
[0122] For example, an amine group may be protected, for example,
as an amide or a urethane, for example, as: a methyl amide
(--NHCO--CH.sub.3); a benzyloxy amide
(--NHCO--OCH.sub.2C.sub.6H.sub.5, --NH-Cbz); as a t-butoxy amide
(--NHCO--OC(CH.sub.3).sub.3, --NH-Boc); a 2-biphenyl-2-propoxy
amide (--NHCO--OC(CH.sub.3).sub.2C.sub.6H.sub.4C.sub.6H.sub.5,
--NH-Bpoc), as a 9-fluorenylmethoxy amide (--NH--Fmoc), as a
6-nitroveratryloxy amide (--NH--Nvoc), as a
2-trimethylsilylethyloxy amide (--NH-Teoc), as a
2,2,2-trichloroethyloxy amide (--NH-Troc), as an allyloxy amide
(--NH-Alloc), as a 2(-phenylsulphonyl)ethyloxy amide (--NH--Psec);
or, in suitable cases, as an N-oxide (>NO.).
[0123] For example, a carboxylic acid group may be protected as an
ester for example, as: an C.sub.1-7 alkyl ester (e.g. a methyl
ester; a t-butyl ester); a C.sub.1-7 haloalkyl ester (e.g., a
C.sub.1-7 trihaloalkyl ester); a triC.sub.1-7 alkylsilyl-C.sub.1-7
alkyl ester; or a C.sub.5-20 aryl-C.sub.1-7 alkyl ester (e.g. a
benzyl ester; a nitrobenzyl ester); or as an amide, for example, as
a methyl amide.
[0124] For example, a thiol group may be protected as a thioether
(--SR), for example, as: a benzyl thioether; an acetamidomethyl
ether (--S--CH.sub.2NHC(.dbd.O)CH.sub.3).
[0125] It may be convenient or desirable to prepare, purify, and/or
handle the active compound in the form of a prodrug. The term
"prodrug", as used herein, pertains to a compound which, when
metabolised (e.g. in vivo), yields the desired active compound.
Typically, the prodrug is inactive, or less active than the active
compound, but may provide advantageous handling, administration, or
metabolic properties. For example, some prodrugs are esters of the
active compound (e.g. a physiologically acceptable metabolically
labile ester). During metabolism, the ester group (--C(.dbd.O)OR)
is cleaved to yield the active drug. Such esters may be formed by
esterification, for example, of any of the carboxylic acid groups
(--C(.dbd.O)OH) in the parent compound, with, where appropriate,
prior protection of any other reactive groups present in the parent
compound, followed by deprotection if required. Examples of such
metabolically labile esters include those wherein R is C1-7 alkyl
(e.g. -Me, -Et); C.sub.1-7 aminoalkyl (e.g. aminoethyl;
2-(N,N-diethylamino)ethyl; 2-(4-morpholino)ethyl); and
acyloxy-C.sub.1-7 alkyl (e.g. acyloxymethyl; acyloxyethyl; e.g.
pivaloyloxymethyl; acetoxymethyl; 1-acetoxyethyl;
1-(1-methoxy-1-methyl)ethyl-carbonxyloxyethyl; 1-(benzoyloxy)ethyl;
isopropoxy-carbonyloxymethyl; 1-isopropoxy-carbonyloxyethyl;
cyclohexyl-carbonyloxymethyl; 1-cyclohexyl-carbonyloxyethyl;
cyclohexyloxy-carbonyloxymethyl; 1-cyclohexyloxy-carbonyloxyethyl;
(4-tetrahydropyranyloxy) carbonyloxymethyl;
1-(4-tetrahydropyranyloxyl)carbonyloxyethyl;
(4-tetrahydropyranyl)carbonyloxymethyl; and
1-(4-tetrahydropyranyl)carbonyloxyethyl).
[0126] Also, some prodrugs are activated enzymatically to yield the
active compound, or a compound which, upon further chemical
reaction, yields the active compound. For example, the prodrug may
be a sugar derivative or other glycoside conjugate, or may be an
amino acid ester derivative.
Selectivity
[0127] The selectivity of the compounds for inhibiting VEGFR3 over
other kinases, such as FAK, and/or VEGFR2 can be demonstrated by
cellular assay results (see, for example, the VEGFR3 and VEGFR2
assays described below).
Further Embodiments
[0128] The following embodiments and preferences may be combined
with one another as appropriate.
A
[0129] In some embodiments, A is optionally substituted phenyl and
can have the structures:
##STR00030##
where the R.sup.1B group cannot be alpha the connection point to
the rest of the compound.
[0130] In these embodiments (i.e. when A is phenyl), it may be
preferred that either there are no R.sup.1B substituents, or a
single R.sup.1B substituent. If there is a single R.sup.1B
substituent it is may be meta or para, so further preferred A
groups include:
##STR00031##
[0131] In some embodiments, A is an optionally substituted 6
membered heteroaryl group. 6 membered heteroaryl groups include,
but are not limited to: pyridyl, isoxazinyl, pyridazinyl,
pyrimidinyl, pyrazinyl and triazinyl.
[0132] In these embodiments, it may be preferred that A is pyridyl,
which can have the structures:
##STR00032##
[0133] In these embodiments, it is preferred that when R.sup.1A is
present R.sup.1C is not an oxo (.dbd.O) group. Of these structures,
A.sup.6 may be further preferred.
[0134] In some embodiments (i.e. when A is 6 membered heteroaryl
group), there may be no R.sup.1C substituents. Thus, when A is
pyridyl and there are no R.sup.1C groups, it may have the
structures:
##STR00033##
[0135] Of these structures A.sup.6A may be further preferred.
[0136] In other embodiments, (i.e. when A is 6 membered heteroaryl
group), there may be no R.sup.1A substituents. Thus, when A is
pyridyl and there are no R.sup.1C groups, the pyridyl group may be
unsubstituted, or it may have one of more R.sup.1C groups. The
pyridyl group may have one of the following structures:
##STR00034##
[0137] Of these structures A.sup.6A1 may be further preferred.
[0138] In some embodiments, A is an optionally substituted 5
membered heteroaryl group. 5 membered heteroaryl groups include,
but are not limited to: pyrrolyl; furanyl; thiophenyl; oxazolyl;
isoxazolyl; oxadiazolyl; oxatriazolyl; thiazolyl; isothiazolyl;
imidazolyl; pyrazolyl; triazolyl and tetrazolyl.
[0139] In these embodiments, it may be preferred that A is
pyrazolyl, which can have the structures:
##STR00035##
[0140] In these embodiments, it is preferred that R.sup.1C is not
an oxo (.dbd.O) group. Of these structures A.sup.16 and A.sup.18
may be further preferred.
[0141] In some embodiments (i.e. when A is a 5 membered heteroaryl
group), there may be no R.sup.1C substituents. Thus, when A is
pyrazolyl and there are no R.sup.1C groups, it may have the
structures:
##STR00036##
[0142] Of these structures A.sup.19A may be further preferred.
[0143] In other embodiments, (i.e. when A is 5 membered heteroaryl
group), there may be no R.sup.1A substituents. Thus, when A is
pyrazolyl and there are no R.sup.1C groups, the pyrazolyl group may
be unsubstituted, or it may have one of more R.sup.1C groups. The
pyrazolyl group may have one of the following structures:
##STR00037##
[0144] Of these structures A.sup.19A1 may be further preferred.
[0145] Thus particularly preferred structures for A include:
##STR00038##
R.sup.1A
[0146] In some embodiments where R.sup.1A is
CH(R.sup.C1)NZ.sup.1Z.sup.3, R.sup.C1 is selected from H and
C.sub.1-2 alkyl, Z.sup.1 may be any one of:
(i) H;
[0147] (ii) C.sub.1-3 alkyl optionally substituted by OH,
preferably Me, or CH.sub.2CH.sub.2OH; (iii) C(.dbd.O)OC.sub.1-4
alkyl, preferably C(.dbd.O)OMe or C(.dbd.O)OtBu; and
(iv) C(.dbd.O)Me.
[0148] and Z.sup.3 may be H, or Z.sup.1 and Z.sup.3 together with
the N to which they are attached form a 4-6 membered heterocycle
containing one N and optionally one O.
[0149] In some of these embodiments, Z.sup.1 may be selected from H
and CH.sub.2CH.sub.2OH and Z.sup.3 may be H. Thus, in these
embodiments, R.sup.1A is selected from: CH.sub.2NH.sub.2;
CH(CH.sub.3)NH.sub.2; CH(C.sub.2H.sub.5)NH.sub.2;
CH.sub.2NHCH.sub.2CH.sub.2OH; CH(CH.sub.3)NHCH.sub.2CH.sub.2OH; and
CH(C.sub.2H.sub.5)NHCH.sub.2CH.sub.2OH.
[0150] In some embodiments where R.sup.1A is
CH(R.sup.C1)NZ.sup.1Z.sup.3, R.sup.C1 may be selected from H and
methyl and Z.sup.3 may be H. Thus, in these embodiments, R.sup.1A
is selected from: CH.sub.2NHZ.sup.1 and CH(CH.sub.3)NHZ.sup.1.
[0151] In some embodiments where R.sup.1A is CH(R.sup.C1)NHZ.sup.1,
Z.sup.1 may be selected from H and CH.sub.2CH.sub.2OH, and R.sup.C1
may be selected from H and methyl. Thus, in these embodiments,
R.sup.1A is selected from: CH.sub.2NH.sub.2;
CH.sub.2NHCH.sub.2CH.sub.2OH; CH(CH.sub.3)NH.sub.2; and
CH(CH.sub.3)NHCH.sub.2CH.sub.2OH.
[0152] In some embodiments where R.sup.1A is
CH(R.sup.C1)NZ.sup.1Z.sup.3, Z.sup.1 and Z.sup.3 may together with
the N to which they are attached form a 4-6 membered heterocycle
containing one N and optionally one O. In these embodiments,
R.sup.1A is, for example, azetidinyl or morpholinyl.
[0153] In some embodiments where R.sup.1A is XNHZ.sup.2, Z.sup.2
may be any one of:
(i) H;
[0154] (ii) C.sub.1-3 alkyl optionally substituted by OH,
preferably Me, or CH.sub.2CH.sub.2OH; (iii) C(.dbd.O)OC.sub.1-3
alkyl, preferably C(.dbd.O)OMe; and
(iv) C(.dbd.O)Me.
[0155] In some embodiments where R.sup.1A is XNHZ.sup.2, Z.sup.2
may be H. Thus, in these embodiments, R.sup.1A has the
structures:
##STR00039##
[0156] In some embodiments where R.sup.1A is XNHZ.sup.2, Z.sup.2
may be C(.dbd.O)OMe. Thus, in these embodiments, R.sup.1A has the
structures:
##STR00040##
[0157] In some embodiments where R.sup.1A is XNHZ.sup.2, X may be
selected from CMe.sub.2, and cyclobutylidene. Thus, in these
embodiments, R.sup.1A has the structures:
##STR00041##
[0158] In some embodiments where R.sup.1A is XNHZ.sup.2, Z.sup.2
may be selected from H and C(.dbd.O)OMe, and X may be selected from
CMe.sub.2, and cyclobutylidene. Thus, in these embodiments,
R.sup.1A has the structures:
##STR00042##
[0159] In some embodiments, R.sup.1A is:
##STR00043##
wherein R.sup.N1 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N1 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N1 is
H, methyl or ethyl.
[0160] In some embodiments, R.sup.1A is:
##STR00044##
wherein R.sup.N2 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N2 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N2 is
H, methyl or ethyl, more preferably H or methyl.
[0161] In some embodiments, R.sup.1A is:
##STR00045##
wherein R.sup.N3 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N3 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N3 is
H, methyl or ethyl, more preferably H or methyl.
[0162] In some embodiments, R.sup.1A is:
##STR00046##
wherein R.sup.N4 is selected from H or methyl. In some of these
embodiments, it may be preferred that R.sup.N4 is H.
[0163] In some embodiments, R.sup.1A is:
##STR00047##
wherein R.sup.N5 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N5 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N5 is
H, methyl or ethyl, more preferably H or methyl.
[0164] In some embodiments, R.sup.1A is:
##STR00048##
wherein R.sup.N6 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N6 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N6 is
H, methyl or ethyl, more preferably H or methyl.
[0165] In some embodiments, R.sup.1A is:
##STR00049##
wherein R.sup.N7 and R.sup.N8 are both H or both methyl. In some of
these embodiments, it may be preferred that R.sup.N7 and R.sup.N8
are both H.
[0166] In some embodiments, R.sup.1A is:
##STR00050##
wherein R.sup.N9 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N9 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N9 is
H, methyl or ethyl, more preferably H or methyl.
[0167] In some embodiments, R.sup.1A is:
##STR00051##
wherein R.sup.N10 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.n10 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N10 is
H, methyl or ethyl, more preferably H or methyl.
[0168] In some embodiments, R.sup.1A is:
##STR00052##
wherein R.sup.N11 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N11 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N11 is
H, methyl or ethyl, more preferably H or methyl.
[0169] In some embodiments, R.sup.1A is:
##STR00053##
wherein R.sup.N12 is selected from H, C.sub.1-4 alkyl (i.e. methyl,
ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl, sec-butyl,
tert-butyl), C.sub.3-4 cycloalkyl (i.e. cyclopropyl,
methylcyclopropyl, cyclobutyl) and C(.dbd.O)Me. In some of these
embodiments, it may be preferred that R.sup.N12 is C(.dbd.O)Me. In
others of these embodiments, it may be preferred that R.sup.N12 is
H, methyl or ethyl, more preferably H or methyl.
[0170] Particularly preferred R.sup.1A groups include:
##STR00054##
[0171] In some embodiments, R.sup.1A is selected from R.sup.1A2 and
R.sup.1A6:
##STR00055##
wherein: [0172] R.sup.N2 is selected from H and methyl; [0173]
R.sup.N5 is selected from H and methyl; and [0174] R.sup.N6 is
selected from H and methyl.
[0175] In some embodiments, R.sup.1A is selected from R.sup.1A2 and
R.sup.1A6:
##STR00056##
wherein: [0176] R.sup.N2 is selected from H and methyl; and [0177]
R.sup.N6 is selected from H and methyl.
R.sup.1B
[0178] In some embodiments, no R.sup.1B substituents are
present.
[0179] In some embodiments, R.sup.1B is preferably C.sub.1-3 alkyl
and more preferably methyl.
[0180] In some embodiments, a single R.sup.1B substituent is
present. It may be C.sub.1-3 alkyl; CF.sub.3; F; Cl;
O--(C.sub.1-3alkyl); and CN. In some of these embodiments, it is
preferably C.sub.1-3 alkyl, and more preferably methyl.
[0181] R.sup.1C
[0182] In some embodiments, no R.sup.1C substituents are
present.
[0183] In the embodiments where R.sup.1C substituents are present,
each R.sup.1C substituent is independently selected from:
(i) C.sub.1-3alkyl optionally substituted with one to three
substituents selected from F, OH and OMe;
(ii) F;
[0184] (iii) Cl; (iv) O--(C.sub.1-3 alkyl);
(v) CN;
[0185] (vi) =o; and (vii) C(.dbd.O)Me.
[0186] In some embodiments, each R.sup.1C substituent is
independently selected from:
(i) C.sub.1-3 alkyl optionally substituted with one to three
substituents selected from F, OH and OMe; (ii) O--(C.sub.1-3
alkyl);
(v) CN; and
[0187] (vii) C(.dbd.O)Me.
[0188] In other embodiments, R.sup.1C is preferably C.sub.1-3 alkyl
optionally substituted with one to three F atoms and more
preferably methyl or CF.sub.3.
[0189] In some embodiments, a single R.sup.1C substituent is
present. It may be C.sub.1-3 alkyl; CF.sub.3; F; Cl;
O--(C.sub.1-3alkyl); CN; C(.dbd.O)Me; and .dbd.O. In some of these
embodiments, it is preferably C.sub.1-3 alkyl optionally
substituted with one to three F substituents, and more preferably
methyl or CF.sub.3.
R.sup.2
[0190] In some embodiments, R.sup.2 is H.
[0191] In some embodiments, R.sup.2 is halo (i.e. F, Cl, Br, I). In
some of these embodiments, the halo group is either F or Cl.
[0192] In some embodiments, R.sup.2 is C.sub.1-4 alkyl (i.e.
methyl, ethyl, prop-1-yl and prop-2-yl, n-butyl, iso-butyl,
sec-butyl, tert-butyl). In some of these embodiments, the C.sub.1-4
alkyl group is methyl or ethyl, and methyl may be preferred.
[0193] In some embodiments, R.sup.2 is selected from CF.sub.3 and
CF.sub.2H. In some of these embodiments, R.sup.2 is CF.sub.3.
[0194] In some embodiments, R.sup.2 is CN.
[0195] In some embodiments, R.sup.2 is methoxy.
[0196] In some embodiments, R.sup.2 is selected from CF.sub.3 and
Cl.
R.sup.3
[0197] In some embodiments, R.sup.3 is substituted phenyl, and
therefore it has the structure:
##STR00057##
where R.sup.6, R.sup.7 and R.sup.8 are independently selected from
H, F, methyl and CF.sub.3. One of R.sup.4A and R.sup.4B is R.sup.4,
and the other is selected from H, F, methyl and CF.sub.3. In some
of these embodiments, the group of R.sup.4A and R.sup.4B that is
not R.sup.4, and R.sup.6, R.sup.7 and R.sup.8 are all H. In others
of these embodiments, one of the group of R.sup.4A and R.sup.4B
that is not R.sup.4, R.sup.6, R.sup.7 and R.sup.8 is not H, and
therefore is F, methyl or CF.sub.3. The group that is not H may
preferably be R.sup.6 or R.sup.7.
[0198] In some embodiments, R.sup.3 is substituted phenyl,
R.sup.4B, R.sup.6, R.sup.7 and R.sup.8 are all H, and R.sup.4A is
R.sup.4.
[0199] In some embodiments, R.sup.3 is a substituted 6 membered
heteroaryl group, where the heteroaryl ring system contains 1 or 2
N heteroatoms. In these embodiments, it may be preferred that
R.sup.3 is pyridyl, which can have the structures:
##STR00058##
where R.sup.6, R.sup.7 and R.sup.8 (if present) are independently
selected from H, F, methyl and CF.sub.3. One of R.sup.4A and
R.sup.4B (if present) is R.sup.4, and the other is selected from H,
F, methyl and CF.sub.3. Of these structures, R.sup.3d and R.sup.3e
may be preferred. In some of these embodiments, the group of
R.sup.4A and R.sup.4B that is not R.sup.4, and R.sup.6, R.sup.7 and
R.sup.8 (if present) are all H. In others of these embodiments, one
of the group of R.sup.4A and R.sup.4B that is not R.sup.4, R.sup.6,
R.sup.7 and R.sup.8 (if present) is not H, and therefore is F,
methyl or CF.sub.3. In some embodiments, it may be preferred that a
F substituent is not alpha to a ring nitrogen atom.
R.sup.4
[0200] In some embodiments R.sup.4 is alpha to the
--C.sub.2H.sub.4-- group. In some embodiments R.sup.4 is beta to
the --C.sub.2H.sub.4-- group.
[0201] The group R.sup.4 is --Y--C(O)(R.sup.N13)Z.sup.4.
[0202] In some embodiments, R.sup.N13 is H.
[0203] In other embodiments, R.sup.N13 is Me.
[0204] In some embodiments, Z.sup.4 is H.
[0205] In other embodiments, Z.sup.4 is Me.
[0206] In other embodiments, Z.sup.4 is OMe.
[0207] Y can be any of --CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--,
--C(CH.sub.3).sub.2--, C.sub.3-5 cycloalkylidene.
[0208] In some embodiments, Y is selected from CHCH.sub.3--,
--CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and C.sub.3-5
cycloalkylidene.
[0209] In some embodiments, Y is selected from
--CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--, and C.sub.3-5
cycloalkylidene.
[0210] In some embodiments, Y is selected from Y.sup.a to
Y.sup.e:
##STR00059##
[0211] In some embodiments, Y is selected from Y.sup.b, Y.sup.c,
Y.sup.d and Y.sup.e.
[0212] In other embodiments, Y is selected from Y.sup.a, Y.sup.b,
Y.sup.c and Y.sup.e.
[0213] In some embodiments Y is selected from Y.sup.a, Y.sup.c,
Y.sup.d and Y.sup.e.
[0214] In some embodiments, R.sup.4 is --Y'--C(O)NH.sub.2, where Y'
is selected from --C(CH.sub.3).sub.2--, --CH(CH.sub.2CH.sub.3)--,
cyclopropylidene and cyclobutylidene.
[0215] In some embodiments, R.sup.4 is:
##STR00060##
[0216] In some embodiments, R.sup.4 is:
##STR00061##
[0217] In some embodiments, R.sup.4 is:
##STR00062##
[0218] In some embodiments, R.sup.4 is:
##STR00063##
[0219] In some embodiments of the present invention, the compounds
are of formula (Ia) which is a compound of formula (I) of the first
aspect or isomers, salts, solvates, protected forms or prodrugs
thereof wherein:
A is selected from optionally substituted phenyl and an optionally
substituted 5 or 6 membered heteroaryl group linked to the NH group
through an aromatic ring carbon atom, in which the heteroaryl ring
system contains 1 or 2 heteroatoms selected from N and O, and; when
A is optionally substituted phenyl, A may bear a substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is
selected from: [0220] (i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where
R.sup.C1 is selected from H, C.sub.1-2 alkyl, Z.sup.1 is selected
from H and C.sub.1-3 alkyl substituted by C(.dbd.O)OC.sub.1-4 alkyl
or C(.dbd.O)Me and Z.sup.3 is H, or Z.sup.1 and Z.sup.3 together
with the N to which they are attached form a 4-6 membered
heterocycle containing one N and optionally one O; [0221] (iii) a
group selected from:
##STR00064##
[0221] wherein: [0222] R.sup.N2 is selected from H, C.sub.1-3 alkyl
and C(.dbd.O)Me; [0223] R.sup.N5 is selected from H and C.sub.1-4
alkyl; [0224] R.sup.N6 is H; when A is an optionally substituted 5
or 6 membered heteroaryl group, A may bear a single substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is as
defined above, and may optionally further bear one or two
substituents R.sup.1C, where each R.sup.1C is independently
selected from: (i) C.sub.1-3 alkyl optionally substituted with one
to three substituents independently selected from F, OH and
O--(C.sub.1-3alkyl); (ii) O--(C.sub.1-3alkyl);
(ii) CN; and
[0225] (iii) C(.dbd.O)Me; R.sup.2 is selected from halo, CH.sub.3
and CF.sub.3; R.sup.3 is substituted phenyl, where R.sup.3 bears a
substituent R.sup.4 either alpha or beta to the --C.sub.2H.sub.4--
group, and may additionally bear a further substituent F; and
R.sup.4 is --Y--C(O)N(R.sup.N13)Z.sup.4, where Y is selected from
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
and C.sub.3-4 cycloalkylidene; R.sup.N13 is H; and Z.sup.4 is
H.
[0226] In some embodiments of the present invention, the compounds
are of formula (Ib) which is a compound of formula (I) of the first
aspect and formula (Ia) defined above or isomers, salts, solvates,
protected forms or prodrugs thereof wherein:
A is selected from optionally substituted phenyl and an optionally
substituted 5 or 6 membered heteroaryl group linked to the NH group
through an aromatic ring carbon atom, in which the heteroaryl ring
system contains 1 or 2 heteroatoms selected from N and O, and; when
A is optionally substituted phenyl, A may bear a substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is
selected from: [0227] (i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where
R.sup.C1 is selected from H, C.sub.1-2 alkyl, Z.sup.1 is selected
from H and C.sub.1-3 alkyl substituted by C(.dbd.O)OC.sub.1-4 alkyl
or C(.dbd.O)Me and Z.sup.3 is H; [0228] (iii) a group selected
from:
##STR00065##
[0228] wherein: [0229] R.sup.N2 is selected from H, C.sub.1-3 alkyl
and C(.dbd.O)Me; [0230] R.sup.N5 is selected from H and C.sub.1
alkyl; [0231] R.sup.N6 is H; when A is an optionally substituted 5
or 6 membered heteroaryl group, A may bear a single substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is as
defined above, and may optionally further bear one or two
substituents R.sup.1C, where each R.sup.1C is independently
selected from: (i) C.sub.1-3 alkyl optionally substituted with one
to three substituents independently selected from F, OH or
O--(C.sub.1-3 alkyl); (ii) O--(C.sub.1-3 alkyl);
(ii) CN; and
[0232] (iii) C(.dbd.O)Me; R.sup.2 is selected from halo, CH.sub.3
and CF.sub.3; R.sup.3 is substituted phenyl, where R.sup.3 bears a
substituent R.sup.4 either alpha or beta to the --C.sub.2H.sub.4--
group, and may additionally bear a further substituent F; and
R.sup.4 is --Y--C(O)N(R.sup.N13)Z.sup.4, where Y is selected from
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
and C.sub.3-4 cycloalkylidene; R.sup.N13 is H; and Z.sup.4 is
H.
[0233] In some embodiments of the present invention, the compounds
are of formula (Ic) which is a compound of formula (I) of the first
aspect and formulae (Ia) and (Ib) defined above or isomers, salts,
solvates, protected forms or prodrugs thereof wherein:
A is selected from substituted phenyl and an optionally substituted
5 or 6 membered heteroaryl group linked to the NH group through an
aromatic ring carbon atom, in which the heteroaryl ring system
contains 1 or 2 heteroatoms selected from N and O, and; when A is
optionally substituted phenyl, A may bear a substituent R.sup.1A
which is not alpha to the NH group, where R.sup.1A is selected
from: (i) CH(R.sup.C1)NZ.sup.1Z.sup.3, where R.sup.C1 is methyl,
Z.sup.1 is H and Z.sup.3 is H; (iii) a group selected from:
##STR00066##
wherein: [0234] R.sup.N2 is selected from H and methyl; [0235]
R.sup.N5 is selected from H and methyl; [0236] R.sup.N6 is H; when
A is an optionally substituted 5 or 6 membered heteroaryl group, A
may bear a single substituent R.sup.1A which is not alpha to the NH
group, where R.sup.1A is as defined above, and may optionally
further bear one or two substituents R.sup.1C, where each R.sup.1C
is independently selected from:
(i) CH.sub.3 or CF.sub.3;
[0237] R.sup.2 is selected from halo and CF.sub.3; R.sup.3 is
substituted phenyl, where R.sup.3 bears a substituent R.sup.4
either alpha or beta to the --C.sub.2H.sub.4-- group, and may
additionally bear a further substituent F; and R.sup.4 is
--Y--C(O)N(R.sup.N13)Z.sup.4, where Y is selected from
--CHCH.sub.3--, --CH(CH.sub.2CH.sub.3)--, --C(CH.sub.3).sub.2--,
and cyclopropylidene; R.sup.N13 is H; and Z.sup.4 is H.
[0238] In some embodiments of the present invention, the compounds
are of formula (Id) which is a compound of formula (I) of the first
aspect and formulae (Ia) to (Ic) defined above or isomers, salts,
solvates, protected forms or prodrugs thereof wherein:
A is selected from substituted phenyl and an optionally substituted
pyrazolyl or pyridyl group linked to the NH group through an
aromatic ring carbon atom, in which the heteroaryl ring system
contains 1 or 2 heteroatoms selected from N and O, and; when A is
optionally substituted phenyl, A may bear a substituent R.sup.1A
which is not alpha to the NH group, where R.sup.1A is:
##STR00067##
wherein: [0239] R.sup.N2 is methyl; when A is an optionally
substituted pyrazolyl or pyridyl group, A may bear a single
substituent R.sup.1A which is not alpha to the NH group, where
R.sup.1A is as defined above, and may optionally further bear one
or two substituents R.sup.1C, where each R.sup.1C is independently
selected from:
(i) CH.sub.3 or CF.sub.3;
[0240] R.sup.2 is selected from Cl and CF.sub.3; R.sup.3 is
substituted phenyl, where R.sup.3 bears a substituent R.sup.4 alpha
to the --C.sub.2H.sub.4-- group, and may additionally bear a
further substituent F; and R.sup.4 is --Y--C(O)N(R.sup.N13)Z.sup.4,
where Y is selected from --CHCH.sub.3-- and C.sub.3
cycloalkylidene; R.sup.N13 is H; and Z.sup.4 is H.
[0241] In some embodiments of the present invention, the compounds
are of formula (Ie) or isomers, salts, solvates, protected forms or
prodrugs thereof:
##STR00068##
wherein: A is selected from: optionally substituted phenyl and
optionally substituted pyridyl, wherein A may bear one substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is
selected from R.sup.1A2 and R.sup.1A6:
##STR00069##
wherein: [0242] R.sup.N2 is selected from H and methyl; [0243]
R.sup.N6 is selected from H and methyl; R.sup.2 is selected from
Cl, methyl and CF.sub.3; and R.sup.4 is --Y--C(O)NH.sub.2, where Y
is selected from --CHCH.sub.3--, --C(CH.sub.3).sub.2--,
cyclopropylidene and cyclobutylidene; with the proviso that: when A
is either:
##STR00070##
[0243] and R.sup.2 is Cl or CF.sub.3, then R.sup.4 is not
--CH(CH.sub.3)C(O)NH.sub.2.
[0244] In some embodiments of the present invention, the compounds
are of formula (If) or isomers, salts, solvates, protected forms or
prodrugs thereof:
##STR00071##
wherein: A is selected from: optionally substituted phenyl and
optionally substituted pyridyl, wherein A may bear one substituent
R.sup.1A which is not alpha to the NH group, where R.sup.1A is
selected from R.sup.1A2 and R.sup.1A6:
##STR00072##
wherein: [0245] R.sup.N2 is selected from H and methyl; [0246]
R.sup.N6 is selected from H and methyl; R.sup.2 is selected from
Cl, methyl and CF.sub.3; and R.sup.4 is --Y--C(O)NH.sub.2, where Y
is selected from --C(CH.sub.3).sub.2--, cyclopropylidene and
cyclobutylidene.
[0247] In some embodiments of the present invention, the compounds
are of formula (Ig) or isomers, salts, solvates, protected forms or
prodrugs thereof:
##STR00073##
wherein A is selected from an optionally substituted 5 or 6
membered heteroaryl group which contains 1 or 2 heteroatoms
selected from N and O, wherein A may bear one substituent R.sup.1A
which is not alpha to the NH group, where R.sup.1A is selected from
R.sup.1A2, R1A3 and R.sup.1A6
##STR00074##
wherein: [0248] R.sup.N2 is selected from H and C.sub.1-4 alkyl;
[0249] R.sup.N3 is selected from H, C.sub.1-3 alkyl, and
C(.dbd.O)Me; [0250] R.sup.N6 is selected from H and C.sub.1-4alkyl;
and may optionally further bear one substituent R.sup.1C selected
from CF.sub.3, C.sub.1-3alkyl, CH.sub.2CF.sub.3, CN,
C(.dbd.O)(C.sub.1-3alkyl), CH(CH.sub.3)NH.sub.2, CHCF.sub.2,
OCH.sub.3, CH(CH.sub.3)NHCH.sub.3, C.sub.1-3alkylOH and
C.sub.1-3alkylOMe; R.sup.2 is selected from halo, C.sub.1-4alkyl
and CF.sub.3; and R.sup.4 is --Y--C(.dbd.O)NH.sub.2, where Y is
selected from --CHCH.sub.3--, --C(CH.sub.3).sub.2--,
cyclopropylidene and cyclobutylidene.
[0251] In one embodiment of formula (Ig), A is selected from a 5
membered heteroaryl group which contains N such as optionally
substituted pyrazolyl, a 5 membered heteroaryl group which contains
N and O such as optionally substituted oxazolyl and a 6 membered
heteroaryl group which contains N such as optionally substituted
pyridinyl, optionally substituted pyrimidinyl and optionally
substituted pyridazinyl, wherein A bears one substituent selected
from R1A2, (R1A3) and R1A6 defined in formula (Ig) above and may
optionally further bear one substituent R1C selected from CF3,
C1-3alkyl, CH2CF3, CN, C(.dbd.O)Me, CH(CH3)NH2, CHCF2, OMe,
CH(CH3)NHCH3, C1-3alkylOH and C1-3alkylOMe;
R2 is selected from Cl, Me and CF3; and
R4 is CHCH3C(.dbd.O)NH2.
[0252] In a further embodiment of formula (Ig), A is selected from
a 5 membered heteroaryl group which contains N such as optionally
substituted pyrazolyl wherein A bears one substituent selected from
R.sup.1A2 defined in formula (Ig) above;
R.sup.2 is CF.sub.3; and
R.sup.4 is CHCH.sub.3C(.dbd.O)NH.sub.2.
[0253] The preferences expressed in relation to compounds of
formula I also apply to compounds of formulae Ia to Ig, where
appropriate.
[0254] Embodiments of the inventions are compounds of the examples,
including compounds 1 to 77. Embodiments of particular interest
include compounds 2, 3, 4, 5, 6, 7, 8, 13, 15, 19, 20, 33, 35, 45,
48, 49, 50, 56, 56-1A, 56-2A, 57, 57-1A, 57-2A, 68, and 70.
General Synthesis Methods
[0255] The compounds of the invention can be prepared by employing
the following general methods and using procedures described in
detail in the experimental section. The reaction conditions
referred to are illustrative and non-limiting.
[0256] The process for the preparation of a compound of formula (I)
or an isomer, salt, solvate, protected form or prodrug thereof,
comprises reacting a compound of formula F1
##STR00075##
with a compound of formula A-NH.sub.2 to displace the group Land
with a compound of the formula HC.ident.R.sup.3 to displace the
group L.sup.2; or with a compound of formula HC.ident.R.sup.3 to
displace the group L.sup.2 and with a compound of formula A-N
H.sub.2 to displace the group L.sup.2, wherein R.sup.2, A and
R.sup.3 areas defined in formula (I) above and L.sup.1 and L.sup.2
are leaving groups.
[0257] It will be appreciated that the compounds of formulae A-N
H.sub.2 and HC.ident.R.sup.3 can be reacted with the compound of
formula F1 separately or sequentially in any order or
simultaneously.
[0258] The leaving groups L.sup.1 and L.sup.2 may be any suitable
leaving groups, such as a halogen atom (F, Cl, Br, I), --SR or
--S0.sub.2R where R is a C.sub.1-4 straight chain or branched alkyl
group. In some embodiments, L.sup.1 and L.sup.2 may be the same or
different and may be selected from the group consisting of Cl, Br,
I, SMe, SO.sub.2Me.
[0259] Compounds of formula I, as described above, can be prepared
by synthetic strategies outlined below, wherein the definitions
above apply:
##STR00076##
[0260] Compounds of formula F1 may be reacted with substituted
commercial or synthetic amino substituted compounds of formula F2
(as prepared in scheme C to N) to form intermediates of formula F3
where L.sup.1 and L.sup.2 may be the same or different and include
Cl, Br, I, SMe, SO.sub.2Me.
[0261] Compounds of the formula F1 may be prepared where L.sup.1
and L.sup.2 are different to allow regioselective substitution or
when L.sup.1=L.sup.2 suitable reaction conditions can be employed
(choice of solvent, reaction temperature, addition of a Lewis acid,
for example ZnCl.sub.2 in Et.sub.2O) to allow L.sup.1 to be
selectively displaced over L.sup.2. Where regiochemical mixtures
and di-substitution are obtained the regioisomers may be separated
by chromatography.
[0262] Compounds of the formula F1 where L.sup.1=L.sup.2 are either
commercially available, for example
2,4-dichloro-5-(trifluoromethyl)pyrimidine,
2,4-dichloro-5-fluoropyrimidine, 2,4,5-trichloropyrimidine,
2,4-dichloro-5-bromopyrimidine, 2,4-dichloro-5-iodopyrimidine,
2,4-dichloro-5-methylpyrimidine, 2,4-dichloro-5-cyanopyrimidine or
may be prepared readily from commercial starting materials. Where
R.sup.2=CF.sub.3 and differentiation of L.sup.1 and L.sup.2 is
desirable, the method outlined in scheme B may be employed.
##STR00077##
[0263] Commercially available
2,4-dichloro-5-(trifluoromethyl)pyrimidine (G1) can be selectively
reacted with sodium thiomethoxide in the presence of zinc(II)
chloride to give
2-thiomethyl-4-chloro-5-(trifluoromethyl)pyrimidine (G2).
2-Thiomethyl-4-chloro-5-(trifluoromethyl)pyrimidine (G2) can be
further reacted, for example by conversion to
2-thiomethyl-4-iodo-5-(trifluoromethyl)pyrimidine (G3) under
Finkelstein conditions and/or by oxidation with m-CPBA to give the
corresponding sulfone if further differentiation of the 2 and
4-position is required or if additional activation is
desirable.
[0264] Examples of commercially available amino compounds of the
formula F2 include, but are not limited to those depicted in table
1.
TABLE-US-00001 TABLE 1 ##STR00078## ##STR00079## ##STR00080##
##STR00081## ##STR00082## ##STR00083## ##STR00084## ##STR00085##
##STR00086##
[0265] It will be appreciated that compounds of the formula F2,
both commercial and synthetic, can be further modified either prior
or post coupling to pyrimidines of the formula F1 via an extensive
range of chemistries including, but not limited to hydrolysis,
alkylation, acylation, electrophilic halogenation and Mitsunobu
coupling.
[0266] In addition to commercially available amino compounds of the
formula F2, numerous analogous nitro containing compounds are also
commercially available including, but not limited to those depicted
in table 2.
TABLE-US-00002 TABLE 2 ##STR00087## ##STR00088## ##STR00089##
[0267] It will be appreciated that such compounds can be reduced
under suitable conditions, for example in the presence of palladium
under an atmosphere of hydrogen, to give amino compounds of the
formula F2.
[0268] Synthetic amino compounds of the invention may be prepared
via a range of procedures. It will be appreciated that heterocyclic
analogues may also be prepared by analogous methods to those
outlined below via substitution of phenyl containing starting
materials with suitable heteroaromatic systems.
##STR00090##
[0269] Commercially available 1-(4-nitrophenyl)piperazine (G4), or
a salt thereof, can be reacted with Boc anhydride to give
tert-butyl 4-(4-nitrophenyl)piperazine-1-carboxylate (G5).
Subsequent reduction via hydrogenation in the presence of a
catalyst, for example palladium on charcoal, gives the
corresponding aniline, tert-butyl
4-(4-aminophenyl)piperazine-1-carboxylate (G6).
##STR00091##
[0270] tert-Butyl 4-(3-aminophenyl)piperazine-1-carboxylate (G9)
can be prepared by coupling of commercially available tert-butyl
piperazine-1-carboxylate (G7) and compounds of the formula F4,
where L.sup.3=I or Br, in a Buchwald type reaction to give
tert-butyl 4-(3-nitrophenyl)piperazine-1-carboxylate (G8).
Reduction with hydrogen in the presence of a catalyst, for example
palladium on charcoal, gives tert-butyl
4-(3-aminophenyl)piperazine-1-carboxylate (G9).
##STR00092##
[0271] The corresponding 4-piperidine analogues of G6 can be
prepared by a sequence of reactions starting with the conversion of
commercially available tert-butyl 4-oxopiperidine-1-carboxylate
(G10) to vinyl triflate G11. Coupling of G11 in a Suzuki type
reaction with (4-nitrophenyl)boronic acid (G12) gives
tetrahydropyridine G13. Subsequent reduction via hydrogenation in
the presence of a catalyst, for example palladium on charcoal,
gives tert-butyl 4-(4-aminophenyl)piperidine-1-carboxylate
(G14).
##STR00093##
[0272] The corresponding 4-(3-aminophenyl)piperidine analogue of G9
can be prepared by a sequence of reactions starting with the
conversion of commercially available tert-butyl
4-oxopiperidine-1-carboxylate (G10) to vinyl triflate G11. Coupling
of G11 in a Suzuki type reaction with (3-nitrophenyl)boronic acid
(G15) gives tetrahydropyridine G16. Subsequent reduction via
hydrogenation in the presence of a catalyst, for example palladium
on charcoal, gives tert-butyl
4-(3-aminophenyl)piperidine-1-carboxylate (G17).
##STR00094##
[0273] The 3-(4-aminophenyl)piperidine regioisomers of G14 can be
prepared by reaction of commercially available compounds of the
formula F5, where L.sup.3=I or Br, with pyridin-3-ylboronic acid
(G18) in a Suzuki type reaction to form 3-(4-nitrophenyl)pyridine
(G19). Reduction of G19 with hydrogen in the presence of a
catalyst, for example platinum oxide, gives
4-(piperidin-3-yl)aniline (G20) which may be protected using Boc
anhydride to give tert-butyl
3-(4-aminophenyl)piperidine-1-carboxylate (G21).
##STR00095##
[0274] The 2-(4-aminophenyl)piperidine regioisomer of G14 can be
prepared by reaction of commercially available compounds of the
formula F5, where L.sup.3=I or Br, with pyridin-2-ylboronic acid
(G22) in a Suzuki type reaction to form 2-(4-nitrophenyl)pyridine
(G23). Reduction of G23 with hydrogen in the presence of a
catalyst, for example platinum oxide, gives
4-(piperidin-2-yl)aniline (G24) which may be protected using Boc
anhydride to give tert-butyl
2-(4-aminophenyl)piperidine-1-carboxylate (G25).
##STR00096##
[0275] Commercially available tert-butyl
3-oxopyrrolidine-1-carboxylate (G26) can be converted to a mixture
of vinyl triflates G27 and G28 in the presence of a triflamide and
a suitable base, for example NaHMDS. Coupling of the mixture with
(4-nitrophenyl)boronic acid (G12) under Suzuki conditions gives
dihydropyrroles G29 and G30. Reduction of this mixture using
hydrogen in the presence of a catalyst, for example 10% palladium
on charcoal, gives tert-butyl
3-(4-aminophenyl)pyrrolidine-1-carboxylate (G31).
##STR00097##
[0276] A metal/silyl mediated coupling of commercially available
tert-butyl 3-iodoazetidine-1-carboxylate (G32) and
4-iodo-nitrobenzene (G33) gives tert-butyl
3-(4-nitrophenyl)azetidine-1-carboxylate (G34). Subsequent
reduction via hydrogenation in the presence of a catalyst, for
example palladium on charcoal, gives the tert-butyl
3-(4-aminophenyl)azetidine-1-carboxylate (G35).
##STR00098##
[0277] tert-Butyl (1-(4-aminophenyl)piperidin-4-yl)carbamate (G39)
can be prepared by nucleophilic aromatic substitution of
commercially available tert-butyl piperidin-4-ylcarbamate (G36) and
1-fluoro-4-nitrobenzene (G37) under thermal conditions to give
tert-butyl (1-(4-nitrophenyl)piperidin-4-yl)carbamate (G38).
Reduction of G38 with hydrogen in the presence of a catalyst, for
example 10% palladium on charcoal gives tert-butyl
(1-(4-aminophenyl)piperidin-4-yl)carbamate (G39).
##STR00099##
[0278] Commercially available 2-bromo-1-(4-nitrophenyl)ethanone
(G40) can be reduced and cyclised to give epoxide G41. Opening of
the epoxide with tosylamide followed by cyclisation with
(2-bromoethyl)diphenylsulfonium trifluoromethanesulfonate gives
morpholine G43. Cleavage of the sulphonamide and subsequent
re-protection with Boc anhydride gives carbamate G45. Reduction
using hydrazine in the presence of iron(III) chloride gives
tert-butyl 2-(4-aminophenyl)morpholine-4-carboxylate (G46).
##STR00100##
tert-Butyl 4-(4-aminobenzyl)piperazine-1-carboxylate (G49) can be
prepared by the nucleophilic displacement of commercially available
1-(bromomethyl)-4-nitrobenzene (G47) with tert-butyl
piperazine-1-carboxylate (G7) to give tert-butyl
4-(4-nitrobenzyl)piperazine-1-carboxylate (G48). Subsequent
reduction with hydrogen in the presence of a catalyst, for example
10% palladium on charcoal, gives tert-butyl
4-(4-aminobenzyl)piperazine-1-carboxylate (G49).
[0279] Compounds of the formula F2 containing benzylamine or
substituted benzylamines may either be purchased with suitable
protecting groups in place to allow selective reaction at the
aniline or synthesised using an Ellman type procedure as out lined
in scheme N.
##STR00101##
[0280] Carbonyl compounds of the formula F6 can be reacted with
2-methylpropane-2-sulfinamide (G50) to give compounds of the
formula F7. Compounds of the formula F7 can be reacted with anions
prepared from suitably protected amino compounds, for example
N-(4-bromophenyl)-1,1,1-trimethyl-N-(trimethylsilyl)silanamine
(G51) treated with n-butyllithium, to give compounds of the formula
F8. Hydrolysis of compounds of the formula F8 under acidic
conditions, for example using aqueous hydrochloric acid, gives
compounds of the formula F9. Where necessary, compounds of the
formula F9 can be further protected to facilitate regiospecific
reactivity. It will be appreciated that Q.sup.1 and Q.sup.2 may be
the same or different and may be fused together to form a ring
structure, for example as in cyclobutanone. Substituents Q.sup.1
and Q.sup.2 form either R.sup.C1 or part of X in compounds of
formula I. It will also be appreciated that anions of suitably
protected amino heterocycles may be added to compounds of the
formula F7 to give heterocyclic analogues of compounds of the
formula F9.
[0281] Where compounds are required where R.sup.3 is aryl or
substituted aryl compounds of the formula F14 may be prepared as
outlined in scheme O.
##STR00102##
[0282] Compounds of the formula F10, F11 and F12, where R.sup.10
are independently H, F, Me or CF.sub.3; Q.sup.3 may be OH, O-alkyl,
NH.sub.2 or substituted N and X=Cl, Br or I, are either
commercially available or may be prepared synthetically. It will be
appreciated that for compounds of the formula F10, F11 and F12 that
the nature of Q.sup.3 can be readily changed. For example, a
carboxylic acid may be converted to a corresponding ester or amide
as required and conversely esters and amides can be hydrolysed to
give carboxylic acids. Compounds of the formula F10 where Q.sup.3
is an ester, may be deprotonated using a suitable base, for example
LDA and the resulting anion quenched with an alkylating agent, for
example methyl iodide, to give compounds of the formula F11 where Y
is a monoalkylated species. A second deprotonation can then be
carried out allowing the introduction of a second alkylating agent,
which may be the same or different to the first alkylating agent
employed, or to facilitate the cyclisation of a ring system where
the first alkylating agent used was di-functional, for example
1,3-dibromopropane. Halogenation of compounds of the formula F11
gives compounds of the formula F12, for example
1-phenyl-1-cyclopropanecarboxylic acid can be readily iodinated in
the presence of Pd(OAc).sub.2 and (diacetoxyiodo)benzene to give
1-(2-iodophenyl)cyclopropanecarboxylic acid. Compounds of the
formula F12 may be reacted under Sonagashira type coupling
conditions to give acetylenes of the formula F13 where R.sup.9=TMS,
TES or C(CH.sub.3).sub.2OH. R.sup.9 may then be removed to generate
compounds of the formula F14. When R.sup.9=TMS or TES potassium
carbonate or tetra-n-butyl ammonium fluoride may be employed to
induce this transformation. When R.sup.9=C(CH.sub.3).sub.2OH,
sodium hydride in refluxing toluene may be used.
[0283] Alternatively, when compounds in which R.sup.3=heteroaryl
are desired heteroaryl analogues of F14 may be prepared as outlined
in Schemes P, Q and R.
##STR00103##
[0284] For pyrazine containing analogues, 2,3-di-chloropyrazine
(G52) can be reacted with ethyl acetate in the presence of LiHMDS
to give ester G53. Deprotonation and alkylation, as described above
for aryl analogues, gives compounds of the formula F15. Coupling of
compounds of the formula F15 with TMS acetylene under Sonagashira
conditions gives acetylenes of the formula F16. Removal of the
trimethylsilyl group using TBAF gives compounds of the formula
F17.
##STR00104##
[0285] For pyrimidine analogues, diethyl succinate (G54) and ethyl
formate (G55) can be condensed to give aldehyde G56 in the presence
of sodium metal. Cyclisation using thiourea gives
4-oxo-2-thioxo-1,2,3,4-tetrahydropyrimidine (G57). Desulfurisation
using Raney-nickel gives pyrimidone G58, which can be converted to
4-chloro pyrimidine G59 using phosphorous oxychloride.
Deprotonation and alkylation, as described above for aryl
analogues, gives compounds of the formula F18. Coupling of
compounds of the formula F18 with TES-acetylene under Sonagashira
conditions, followed by removal of the triethylsilyl group using
TBAF gives compounds of the formula F20. It will be appreciated
that the regioisomeric pyrimidine can be accessed by analogous
series of reactions from the isomer of G57.
##STR00105##
[0286] For 3-pyridyl acetates, 2-(pyridin-3-yl)acetonitrile (G60)
can be oxidised to N-oxide G61. Chlorination with phosphorous
oxychloride gives 2-chloropyridine G62 which can be hydrolysed with
sodium hydroxide to acetic acid G63. Ester formation using methanol
gives 2-chloropyridine ester G64. Deprotonation and alkylation, as
described above for aryl analogues, gives compounds of the formula
F21. Coupling of compounds of the formula F21 with TES-acetylene
under Sonagashira conditions, followed by removal of the
triethylsilyl group using TBAF gives compounds of the formula F23.
It will be appreciated that the other regioisomeric pyridine
analogues can be prepared using an analogous sequence starting from
other commercially available pyridyl acetates.
##STR00106##
[0287] Pyrimidines of the formula F3 may be reacted with terminal
acetylenes of the formula F14 to give acetylenes of the formula F24
in a Sonagashira type coupling. The acetylene in compounds of the
formula F24 may be reduced to an alkane of the formula F25 using
hydrogen gas in the presence of a transition metal catalyst. The
exact choice of catalyst and conditions employed is dependant on
the nature of R.sup.2. For example, where R.sup.2=F, CF.sub.3,
methyl or methoxy, 10% Pd/C may be used, where R.sup.2=Cl, platinum
oxide is employed. Functional group manipulation may be carried on
compounds of the formula F25 if necessary. For example, compounds
of the formula F25 where Q.sup.3=O-alkyl (i.e. esters) may then be
deprotected to give carboxylic acids of the formula F25 where
Q.sup.3=OH. In esters where Q.sup.3=OMe, LiOH solutions may be
employed. Where Q.sup.3=Ot-Bu, acidic solutions, for example TFA in
DCM may be used to facilitate hydrolysis. It will be appreciated
that under acidic conditions Boc protecting groups in A will also
be cleaved.
[0288] Compounds of the formula F25 where Q.sup.3=OH may then be
converted to amides and substituted amides as described in formula
(1) using a suitable choice of amine in the presence of a coupling
agents for example EDCl.HCl or HATU.
[0289] It will be appreciated that heteroaromatic analogues of
compounds of the formula F14 (as described in schemes P, Q and R)
may be coupled in an analogous manner to that described in scheme S
and then further elaborated to amides as described above.
[0290] Compounds of the formula F25, in which Q.sup.3=an amide or
substituted amide may then be further modified by derivitisation of
amine functionality present in A. For example, compounds of the
formula F25 where A was prepared as described in schemes C to M, in
which a tert-butyl carbamate is present, may be hydrolysed in the
presence of mild acid, for example TFA, to give the parent amine.
The amine functionality maybe further derivatised by reductive
alkylation with formaldehyde in the presence of sodium
triacetoxyborohydride to give N-Me analogues; by reductive
alkylation with acetaldehyde in the presence of sodium
triacetoxyborohydride to give N-Et analogues or the N-acetyl
analogues may be prepared by reaction with a suitable acylating
agent, for example acetic anhydride.
[0291] Alternatively, a complementary approach to that described
scheme S can be employed, where R.sup.2 is not CF.sub.3, whereby
pyrimidines of the formula F1 are initially coupled to acetylenes
of the formula F14 as detailed in scheme T.
##STR00107##
[0292] Pyrimidines of the formula F1 may be coupled to acetylenes
of the formula F14 to give acetylenes of the formula F26 in a
Sonagashira type coupling. Depending on the nature of R.sup.2 these
couplings may either be regioselective, or where mixtures are
obtained, regioisomers may be separated by chromatography. The
acetylene in compounds of the formula F26 may be reduced to an
alkane of the formula F27 using hydrogen gas in the presence of a
transition metal catalyst. The exact choice of catalyst and
conditions employed is dependant on the nature of R.sup.2. For
example, where R.sup.2=Me, 10% Pd/C may be used, where R.sup.2=Cl,
platinum oxide is employed. The desired amide may already be
present in compounds of the formula F14, or alternatively an ester
may be used and subsequently derivatised as described above.
##STR00108##
[0293] Compounds of the formula F27 may then be reacted with amino
compounds of the formula F2, prepared as described above, to give
compounds of the formula F25. Such couplings may either be mediated
under acidic conditions, for example using trifluoroacetic acid in
trifluoroethanol or using palladium catalysis in a Buchwald/Hartwig
type coupling.
[0294] Compounds of the formula 25 may then be further elaborated
as desired as described above.
##STR00109##
[0295] Aldehydes or ketones of formula F18 where R.sup.11 is a
hydrogen, an alkyl group or similar may be substituted with amines
to form compounds of formula F19. It will be appreciated that in
some cases Q.sup.4=Q.sup.5=H forming a primary amine. Alternatively
Q.sup.4=H and Q.sup.5=R to form a secondary amine. In some cases
Q.sup.4=Q.sup.5=R, which will provide a secondary amine that may
also have Q.sup.4 and Q.sup.5 fused together to form a ring
structure, for example but not limited to azetidine, pyrrolidine,
piperazine, morpholine and piperidine.
Use of Compounds of the Invention
[0296] The present invention provides active compounds,
specifically, active 2,4,5-substituted pyrimidines.
[0297] The term "active", as used herein, pertains to compounds
which are capable of inhibiting VEGFR3 activity and specifically
includes both compounds with intrinsic activity (drugs) as well as
prodrugs of such compounds, which prodrugs may themselves exhibit
little or no intrinsic activity.
[0298] Assays which may be used in order to assess the VEGFR3
inhibition offered by a particular compound are described in the
examples below.
[0299] The present invention further provides a method of
inhibiting VEGFR3 activity in a cell, comprising contacting said
cell with an effective amount of an active compound, preferably in
the form of a pharmaceutically acceptable composition. Such a
method may be practised in vitro or in vivo.
[0300] The present invention further provides active compounds
which inhibit VEGFR3 activity, as well as methods of inhibiting
VEGFR3 activity, comprising contacting a cell with an effective
amount of an active compound, whether in vitro or in vivo.
[0301] Active compounds may also be used as part of an in vitro
assay, for example, in order to determine whether a candidate host
is likely to benefit from treatment with the compound in
question.
[0302] The invention further provides active compounds for use in a
method of treatment of the human or animal body. Such a method may
comprise administering to such a subject a
therapeutically-effective amount of an active compound, preferably
in the form of a pharmaceutical composition.
[0303] The term "treatment", as used herein in the context of
treating a condition, pertains generally to treatment and therapy,
whether of a human or an animal (e.g. in veterinary applications),
in which some desired therapeutic effect is achieved, for example,
the inhibition of the progress of the condition, and includes a
reduction in the rate of progress, a halt in the rate of progress,
amelioration of the condition, and cure of the condition. Treatment
as a prophylactic measure (i.e. prophylaxis) is also included.
[0304] The term "therapeutically-effective amount" as used herein,
pertains to that amount of an active compound, or a material,
composition or dosage from comprising an active compound, which is
effective for producing some desired therapeutic effect,
commensurate with a reasonable benefit/risk ratio.
Cancer
[0305] The present invention provides active compounds which are
anticancer agents. One of ordinary skill in the art is readily able
to determine whether or not a candidate compound treats a cancerous
condition for any particular cell type, either alone or in
combination.
[0306] The invention provides the use of the active compounds for
the treatment of cancer in the human or animal body. The invention
further provides active compounds for use in a method of treatment
of cancer in the human or animal body. Such a use or method may
comprise administering to such a subject a
therapeutically-effective amount of an active compound, preferably
in the form of a pharmaceutical composition.
[0307] Examples of cancers include, but are not limited to, bone
cancer, brain stem glioma, breast Cancer, cancer of the adrenal
gland, cancer of the anal region, cancer of the bladder, cancer of
the endocrine system, cancer of the oesophagus, cancer of the head
or neck, cancer of the kidney or ureter, cancer of the liver,
cancer of the parathyroid gland, cancer of the penis, cancer of the
small intestine, cancer of the thyroid gland, cancer of the
urethra, carcinoma of the cervix, carcinoma of the endometrium,
carcinoma of the fallopian tubes, carcinoma of the renal pelvis,
carcinoma of the vagina, carcinoma of the vulva, chronic or acute
leukemia, colon cancer, melanoma such as cutaneous or intraocular
melanoma, haemetological malignancies, Hodgkin's disease, lung
cancer, lymphocytic lymphomas, neoplasms of the central nervous
system (CNS), ovarian cancer, pancreatic cancer, pituitary adenoma,
primary CNS lymphoma, prostate cancer, rectal cancer, renal cell
carcinoma, sarcoma of soft tissue, skin cancer, spinal axis tumors,
stomach cancer and uterine cancer. In some embodiments, the cancer
is melanoma, breast cancer or head and neck cancer.
[0308] Any type of cell may be treated, including but not limited
to, lung, gastrointestinal (including, e.g., bowel, colon), breast
(mammary), ovarian, prostate, liver (hepatic), kidney (renal),
bladder, pancreas, brain, and skin.
[0309] Compounds of the present invention may also be useful in
inhibiting lymphangiogenesis and/or suppressing lymph node
metastasis. Compounds of the present invention may also be useful
in preventing the spread of cancer and in the prevention of
metastisis.
[0310] In one embodiment there is provided the use of a compound of
formula (I) or an isomer, salt, solvate, protected form or prodrug
thereof to prevent the spread of cancer or prevent metastasis.
There is also provided a compound of formula (I) or an isomer,
salt, solvate, protected form or prodrug thereof for use in a
method for preventing the spread of cancer or preventing of
metastasis.
[0311] In another embodiment there is provided an anti-cancer
treatment comprising a compound of formula (I) or an isomer, salt,
solvate, protected form or prodrug thereof and an anti-tumour
agent.
[0312] The anti cancer treatment defined herein may be applied as a
sole therapy or may involve, in addition to the compound of the
invention, conventional surgery or radiotherapy or chemotherapy.
Such chemotherapy may include one or more of the following
categories of anti-tumour agents:--
(i) other antiproliferative/antineoplastic drugs and combinations
thereof, as used in medical oncology, such as alkylating agents
(for example cisplatin, oxaliplatin, carboplatin, cyclophosphamide,
nitrogen mustard, melphalan, chlorambucil, busulphan, temozolamide
and nitrosoureas); antimetabolites (for example gemcitabine and
antifolates such as fluoropyrimidines like 5 fluorouracil and
tegafur, raltitrexed, methotrexate, cytosine arabinoside, and
hydroxyurea); antitumour antibiotics (for example anthracyclines
like adriamycin, bleomycin, doxorubicin, daunomycin, epirubicin,
idarubicin, mitomycin-C, dactinomycin and mithramycin); antimitotic
agents (for example vinca alkaloids like vincristine, vinblastine,
vindesine and vinorelbine and taxoids like taxol and docetaxel
(Taxotere) and polokinase inhibitors); and topoisomerase inhibitors
(for example epipodophyllotoxins like etoposide and teniposide,
amsacrine, topotecan and camptothecin); (ii) cytostatic agents such
as antioestrogens (for example tamoxifen, fulvestrant, toremifene,
raloxifene, droloxifene and iodoxyfene), antiandrogens (for example
bicalutamide, flutamide, nilutamide and cyproterone acetate), LHRH
antagonists or LHRH agonists (for example goserelin, leuprorelin
and buserelin), progestogens (for example megestrol acetate),
aromatase inhibitors (for example as anastrozole, letrozole,
vorazole and exemestane) and inhibitors of 5*-reductase such as
finasteride; (iii) anti-invasion agents (for example c-Src kinase
family inhibitors like
4-(6-chloro-2,3-methylenedioxyanilino)-7-[2-(4-methylpiperazin-1-yl)ethox-
y]-5-tetrahydropyran-4-yloxyquinazoline (AZD0530; International
Patent Application WO 01/94341),
N-(2-chloro-6-methylphenyl)-2-{6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-met-
hylpyrimidin-4-ylamino}thiazole-5-carboxamide (dasatinib,
BMS-354825; J. Med. Chem., 2004, 47, 6658-6661 and
4-((2,4-dichloro-5-methoxyphenyl)amino)-6-methoxy-7-(3-(4-methylpiperazin-
-1-yl)propoxy)quinoline-3-carbonitrile (bosutinib, SKI-606; Cancer
research (2003), 63(2), 375-81), and metalloproteinase inhibitors
like marimastat, inhibitors of urokinase plasminogen activator
receptor function or antibodies to Heparanase); (iv) inhibitors of
growth factor function: for example such inhibitors include growth
factor antibodies and growth factor receptor antibodies (for
example the anti erbB2 antibody trastuzumab [HerceptinT], the
anti-EGFR antibody panitumumab, the anti erbB1 antibody cetuximab
[Erbitux, C225] and any growth factor or growth factor receptor
antibodies disclosed by Stern et al. Critical reviews in
oncology/haematology, 2005, Vol. 54, pp 11-29); such inhibitors
also include tyrosine kinase inhibitors, for example inhibitors of
the epidermal growth factor family (for example EGFR family
tyrosine kinase inhibitors such as
N-(3-chloro-4-fluorophenyl)-7-methoxy-6-(3-morpholinopropoxy)quinazolin-4-
-amine (gefitinib, ZD1839),
N-(3-ethynylphenyl)-6,7-bis(2-methoxyethoxy)quinazolin-4-amine
(erlotinib, OSI 774) and
6-acrylamido-N-(3-chloro-4-fluorophenyl)-7-(3-morpholinopropoxy)-quinazol-
in-4-amine (CI 1033), erbB2 tyrosine kinase inhibitors such as
lapatinib, inhibitors of the hepatocyte growth factor family,
inhibitors of the platelet-derived growth factor family such as
imatinib, inhibitors of serine/threonine kinases (for example
Ras/Raf signalling inhibitors such as farnesyl transferase
inhibitors, for example sorafenib (BAY 43-9006)), inhibitors of
cell signalling through MEK and/or AKT kinases, inhibitors of the
hepatocyte growth factor family, c-kit inhibitors, abl kinase
inhibitors, IGF receptor (insulin-like growth factor) kinase
inhibitors; aurora kinase inhibitors (for example AZD1152,
PH739358, VX-680, MLN8054, R763, MP235, MP529, VX-528 AND AX39459)
and cyclin dependent kinase inhibitors such as CDK2 and/or CDK4
inhibitors; (v) antiangiogenic and antilymphangiogenic agents such
as those which inhibit the effects of vascular endothelial growth
factor, [for example the anti vascular endothelial cell growth
factor A (VEGFA) antibody bevacizumab (AvastinT), the anti vascular
endothelial cell growth factor A (VEGFA) antibody ranibizumab, the
anti-VEGF aptamer pegaptanib, the anti vascular endothelial growth
factor receptor 3 (VEGFR3) antibody IMC-3C5, the anti vascular
endothelial cell growth factor C (VEGFC) antibody VGX-100, the anti
vascular endothelial cell growth factor D (VEGFD) antibody VGX-200,
the soluble form of the vascular endothelial growth factor receptor
3 (VEGFR3) VGX-300 and VEGF receptor tyrosine kinase inhibitors
such as
4-(4-bromo-2-fluoroanilino)-6-methoxy-7-(1-methylpiperidin-4-ylmethoxy)qu-
inazoline (vandetanib; ZD6474; Example 2 within WO 01/32651),
4-(4-fluoro-2-methylindol-5-yloxy)-6-methoxy-7-(3-pyrrolidin-1-ylpropoxy)-
quinazoline (cediranib; AZD2171; Example 240 within WO 00/47212),
vatalanib (PTK787; WO 98/35985), pazopanib (GW786034), axitinib
(AG013736), sorafenib and sunitinib (SU11248; WO 01/60814),
compounds such as those disclosed in International Patent
Applications WO97/22596, WO 97/30035, WO 97/32856 and WO 98/13354
and compounds that work by other mechanisms (for example linomide,
inhibitors of integrin avb3 function and angiostatin)]; (vi)
vascular damaging agents such as Combretastatin A4 and compounds
disclosed in International Patent Applications WO 99/02166, WO
00/40529, WO 00/41669, WO 01/92224, WO 02/04434 and WO 02/08213;
(vii) antisense therapies, for example those which are directed to
the targets listed above, such as ISIS 2503, an anti-ras antisense;
(viii) gene therapy approaches, including for example approaches to
replace aberrant genes such as aberrant p53 or aberrant BRCA1 or
BRCA2, GDEPT (gene directed enzyme pro drug therapy) approaches
such as those using cytosine deaminase, thymidine kinase or a
bacterial nitroreductase enzyme and approaches to increase patient
tolerance to chemotherapy or radiotherapy such as multi drug
resistance gene therapy; and (ix) immunotherapy approaches,
including for example ex vivo and in vivo approaches to increase
the immunogenicity of patient tumour cells, such as transfection
with cytokines such as interleukin 2, interleukin 4 or granulocyte
macrophage colony stimulating factor, approaches to decrease T cell
anergy, approaches using transfected immune cells such as cytokine
transfected dendritic cells, approaches using cytokine transfected
tumour cell lines and approaches using anti idiotypic
antibodies
[0313] A combination of particular interest is with docetaxel.
Other possible combinations of interest include with gemcitabine,
cisplatin and the camptothecin prodrug irinotecan.
Diseases Ameliorated by the Control and/or Inhibition of
Lymphangiogenesis
[0314] The present invention provides active compounds which are
useful in preventing and/or treating diseases or conditions
ameliorated by the control and/or inhibition of
lymphangiogenesis.
[0315] In one embodiment there is provided the use of a compound of
formula (I) or an isomer, salt, solvate, protected form or prodrug
thereof to inhibit, suppress or reduce lymphangiogenesis. There is
also provided a compound of formula (I) or an isomer, salt,
solvate, protected form or prodrug thereof for use in the method of
inhibiting, suppressing or reducing lymphangiogenesis.
[0316] As discussed above, these diseases or conditions may
include:
(a) eye diseases, for example corneal graft rejection and age
related macular degeneration; (b) skin inflammations, such as skin
lesions in patients with psoriasis; (c) rejection in renal
transplantation.
Administration
[0317] The active compound or pharmaceutical composition comprising
the active compound may be administered to a subject by any
convenient route of administration, whether
systemically/peripherally or at the site of desired action,
including but not limited to, oral (e.g. by ingestion); topical
(including e.g. transdermal, intranasal, ocular, buccal, and
sublingual); pulmonary (e.g. by inhalation or insufflation therapy
using, e.g. an aerosol, e.g. through mouth or nose); rectal;
vaginal; parenteral, for example, by injection, including
subcutaneous, intradermal, intramuscular, intravenous,
intraarterial, intracardiac, intrathecal, intraspinal,
intracapsular, subcapsular, intraorbital, intraperitoneal,
intratracheal, subcuticular, intraarticular, subarachnoid,
intravitreal and intrasternal; by implant of a depot, for example,
subcutaneously, intravitreal or intramuscularly. The subject may be
a eukaryote, an animal, a vertebrate animal, a mammal, a rodent
(e.g. a guinea pig, a hamster, a rat, a mouse), murine (e.g. a
mouse), canine (e.g. a dog), feline (e.g. a cat), equine (e.g. a
horse), a primate, simian (e.g. a monkey or ape), a monkey (e.g.
marmoset, baboon), an ape (e.g. gorilla, chimpanzee, orang-utan,
gibbon), or a human.
Formulations
[0318] While it is possible for the active compound to be
administered alone, it is preferable to present it as a
pharmaceutical composition (e.g. formulation) comprising at least
one active compound, as defined above, together with one or more
pharmaceutically acceptable carriers, adjuvants, excipients,
diluents, fillers, buffers, stabilisers, preservatives, lubricants,
or other materials well known to those skilled in the art and
optionally other therapeutic or prophylactic agents.
[0319] Thus, the present invention further provides pharmaceutical
compositions, as defined above, and methods of making a
pharmaceutical composition comprising admixing at least one active
compound, as defined above, together with one or more
pharmaceutically acceptable carriers, excipients, buffers,
adjuvants, stabilisers, or other materials, as described
herein.
[0320] The term "pharmaceutically acceptable" as used herein
pertains to compounds, materials, compositions, and/or dosage forms
which are, within the scope of sound medical judgement, suitable
for use in contact with the tissues of a subject (e.g. human)
without excessive toxicity, irritation, allergic response, or other
problem or complication, commensurate with a reasonable
benefit/risk ratio. Each carrier, excipient, etc. must also be
"acceptable" in the sense of being compatible with the other
ingredients of the formulation.
[0321] Suitable carriers, excipients, etc. can be found in standard
pharmaceutical texts, for example, Remington's Pharmaceutical
Sciences, 18th edition, Mack Publishing Company, Easton, Pa.,
1990.
[0322] The formulations may conveniently be presented in unit
dosage form and may be prepared by any methods well known in the
art of pharmacy. Such methods include the step of bringing into
association the active compound with the carrier which constitutes
one or more accessory ingredients. In general, the formulations are
prepared by uniformly and intimately bringing into association the
active compound with liquid carriers or finely divided solid
carriers or both, and then if necessary shaping the product.
[0323] Formulations may be in the form of liquids, solutions,
suspensions, emulsions, elixirs, syrups, tablets, losenges,
granules, powders, capsules, cachets, pills, ampoules,
suppositories, pessaries, ointments, gels, pastes, creams, sprays,
mists, foams, lotions, oils, boluses, electuaries, or aerosols.
[0324] Formulations suitable for oral administration (e.g. by
ingestion) may be presented as discrete units such as capsules,
cachets or tablets, each containing a predetermined amount of the
active compound; as a powder or granules; as a solution or
suspension in an aqueous or non-aqueous liquid; or as an
oil-in-water liquid emulsion or a water-in-oil liquid emulsion; as
a bolus; as an electuary; or as a paste.
[0325] A tablet may be made by conventional means, e.g.,
compression or moulding, optionally with one or more accessory
ingredients. Compressed tablets may be prepared by compressing in a
suitable machine the active compound in a free-flowing form such as
a powder or granules, optionally mixed with one or more binders
(e.g. povidone, gelatin, acacia, sorbitol, tragacanth,
hydroxypropylmethyl cellulose); fillers or diluents (e.g. lactose,
microcrystalline cellulose, calcium hydrogen phosphate); lubricants
(e.g. magnesium stearate, talc, silica); disintegrants (e.g. sodium
starch glycolate, cross-linked povidone, cross-linked sodium
carboxymethyl cellulose); surface-active or dispersing or wetting
agents (e.g. sodium lauryl sulfate); and preservatives (e.g. methyl
p-hydroxybenzoate, propyl p-hydroxybenzoate, sorbic acid). Moulded
tablets may be made by moulding in a suitable machine a mixture of
the powdered compound moistened with an inert liquid diluent. The
tablets may optionally be coated or scored and may be formulated so
as to provide slow or controlled release of the active compound
therein using, for example, hydroxypropylmethyl cellulose in
varying proportions to provide the desired release profile. Tablets
may optionally be provided with an enteric coating, to provide
release in parts of the gut other than the stomach.
[0326] Formulations suitable for topical administration (e.g.
transdermal, intranasal, ocular, buccal, and sublingual) may be
formulated as an ointment, cream, suspension, lotion, powder,
solution, past, gel, spray, aerosol, or oil. Alternatively, a
formulation may comprise a patch or a dressing such as a bandage or
adhesive plaster impregnated with active compounds and optionally
one or more excipients or diluents.
[0327] Formulations suitable for topical administration in the
mouth include losenges comprising the active compound in a
flavoured basis, usually sucrose and acacia or tragacanth;
pastilles comprising the active compound in an inert basis such as
gelatin and glycerin, or sucrose and acacia; and mouthwashes
comprising the active compound in a suitable liquid carrier.
[0328] Formulations suitable for topical administration to the eye
also include eye drops wherein the active compound is dissolved or
suspended in a suitable carrier, especially an aqueous solvent for
the active compound.
[0329] Formulations suitable for nasal administration, wherein the
carrier is a solid, include a coarse powder having a particle size,
for example, in the range of about 20 to about 500 microns which is
administered in the manner in which snuff is taken, i.e. by rapid
inhalation through the nasal passage from a container of the powder
held close up to the nose. Suitable formulations wherein the
carrier is a liquid for administration as, for example, nasal
spray, nasal drops, or by aerosol administration by nebuliser,
include aqueous or oily solutions of the active compound.
[0330] Formulations suitable for administration by inhalation
include those presented as an aerosol spray from a pressurised
pack, with the use of a suitable propellant, such as
dichlorodifluoromethane, trichlorofluoromethane,
dichoro-tetrafluoroethane, carbon dioxide, or other suitable
gases.
[0331] Formulations suitable for topical administration via the
skin include ointments, creams, and emulsions. When formulated in
an ointment, the active compound may optionally be employed with
either a paraffinic or a water-miscible ointment base.
Alternatively, the active compounds may be formulated in a cream
with an oil-in-water cream base. If desired, the aqueous phase of
the cream base may include, for example, at least about 30% w/w of
a polyhydric alcohol, i.e., an alcohol having two or more hydroxyl
groups such as propylene glycol, butane-1,3-diol, mannitol,
sorbitol, glycerol and polyethylene glycol and mixtures thereof.
The topical formulations may desirably include a compound which
enhances absorption or penetration of the active compound through
the skin or other affected areas. Examples of such dermal
penetration enhancers include dimethylsulfoxide and related
analogues.
[0332] When formulated as a topical emulsion, the oily phase may
optionally comprise merely an emulsifier (otherwise known as an
emulgent), or it may comprises a mixture of at least one emulsifier
with a fat or an oil or with both a fat and an oil. Preferably, a
hydrophilic emulsifier is included together with a lipophilic
emulsifier which acts as a stabiliser. It is also preferred to
include both an oil and a fat. Together, the emulsifier(s) with or
without stabiliser(s) make up the so-called emulsifying wax, and
the wax together with the oil and/or fat make up the so-called
emulsifying ointment base which forms the oily dispersed phase of
the cream formulations.
[0333] Suitable emulgents and emulsion stabilisers include Tween
60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl
monostearate and sodium lauryl sulphate. The choice of suitable
oils or fats for the formulation is based on achieving the desired
cosmetic properties, since the solubility of the active compound in
most oils likely to be used in pharmaceutical emulsion formulations
may be very low. Thus the cream should preferably be a non-greasy,
non-staining and washable product with suitable consistency to
avoid leakage from tubes or other containers. Straight or branched
chain, mono- or dibasic alkyl esters such as di-isoadipate,
isocetyl stearate, propylene glycol diester of coconut fatty acids,
isopropyl myristate, decyl oleate, isopropyl palmitate, butyl
stearate, 2-ethylhexyl palmitate or a blend of branched chain
esters known as Crodamol CAP may be used, the last three being
preferred esters. These may be used alone or in combination
depending on the properties required.
[0334] Alternatively, high melting point lipids such as white soft
paraffin and/or liquid paraffin or other mineral oils can be
used.
[0335] Formulations suitable for rectal administration may be
presented as a suppository with a suitable base comprising, for
example, cocoa butter or a salicylate.
[0336] Formulations suitable for vaginal administration may be
presented as pessaries, tampons, creams, gels, pastes, foams or
spray formulations containing in addition to the active compound,
such carriers as are known in the art to be appropriate.
[0337] Formulations suitable for parenteral administration (e.g. by
injection, including cutaneous, subcutaneous, intramuscular,
intravenous and intradermal), include aqueous and non-aqueous
isotonic, pyrogen-free, sterile injection solutions which may
contain anti-oxidants, buffers, preservatives, stabilisers,
bacteriostats, and solutes which render the formulation isotonic
with the blood of the intended recipient; and aqueous and
non-aqueous sterile suspensions which may include suspending agents
and thickening agents, and liposomes or other microparticulate
systems which are designed to target the compound to blood
components or one or more organs. Examples of suitable isotonic
vehicles for use in such formulations include Sodium Chloride
Injection, Ringer's Solution, or Lactated Ringer's Injection.
Typically, the concentration of the active compound in the solution
is from about 1 ng/mL to about 10 .mu.g/mL, for example from about
10 ng/ml to about 1 .mu.g/mL. The formulations may be presented in
unit-dose or multi-dose sealed containers, for example, ampoules
and vials, and may be stored in a freeze-dried (lyophilised)
condition requiring only the addition of the sterile liquid
carrier, for example water for injections, immediately prior to
use. Extemporaneous injection solutions and suspensions may be
prepared from sterile powders, granules, and tablets. Formulations
may be in the form of liposomes or other microparticulate systems
which are designed to target the active compound to blood
components or one or more organs.
Dosage
[0338] It will be appreciated that appropriate dosages of the
active compounds, and compositions comprising the active compounds,
can vary from patient to patient. Determining the optimal dosage
will generally involve the balancing of the level of therapeutic
benefit against any risk or deleterious side effects of the
treatments of the present invention. The selected dosage level will
depend on a variety of factors including, but not limited to, the
activity of the particular compound, the route of administration,
the time of administration, the rate of excretion of the compound,
the duration of the treatment, other drugs, compounds, and/or
materials used in combination, and the age, sex, weight, condition,
general health, and prior medical history of the patient. The
amount of compound and route of administration will ultimately be
at the discretion of the physician, although generally the dosage
will be to achieve local concentrations at the site of action which
achieve the desired effect without causing substantial harmful or
deleterious side-effects.
[0339] Administration in vivo can be effected in one dose,
continuously or intermittently (e.g. in divided doses at
appropriate intervals) throughout the course of treatment. Methods
of determining the most effective means and dosage of
administration are well known to those of skill in the art and will
vary with the formulation used for therapy, the purpose of the
therapy, the target cell being treated, and the subject being
treated. Single or multiple administrations can be carried out with
the dose level and pattern being selected by the treating
physician.
[0340] In general, a suitable dose of the active compound is in the
range of about 100 .mu.g to about 250 mg per kilogram body weight
of the subject per day. Where the active compound is a salt, an
ester, prodrug, or the like, the amount administered is calculated
on the basis of the parent compound and so the actual weight to be
used is increased proportionately.
EXAMPLES
[0341] The following examples are provided solely to illustrate the
present invention and are not intended to limit the scope of the
invention, as described herein. In the examples below, in case the
structures contain one or more stereogenic centres and the
sterereochemistry is depicted in the diagram, the respective
stereochemistry is assigned in an arbitrary absolute configuration.
These structures depict single enantiomers as well as mixtures of
enantiomers in all ratios, and/or mixtures of diastereoisomers in
all ratios.
Acronyms
[0342] For convenience, many chemical moieties are represented
using well known abbreviations, including but not limited to,
methyl (Me), ethyl (Et), n-propyl (nPr), iso-propyl (iPr), n-butyl
(nBu), tert-butyl (tBu), n-hexyl (nHex), cyclohexyl (cHex), phenyl
(Ph), methoxy (MeO), ethoxy (EtO), trimethylsilyl (TMS),
tert-butyloxycarbonyl (Boc), and acetyl (Ac).
[0343] For convenience, many chemical compounds are represented
using well known abbreviations, including but not limited to,
methanol (MeOH), ethanol (EtOH), ether or diethyl ether
(Et.sub.2O), ethyl acetate (EtOAc), triethylamine (Et.sub.3N),
dichloromethane (methylene chloride, DCM), trifluoroacetic acid
(TFA), N,N-dimethylformamide (DMF), sodium sulphate
(Na.sub.2SO.sub.4), tetrahydrofuran (THF), dimethylsulfoxide
(DMSO), magnesium sulphate (MgSO.sub.4), sodium hydrogen carbonate
(NaHCO.sub.3), tert-butanol (t-BuOH),
1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride salt
(EDCl.HCl), tetra-n-butylammonium fluoride (TBAF),
meta-chloroperbenzoic acid (mCPBA), hexamethyldisilazane sodium
salt (NaHMDS), N,N-diisopropylethylamine (DI PEA),
1-hydroxybenzotriazole (HOBt),
O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate (HATU),
tris(dibenzylideneacetone)dipalladium(0) (Pd.sub.2(dba).sub.3),
trans-dichlorobis(triphenylphosphine)palladium(II)
(PdCl.sub.2(PPh.sub.3).sub.2), palladium(II) acetate
(Pd(OAc).sub.2)tri-tert-butyl phosphonium tetrafluoroborate
(t-Bu.sub.3PH.BF.sub.4),
4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (Xantphos),
triphenylphosphine (PPh.sub.3) and 1,2-dichloroethane (DCE).
General Experimental Details
[0344] Unless otherwise stated the following generalisations
apply.
[0345] .sup.1NMR spectra were recorded on either a Bruker Avance
DRX300 (300 MHz) or a Bruker Ultrashield plus (400 MHz). The
multiplicity of a signal is designated by the following
abbreviations: s, singlet; d, doublet; dd, doublet of doublets; t,
triplet; tt, triplet of triplets; td, triplet of doublets; q,
quartet; br, broad; m, multiplet. All observed coupling constants,
J, are reported in Hertz.
[0346] LC/MS data was generated using either an Agilent 6100 Series
Single Quad LC/MS (LCMS-A) or Waters ZQ 3100 system (LCMS-B) or
Finnigan LCG Advantage Max (LCMS-C) or Agilent 1200 Infinity Series
(LCMS-D).
LCMS Method A (LCMS-A)
Instrument: Agilent 6100 Series Single Quad LC/MS
Agilent 1200 Series HPLC
[0347] Pump: 1200 Series G1311A Quaternary pump
Autosampler: 1200 Series G1329A Thermostatted Autosampler
Detector: 1200 Series G1314B Variable Wavelength Detector
LC Conditions:
[0348] Reverse Phase HPLC analysis
Column: Luna C8(2) 5.mu. 50.times.4.6 mm 100 A
[0349] Column temperature: 30.degree. C.
Injection Volume: 5 .mu.L
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0350] Gradient: 5-100% B over 10 min
Detection: 254 nm or 214 nm
MS Conditions:
Ion Source: Quadrupole
Ion Mode: Multimode-ES
[0351] Drying gas temp: 300.degree. C. Vaporizer temperature:
200.degree. C. Capillary voltage (V): 2000 (positive) Capillary
voltage (V): 4000 (negative)
Scan Range: 100-1000
[0352] Step size: 0.1 sec Acquisition time: 10 min
LCMS Method B (LCMS-B)
Instrument: Waters ZQ 3100 Mass Detector
Waters 2545-Pump
Waters SFO System Fluidics Organizer
Waters 2996 Diode Array Detector
Waters 2767 Sample Manager
LC Conditions:
[0353] Reverse Phase HPLC analysis
Column: XBridge.TM. C18 5 .mu.m 4.6.times.100 mm
Injection Volume: 10 .mu.L
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0354] Gradient: 10-100% B over 10 min Flow rate: 1.5 ml/min
Detection: 100-600 nm
MS Conditions
Ion Source: Single-quadrupole
[0355] Ion Mode: ES positive
Source Temp: 150.degree. C.
Desolvation Temp: 350.degree. C.
[0356] Detection: Ion counting
Capillary (KV): 3.00
Cone (V): 30
Extractor (V): 3
RF Lens (V): 0.1
Scan Range: 100-1000 Amu
Scan Time: 0.5 sec
[0357] Acquisition time: 10 min
Gas Flow: 100 L/hr
Desolvation: 650 L/hr
LCMS Method C (LCMS-C)
Instrument: Finnigan LCG Advantage Max
Finnigan Surveyor LC Pump
Finnigan Surveyor Autosampler
Finnigan Surveyor PDA Detector
LC Conditions:
[0358] Reverse Phase HPLC analysis
Column: Gemini 3 .mu.m C18 20.times.4.0 mm 110 A
Injection Volume: 10 .mu.L
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0359] Gradient: 10-100% B over 10 min
Detection: 100-600 nm
MS Conditions
[0360] Ion Source: Ion trap Ion Mode: ES positive
Temp: 300.degree. C.
[0361] Detection: Ion counting
Scan Range: 80-1000 Amu
Scan Time: 0.2 sec
[0362] Acquisition time: 10 min
LCMS Method D (LCMS-D)
Instrument: Agilent 1200 Infinity Series
[0363] Pump: 1260 Infinity G1312B Binary pump
Auto Sampler: 1260 Infinity G1367E 1260 HiP ALS
Detector: 1290 Infinity G4212A 1290 DAD
LC Conditions:
[0364] Reverse Phase HPLC analysis
Column: Poroshell 120 EC-C18
[0365] Column temperature: 35.degree. C.
Injection Volume: 1 .mu.L
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0366] Gradient: 5-100% B over 3.8 mins Detection: monitored at 254
nm and 214 nm
MS Conditions:
Ion Source: Quadrupole
Ion Mode: API-ES
[0367] Drying gas temp: 350.degree. C. Capillary voltage (V): 3000
(positive) Capillary voltage (V): 3000 (negative)
Scan Range: 100-1000
[0368] Step size: 0.1 sec Acquisition time: 5 min
HPLC
Instrument: Waters Alliance HT
Detector: Waters 2996 Diode Array
Column: Reverse Phase Xbridge.TM. Prep C18 5 um 10.times.100 mm
Injection Volume: 50 uL
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0369] Flow rate: 4.0 ml/min
Detection: 100-600 nm
[0370] Sample preparation: MeCN and MeOH
Preparative Mass-Directed LC (Prep-LCMS)
Instrument: Waters ZQ 3100-Mass Detector
Waters 2545-Pump
Waters SFO System Fluidics Organizer
Waters 2996 Diode Array Detector
Waters 2767 Sample Manager
LC Conditions:
[0371] Reverse Phase HPLC analysis
Column: XBridge.TM. C18 5 .mu.m 19.times.50 mm
Injection Volume 500 .mu.L
Solvent A: Water 0.1% Formic Acid
Solvent B: Acetonitrile 0.1% Formic Acid
[0372] Gradient: 25-100% B over 10 min Flow rate: 19 mL/min
Detection: 100-600 nm
MS Conditions:
Ion Source: Single-quadrupole
[0373] Ion Mode: ES positive
Source Temp: 150.degree. C.
Desolvation Temp: 350.degree. C.
[0374] Detection: Ion counting
Capillary (KV)-3.00
Cone (V): 30
Extractor (V): 3
RF Lens (V): 0.1
Scan Range: 100-1000 Amu
Scan Time: 0.5 sec
[0375] Acquisition time: 10 min
Gas Flow
Desolvation L/hour-650
Cone L/hour-100
Chiral Separation and Characterisation Methods
Method A
SFC
[0376] Column: Chiralpak IC (250.times.21 mm), 5.mu. Flow Rate: 40
mL/min
Mobile Phase: CO.sub.2: 0.1% DEA in MeOH
Method B
SFC
[0377] Column: Chiralpak-IC (250.times.21 mm), 5.mu. Flow Rate: 40
mL/min
Mobile Phase: CO.sub.2: 0.5% DEA in IPA
Method C
SFC
Column: Chiralcel-ODH
[0378] Flow Rate: 3 mL/min
Mobile Phase: CO.sub.2: 0.5% DEA in IPA
Method D
HPLC
[0379] Column: Chiralpak IC (250.times.10 mm), 5.mu. Mobile Phase:
n-Hexane: ethanol (80:20) Flow rate: 5.0 mL/min
Method E
SFC
[0380] Column: Lux cellulose-4 (250.times.4.6 mm), 5.mu. Mobile
Phase: CO.sub.2: methanol (70:30) Flow rate: 3.0 mL/min
Method F
SFC
[0381] Column: Lux cellulose-2 (250.times.4.6 mm), 5.mu. Flow Rate:
1.2 mL/min Mobile Phase: CO.sub.2: 0.5% DEA in methanol
Method G
HPLC
[0382] Column: Chiralpak ADH (250.times.4.6 mm), 5.mu. Mobile
Phase: n-Hexane: ethanol (80:20) Flow rate: 1 mL/min
[0383] Analytical thin-layer chromatography was performed on Merck
silica gel 60F254 aluminium-backed plates which were visualised
using fluorescence quenching under UV light or using an acidic
anisaldehyde or a basic potassium permanganate dip. Flash
chromatography was performed using either a Teledyne Isco
CombiFlash Rf purification system using standard RediSep.RTM.
cartridges or a Biotage Isolera purification system using either
Grace, RediSep.RTM. or Biotage silica cartridges. Microwave
irradiation was achieved using a CEM Explorer SP Microwave
Reactor.
[0384] Where necessary, anhydrous solvents were prepared using a
Braun purification system or purchased from Sigma-Aldrich.
SYNTHESIS OF KEY INTERMEDIATES
Key Intermediate 1
2-(2-Ethynylphenyl)butanamide (K1)
##STR00110##
[0385] (a) 2-(2-Iodophenyl)butanoic acid (I1)
[0386] Lithium diisopropylamide solution (2.0 M in
THF/heptane/ethylbenzene, 3.82 mL, 7.63 mmol) was added to dry THF
(10 mL) under an atmosphere of nitrogen and cooled to 0.degree. C.
A solution of 2-(2-iodophenyl)acetic acid (500 mg, 1.91 mmol) in
dry THF (15 mL) was then added dropwise. This solution was stirred
for 40 minutes at 0.degree. C. before the addition of iodoethane
(0.92 mL, 11 mmol). The solution was returned to room temperature
and stirred for 4 hours. The resulting mixture was quenched with
the addition of H.sub.2O (10 mL) and then 2 M HCl (20 mL). The
aqueous layer was extracted with EtOAc (3.times.30 mL), the organic
layers were combined and washed with brine, dried over MgSO.sub.4
and the solvent was evaporated under reduced pressure. The residue
was adsorbed onto silica gel and purified using column
chromatography (Biotage Isolera, SiO.sub.2 cartridge, 0-40% EtOAc
in petroleum benzine 40-60.degree. C.) to give the title compound
I1 as a pale yellow oil (479 mg, 87%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 12.49 (s, 1H), 7.88 (dd, J=7.9, 1.2 Hz, 1H),
7.39 (td, J=7.6, 1.2 Hz, 1H), 7.32 (dd, J=7.8, 1.7 Hz, 1H), 7.01 (J
m, 1H), 3.77 (t, J=7.5 Hz, 1H), 1.98-1.86 (m, 1H), 1.73-1.60 (m,
1H), 0.85 (t, J=7.3 Hz, 3H).
(b) 2-(2-Iodophenyl)butanamide (I2)
[0387] HOBt (325 mg, 2.40 mmol), EDCl.HCl (461 mg, 2.40 mmol) and
DIPEA (1.40 mL, 8.02 mmol) were added to a stirred solution of
2-(2-iodophenyl)butanoic acid (I1) (465 mg, 1.60 mmol) in dry THF
(6 mL) and dry DMF (1 mL) under an atmosphere of nitrogen. After 10
minutes ammonium carbonate (770 mg, 8.02 mmol) was added in one
portion and the resulting mixture stirred at room temperature for
24 hours. The volatiles were removed in vacuo and EtOAc (50 mL) and
saturated aqueous NaHCO.sub.3 (50 mL) were added to the residue.
The aqueous phase was extracted with EtOAc (2.times.50 mL), then
the combined organic extracts were washed with brine and dried over
MgSO.sub.4. The solvent was removed under reduced pressure and the
resulting solid was purified by silica gel column chromatography
(Biotage Isolera, SiO.sub.2 cartridge, 0-50% EtOAc in petroleum
benzine 40-60.degree. C.) to give the title compound I2 as a white
solid (398 mg, 86%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta.
7.85 (dd, J=7.9, 1.2 Hz, 1H), 7.46 (dd, J=7.8, 1.7 Hz, 1H), 7.39
(s, 1H), 7.35 (td, J=7.6, 1.2 Hz, 1H), 7.02-6.92 (m, 2H), 3.59 (dd,
J=8.8, 6.0 Hz, 1H), 1.92-1.78 (m, 1H), 1.65-1.51 (m, 1H), 0.86 (t,
J=7.3 Hz, 3H). LCMS-A: rt 5.463 min; m/z 290 [M+H].sup.+.
(c) 2-(2-((Trimethylsilyl)ethynyl)phenyl)butanamide (I3)
[0388] Ethynyltrimethylsilane (0.11 mL, 0.80 mmol) and dry DMF (9.0
mL) was added to a mixture of 2-(2-iodophenyl)butanamide (I2) (193
mg, 0.668 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (23 mg, 0.033 mmol),
t-Bu.sub.3PH.BF.sub.4 (10 mg, 0.033 mmol) and CuI (6 mg, 0.033
mmol) under nitrogen. The resulting solution was degassed with a
stream of nitrogen for 10 minutes before the addition of Et.sub.3N
(3.0 mL). The resulting mixture was stirred at 60.degree. C. under
nitrogen for 16 hours then adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound I3 as a tan coloured solid (151 mg, 87); .sup.1H NMR (400
MHz, d.sub.6-DMSO) .delta. 7.46-7.39 (m, 2H), 7.34 (td, J=7.7, 1.4
Hz, 1H), 7.26 (br, 1H), 7.21 (td, J=7.5, 1.4 Hz, 1H), 6.97 (br,
1H), 3.80 (dd, J=8.9, 5.7 Hz, 1H), 1.95-1.80 (m, 1H), 1.69-1.55 (m,
1H), 0.85 (t, J=7.3 Hz, 3H), 0.25 (s, 9H). LCMS-A: rt 6.267 min;
m/z 260 [M+H].sup.+.
(d) 2-(2-Ethynylphenyl)butanamide (K1)
[0389] To a solution of
2-(2-((trimethylsilyl)ethynyl)phenyl)butanamide (I3) (149 mg, 0.574
mmol) in DCM (10.0 mL) at 0.degree. C. under nitrogen was added
TBAF (1.0 M in THF, 0.86 mL, 0.86 mmol). The resulting mixture was
stirred at 0.degree. C. for 5 minutes then poured into water (50
mL). The organic phase was separated and the aqueous layer was
extracted with DCM (2.times.40 mL). The combined organic layers
were washed with brine (40 mL), dried over MgSO.sub.4 and the
solvent was removed in vacuo. The residue was adsorbed onto silica
gel and purified by column chromatography (Biotage Isolera,
SiO.sub.2 cartridge, 0-75% EtOAc in petroleum benzine 40-60.degree.
C.) to give the title compound K1 as an off-white solid (104 mg,
97%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 7.50-7.43 (m,
2H), 7.35 (td, J=7.7, 1.5 Hz, 1H), 7.31 (s, 1H), 7.23 (td, J=7.5,
1.3 Hz, 1H), 6.93 (s, 1H), 4.37 (s, 1H), 3.85 (dd, J=8.6, 6.4 Hz,
1H), 1.97-1.83 (m, 1H), 1.61 (J m, 1H), 0.84 (t, J=7.3 Hz, 3H).
LCMS-A: rt 5.265 min; m/z 188 [M+H].sup.+.
Key Intermediate 2
Methyl 1-(2-ethynylphenyl)cyclopropanecarboxylate (K2)
##STR00111##
[0390] (a) Methyl 1-(2-bromophenyl)cyclopropanecarboxylate (I4)
[0391] A solution of 1-(2-bromophenyl)cyclopropanecarboxylic acid
(500 mg, 2.07 mmol) in MeOH (10 mL) was treated with a solution of
concentrated aqueous HCl (0.5 mL). The resulting mixture was
stirred for 16 hours at room temperature and then heated to reflux
and stirred for a further 24 hours. The volatiles were evaporated
and the residue was dissolved in EtOAc. The organic layer was
washed with saturated solution of NaHCO.sub.3, brine and then dried
over MgSO.sub.4. The solvent was removed in vacuo to give the title
compound I4 (400 mg, 76%) as an orange oil; .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.60-7.54 (m, 1H), 7.32-7.24 (m, 2H), 7.15 (m,
1H), 3.63 (s, 3H), 1.75 (d, J=3.1 Hz, 2H), 1.21 (q, J=4.0 Hz,
2H).
(b) Methyl
1-(2-((trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxylate
(I5)
[0392] A solution of methyl
1-(2-bromophenyl)cyclopropanecarboxylate (I4) (658 mg, 2.58 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (91 mg, 0.13 mmol)
t-Bu.sub.3PH.BF.sub.4 (37 mg, 0.13 mmol), CuI (25 mg, 0.13 mmol)
and ethynyltrimethylsilane (0.44 mL, 3.1 mmol) in dry DMF (5.0 mL)
was degassed with nitrogen for 10 minutes before the addition of
Et.sub.3N (2.0 mL). After stirring at 65.degree. C. under nitrogen
for 16 hours the resulting mixture was adsorbed onto silica gel and
purified by column chromatography (Biotage Isolera, SiO.sub.2
cartridge, 0-20% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound I5 as an orange oil (569 mg, 81%); .sup.1H
NMR (400 MHz, d.sub.6-DMSO) .delta. 7.67-7.12 (m, 4H), 3.57-3.47
(m, 3H), 1.64-1.49 (m, 2H), 1.26-1.16 (m, 2H), 0.22-0.13 (m, 9H).
LCMS-A: rt 6.977 min; m/z 273 [M+H].sup.+.
(c) Methyl 1-(2-ethynylphenyl)cyclopropanecarboxylate (K2)
[0393] To a solution of methyl
1-(2-((trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxylate (15)
(568 mg, 2.09 mmol) in DCM (20.0 mL) at 0.degree. C. was added TBAF
(1.0 M in THF, 3.13 mL, 3.13 mmol). The resulting mixture was
stirred at 0.degree. C. for 10 minutes then poured into water (50
mL). The aqueous layer was extracted with DCM (2.times.50 mL), then
the combined organic layers were washed with brine, dried over
MgSO.sub.4 and the solvent was removed in vacuo. The residue was
adsorbed onto silica gel and purified by column chromatography
(Biotage Isolera, SiO.sub.2 cartridge, 0-25% EtOAc in petroleum
benzine 40-60.degree. C.) to give the title compound K2 as an
off-white solid (140 mg, 34%); .sup.1H NMR (400 MHz, d.sub.6-DMSO)
.delta. 7.46 (dt, J=7.5, 0.8 Hz, 1H), 7.40-7.25 (m, 3H), 4.32 (s,
1H), 3.52 (s, 3H), 1.54 (q, J=4.1 Hz, 2H), 1.25-1.19 (m, 2H).
LCMS-A: rt 5.921 min; m/z 201 [M+H].sup.+.
Key Intermediate 3
tert-Butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)-
piperidine-1-carboxylate (K3)
##STR00112##
[0395] 2,4-Dichloro-5-(trifluoromethyl)pyrimidine (4.122 g, 19.00
mmol) was stirred in a 1:1 t-BuOH:DCE mixture (400 mL) at room
temperature. A 1.0 M ZnCl.sub.2 solution in Et.sub.2O (21.71 mL,
21.71 mmol) was added cautiously and the resulting mixture was
stirred for 10 minutes. 1-Boc-4-(4-aminophenyl)piperidine (5.00 g,
18.1 mmol) was added followed by Et.sub.3N (6.052 mL, 43.42 mmol)
and stirring continued at room temperature overnight. The volatiles
were evaporated to dryness and the resulting residue was suspended
in water (500 mL). After sonication for 30 minutes the suspension
was filtered and the filter cake was washed with water (2.times.100
mL) and dried under a high vacuum to yield the title compound K3 as
a tan solid (8.11 g, 98%); .sup.1H NMR (400 MHz, d.sub.6-DMSO)
.delta. 10.61 (s, 1H), 8.78 (s, 1H), 7.59 (d, J=8.4 Hz, 2H), 7.23
(d, J=8.5 Hz, 2H), 4.07 (d, J=11.1 Hz, 2H), 2.80 (s, 2H), 2.65 (t,
J=12.0 Hz, 1H), 1.74 (d, J=12.3 Hz, 2H), 1.42 (s, 11H). LCMS-A: rt
6.834 min; m/z 457 [M+H].sup.+.
Key Intermediate 4
tert-Butyl
3-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)-
azetidine-1-carboxylate (K4)
##STR00113##
[0396] (a) tert-Butyl-3-(4-nitrophenyl)azetidine-1-carboxylate
(16)
[0397] 1,2-Dibromoethane (0.146 mL, 1.69 mmol) was added to a
vigorously stirred suspension of zinc dust (0.901 g, 13.8 mmol) in
THF (3.5 mL) under a nitrogen atmosphere and the resulting
suspension heated at 80.degree. C. for 10 minutes. Trimethylsilyl
chloride (0.202 mL, 1.59 mmol) in THF (1.75 mL) was added at room
temperature and after stirring for 4 minutes a solution of
tert-butyl 3-iodoazetidine-1-carboxylate (3.00 g, 10.6 mmol) in THF
(3.5 mL) was added dropwise over a period of 15 minutes. The
resulting mixture was stirred at room temperature for 2 hours then
Pd.sub.2(dba).sub.3 (0.155 g, 0.170 mmol) and tri-2-furylphosphine
(0.143 g, 0.615 mmol) were added followed by 1-iodo-4-nitrobenzene
(2.90 g, 11.7 mmol) in THF (18 mL). The resulting mixture was
heated at 55.degree. C. for 3 hours then quenched at room
temperature with a saturated aqueous sodium chloride solution (15
mL). The aqueous phase was extracted with DCM (2.times.15 mL) then
the combined organic fractions were dried (magnesium sulfate),
filtered and evaporated in vacuo. The residue was purified using
silica gel column chromatography (CombiFlash Rf, 40 g SiO.sub.2
Cartridge, 10-40% EtOAc in cyclohexane) to give the title compound
I6 as an orange oil (2.14 g, 72%); .sup.1H NMR (300 MHz,
CDCl.sub.3) .delta. 8.24 (dd, J=6.8, 1.9 Hz, 2H), 7.51 (d, J=8.6
Hz, 2H), 4.41 (t, J=8.7 Hz, 2H), 3.98 (dd, J=8.5, 5.7 Hz, 2H),
3.89-3.81 (s, 1H), 1.49 (s, 9H).
(b) tert-Butyl 3-(4-aminophenyl)azetidine-1-carboxylate (I7)
[0398] A suspension of 10% Pd/C (0.320 g) and
tert-butyl-3-(4-nitrophenyl)azetidine-1-carboxylate (I6) (2.14 g,
7.68 mmol) in EtOAc (16 mL) was stirred under a hydrogen atmosphere
for 18 hours. Additional 10% Pd/C (1.00 g) was added and stirring
continued for a further 20 hours. The resulting mixture was
filtered through a pad of Celite, washing with EtOAc, and the
filtrate concentrated in vacuo to give the title compound I7 as a
light yellow/cream solid (1.80 g, 94%); .sup.1H NMR (300 MHz,
CDCl.sub.3) .delta. 7.12 (d, J=8.3 Hz, 2H), 6.69 (dd, J=6.5, 1.9
Hz, 2H), 4.29 (t, J=8.7 Hz, 2H), 3.93 (dd, J=8.4, 6.1 Hz, 2H), 3.65
(brs, 2H), 1.55-1.68 (m, 1H), 1.48 (s, 9H). LCMS-B: rt 4.964 min;
m/z 249 [M+H].sup.+.
(c) tert-Butyl
3-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)azetidine--
1-carboxylate (K4)
[0399] Zinc chloride (1.0 M in Et.sub.2O) (4.83 mL, 4.83 mmol) was
added to a solution of 2,4-dichloro-5-(trifluoromethyl)pyrimidine
(0.769 g, 3.54 mmol) in 1:1 dichloroethane/tert-butanol (64 mL) at
room temperature under nitrogen. After stirring for 10 minutes,
tert-butyl 3-(4-aminophenyl)azetidine-1-carboxylate (I7) (0.800 g,
3.22 mmol) was added followed by Et.sub.3N (1.08 mL, 7.73 mmol).
The resulting mixture was stirred at room temperature for 20 hours
then the volatiles removed in vacuo. Water was added to the solid
residue and the resulting suspension sonicatated for 2 minutes. The
suspension was filtered, and the filter cake dried then adsorbed
onto silica gel and purified using column chromatography
(CombiFlash Rf, 40 g SiO.sub.2 Cartridge, 10-40% EtOAc in
cyclohexane) to give a white solid. The solid was suspended in MeOH
(7 mL) and sonicated for 30 seconds. The resulting suspension was
filtered and the filter cake was washed with MeOH (3 mL) then dried
to give the title compound K4 as a white solid (0.777 g, 56%);
.sup.1H NMR (300 MHz, d.sub.6-DMSO) .delta. 10.6 (s, 1H), 8.79 (s,
1H), 7.66 (d, J=8.4 Hz, 2H), 7.33 (d, J=8.4 Hz, 2H), 4.23 (t, J=7.6
Hz, 2H), 3.80 (s, 3H), 1.40 (s, 9H). LCMS-B: rt 8.810 min; m/z 429
[M+H].sup.+.
Key Intermediate 5
tert-Butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-
-2-yl)piperidine-1-carboxylate (K5)
##STR00114##
[0400] (a) tert-Butyl
5-nitro-5',6'-dihydro-[2,4'-bipyridine]-1'(2'H)-carboxylate
(I8)
[0401] To a mixture of N-Boc-1,2,3,6-tetrahydropyridine-4-boronic
acid pinacol ester (1.52 g, 4.93 mmol), 2-bromo-5-nitropyridine
(1.00 g, 4.93 mmol) and PdCl.sub.2(PPh.sub.3).sub.2 (173 mg, 0.246
mmol) under nitrogen was added 1,4-dioxane (30 mL) followed by 3 M
aqueous sodium carbonate (4.93 mL, 14.8 mmol). The resulting
mixture was degassed with nitrogen for 10 minutes then heated at
reflux for 16 hours. On cooling EtOAc (150 mL) was added and the
resulting solution was washed with water (3.times.50 mL), brine (50
mL) then dried (Na.sub.2SO.sub.4). The volatiles were evaporated
under reduced pressure to give a brown solid that was purified
using silica gel column chromatography (Biotage Isolera, 40 g
SiO.sub.2 cartridge, 0-30% EtOAc in petroleum benzine 40-60.degree.
C.) to give the title compound I8 as a yellow solid (1.43 g, 95%);
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 9.37 (dd, J=2.6, 0.5 Hz,
1H), 8.43 (dd, J=8.8, 2.7 Hz, 1H), 7.53 (d, J=8.8 Hz, 1H),
6.95-6.83 (m, 1H), 4.20 (d, J=3.0 Hz, 2H), 3.67 (t, J=5.6 Hz, 2H),
2.70-2.63 (m, 2H), 1.49 (s, 9H). LCMS-A: rt 6.140 min; m/z 304
[M-H].sup.-.
(b) tert-Butyl 4-(5-aminopyridin-2-yl)piperidine-1-carboxylate
(I9)
[0402] A slurry of 10% Pd/C (500 mg) in DMF (5 mL) was added to a
solution of tert-butyl
5-nitro-5',6'-dihydro-[2,4'-bipyridine]-1'(2'H)-carboxylate (I8)
(1.40, 4.59 mmol) in DMF (45 mL) and the resulting mixture was
stirred under a hydrogen atmosphere for 16 hours at room
temperature. EtOAc (100 mL) was added and the resulting suspension
was filtered through a Celite pad, washing with EtOAc (150 mL). The
volatiles were removed under reduced pressure and the residue was
purified by silica gel column chromatography (Biotage Isolera, 40 g
Si cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.
and then 0-20% MeOH in EtOAc) to give the title compound I9 as a
yellow oil (1.18 g, 93%); .sup.1H NMR (400 MHz, CDCl.sub.3) .delta.
8.03 (dd, J=2.4, 1.0 Hz, 1H), 7.00-6.87 (m, 2H), 4.22 (brs, 2H),
3.59 (brs, 2H), 2.85-2.67 (m, 3H), 1.86 (m, 2H), 1.72-1.59 (m, 2H),
1.46 (s, 9H). LCMS-A: rt 4.416 min; m/z 278 [M+H].sup.+.
(c) tert-Butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-2-yl)pipe-
ridine-1-carboxylate (K5)
[0403] A 1.0 M ZnCl.sub.2 solution in Et.sub.2O (1.14 mL, 1.14
mmol) was added cautiously to a stirred solution of
2,4-dichloro-5-(trifluoromethyl)pyrimidine (217 mg, 0.999 mmol) in
a 1:1 t-BuOH:DCE mixture (50 mL) at room temperature. After
completion of the addition stirring was continued for 20 minutes
then tert-butyl 4-(5-aminopyridin-2-yl)piperidine-1-carboxylate
(I9) (263 mg, 0.952 mmol) followed by Et.sub.3N (0.159 mL, 1.14
mmol) were added. The resulting mixture was stirred at room
temperature for 48 hours then the volatiles were evaporated under
reduced pressure and the resulting residue was purified using
silica gel column chromatography (Biotage Isolera, 24 g SiO.sub.2
cartridge, 0-55% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound K5 as a white solid (155 mg, 36%); .sup.1H
NMR (400 MHz, d.sub.6-DMSO) .delta. 10.75 (s, 1H), 8.82 (s, 1H),
8.73 (d, J=2.4 Hz, 1H), 8.00 (dd, J=8.5, 2.4 Hz, 1H), 7.30 (d,
J=8.5 Hz, 1H), 4.11-4.01 (m, 2H), 2.93-2.75 (m, 3H), 1.81 (d,
J=11.1 Hz, 2H), 1.63-1.50 (m, 2H), 1.41 (s, 9H). LCMS-A: rt 5.604
min; m/z 458, 460 [M+H].sup.+.
Key Intermediate 6
1-(2-Ethynylphenyl)cyclopropanecarboxamide (K6)
##STR00115##
[0404] (a) 1-(2-Iodophenyl)cyclopropanecarboxylic acid (I10)
[0405] A solution of 1-phenyl-1-cyclopropanecarboxylic acid (2.00
g, 12.3 mmol), Pd(OAc).sub.2 (0.138 g, 0.617 mmol), iodine (2.34 g,
9.24 mmol) and (diacetoxyiodo)benzene (2.97 g, 9.24 mmol) in DMF
(10 mL) was stirred at 60.degree. C. for 18 hours covered aluminium
foil. Additional iodine (2.34 g, 9.24 mmol) and
(diacetoxyiodo)benzene (2.97 g, 9.24 mmol) were added and stirring
was continued at 60.degree. C. for a further 8 hours. A final
addition of iodine (2.34 g, 9.24 mmol) and (diacetoxyiodo)benzene
(2.97 g, 9.24 mmol) was performed and stirring was continued at
60.degree. C. for a further 16 hours. The resulting mixture was
partitioned between EtOAc and water and the aqueous phase was
extracted several times with EtOAc. The combined organic extracts
were washed with a 10% aqueous solution of sodium metabisulfate
(3.times.30 mL), 10% aqueous citric acid (2.times.30 mL), water,
brine, dried (MgSO.sub.4), filtered and evaporated. The residue was
purified using silica gel column chromatography (CombiFlash Rf, 80
g SiO.sub.2 Cartridge, 30-40% EtOAc in cyclohexane) to give the
title compound I10 as a cream solid (3.11 g, 87%); .sup.1H NMR (300
MHz, CDCl.sub.3) .delta. 7.88 (d, J=8.0 Hz, 1H), 7.28-7.35 (m 2H),
6.97-7.03 (m, 1H), 2.13 (brs, 2H), 1.30 (brs, 2H).
(b) 1-(2-Iodophenyl)cyclopropanecarboxamide (I11)
[0406] HOBt (1.89 g, 14.0 mmol) and EDCl.HCl (2.69 g, 14.0 mmol)
were added to a solution of 1-(2-iodophenyl)cyclopropanecarboxylic
acid (I10) (3.11 g, 10.8 mmol) in DMF (6 mL), THF (29 mL) and
Et.sub.3N (9.4 mL, 54 mmol). After 15 minutes ammonium carbonate
(7.95 g, 86.4 mmol) was added and the resulting mixture was stirred
at room temperature for 20 hours. The volatiles were removed in
vacuo and water (150 mL) was added to the residue resulting in the
formation of a suspension. The suspension was sonicated for several
minutes, filtered and the filter cake dried to give the title
compound I11 as an off white solid (2.55 g, 82%); .sup.1H NMR (300
MHz, d.sub.6-DMSO) .delta. 7.87 (d, J=7.6 Hz, 1H), 7.36-7.38 (m,
2H), 7.02-7.08 (m, 1H), 6.96 (brs, 1H), 6.05 (brs, 1H), 1.52 (brs,
2H), 0.96 (brs, 2H). LCMS-B: rt 5.800 min; m/z 288 [M+H].sup.+.
(c) 1-(2-((Trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxamide
(I12)
[0407] A solution of 1-(2-iodophenyl)cyclopropanecarboxamide (I11)
(1.53 g, 85.3 mmol), ethynyltrimethylsilane (1.05 mL, 7.46 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (0.037 g, 0.053 mmol),
t-Bu.sub.3PH.BF.sub.4 (0.031 g, 0.11 mmol) and copper(I) iodide
(0.020 g, 0.11 mmol) in DMF (10 mL) was stirred under a nitrogen
atmosphere at 50.degree. C. for 26 hours. The volatiles were
evaporated in vacuo and the resulting black residue was purified
using silica gel column chromatography (CombiFlash Rf, 24 g
SiO.sub.2 Cartridge, 5-20% EtOAc in cyclohexane) to give the title
compound I12 as a dark yellow amorphous solid (1.09 g, 79%);
.sup.1H NMR (300 MHz, CDCl.sub.3) .delta. 7.52 (d, J=7.1 Hz, 1H),
7.26-7.50 (m, 3H), 5.26 (brs, 2H), 1.72 (dq, J=4.1 Hz, 2H) 1.16
(dq, J=3.9 Hz, 2H), 0.26 (s, 9H). LCMS-B: rt 7.446 min; m/z 258
[M+H].sup.+.
(d) 1-(2-Ethynylphenyl)cyclopropanecarboxamide (K6)
[0408] A cooled (5.degree. C. water/ice bath) solution of
1-(2-((trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxamide (I12)
(1.09 g, 4.23 mmol) in DCM (20 mL) containing acetic acid (0.315
mL, 5.50 mmol) was slowly treated with a 1 M solution of TBAF in
THF (5.08 mL, 5.08 mmol). The resulting mixture was stirred at room
temperature for 2 hours then water (30 mL) was added. The aqueous
layer was extracted with DCM (2.times.15 mL), then the combined
organic extracts were washed with water, brine, dried (MgSO.sub.4),
filtered and evaporated in vacuo to give the title compound as a
tan solid (0.780 g, 99%); .sup.1H NMR (300 MHz, CDCl.sub.3) .delta.
7.57 (d, J=6.7 Hz, 1H), 7.31-7.45 (m, 3H), 5.46 (brs, 1H), 5.27
(brs, 1H), 3.38 (s, 1H), 1.75 (dq, J=4.0 Hz, 2H) 1.18 (dq, J=3.7
Hz, 2H). LCMS-B: rt 5.489 min; m/z 186 [M+H].sup.+.
Synthesis of Key Intermediate 7
2-(2-Ethynylphenyl)propanamide (K7)
##STR00116##
[0409] Method 1
(a) 2-(2-Iodophenyl)propanamide (I13)
[0410] A solution of 2-(2-iodophenyl)propanoic acid (9.73 g, 35.2
mmol) in THF (24 mL), DMF (4 mL) and DIPEA (30.7 mL, 0.176 mol) was
stirred with HOBt (7.14 g, 52.9 mmol) and EDCl.HCl (10.1 g, 52.9
mmol) for 10 minutes at room temperature under N.sub.2. Ammonium
carbonate (16.9 g, 0.176 mol) was then added and the reaction
mixture was stirred overnight at room temperature. The volatiles
were removed under reduced pressure before sat. aq. NaHCO.sub.3
(100 mL) was added to the residue. The aqueous phase was extracted
with EtOAc (3.times.100 mL) and the combined organics were washed
with brine, dried over MgSO.sub.4 and the volatiles were removed in
vacuo to give the impure title compound I13 as a pale yellow solid
(.about.95% purity, 10.05 g, >95%). LCMS-D: rt 3.12 min; m/z 276
[M+H].sup.+.
Alternative synthesis of 2-(2-Iodophenyl)propanamide (I13)
[0411] Thionyl chloride (9.00 mL, 123 mmol) was added to
2-iodo-phenacetic acid (16.0 g, 58.0 mmol) and the mixture was
stirred at 40.degree. C. for 2 hours. The volatiles were removed in
vacuo before EtOAc (40 mL) and 28% ammonium hydroxide aqueous
solution (17.9 mL. 118 mmol) were added. The mixture was stirred at
room temperature overnight before EtOAc (200 mL) and water (100 mL)
were added. The layers were separated and the aqueous phase was
extracted with EtOAc (2.times.100 mL). The combined organics were
washed with brine (100 mL), dried (Na.sub.2SO.sub.4) and
concentrated in vacuo to give the title compound I13 as a solid
(15.7 g, 98%).
(b) 2-(2-((Triethylsilyl)ethynyl)phenyl)propanamide (I14)
[0412] A mixture of 2-(2-iodophenyl)propanamide 113 (12.4 g, 45.2
mmol), ethynyltriethylsilane (9.72 mL, 54.2 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (0.634 g, 0.904 mmol),
tri-tert-butylphosphonium tetrafluoroborate (0.262 g, 0.904 mmol)
and CuI (0.172 g, 0.904 mmol) in DMF (46 mL) was stirred under a
nitrogen atmosphere at 70.degree. C. for 4 hours. The volatiles
were removed in vacuo and the crude residue was purified by silica
gel column chromatography (Combiflash Rf, 0-30% EtOAc in
cyclohexane) to give the title compound I14 as an orange oil (8.9
g, 68%). LCMS-B: rt 3.845 min; m/z 288 [M+H].sup.+.
(c) 2-(2-Ethynylphenyl)propanamide (K7)
[0413] A cooled (0.degree. C. water/ice bath) solution of
2-(2-((triethylsilyl)ethynyl)phenyl)propanamide 114 (7.17 g, 24.9
mmol) in THF (40 mL) was slowly treated with a 1.0 M solution of
tetrabutylammonium fluoride in THF (26.2 mL, 26.2 mmol) under a
nitrogen atmosphere. The resulting mixture was stirred at 0.degree.
C. for 5 minutes before a sat. aq. NaHCO.sub.3 solution was added.
The mixture was extracted with EtOAc (3 times), the combined
organic extracts were washed with brine, dried (Na.sub.2SO.sub.4)
and evaporated in vacuo to give the crude product. The crude
product was purified by silica gel column chromatography
(Combiflash Rf, 30-80% EtOAc in cyclohexane) to give the title
compound K7 as a cream solid (3.69 g, 85%). LCMS-B: rt 3.031 min;
m/z 174 [M+H].sup.+.
Method 2
##STR00117##
[0414] (a) 2-(2-((Trimethylsilyl)ethynyl)phenyl)propanamide
(I15)
[0415] A mixture of 2-(2-iodophenyl)propanamide I13 (16.5 g, 0.060
mol), CuI (0.229 g, 1.20 mmol), tri-tert-butylphosphonium
tetrafluoroborate (0.348 g, 1.20 mmol), PdCl.sub.2(PPh.sub.3).sub.2
(0.421 g, 0.600 mmol) and trimethylsilylacetylene (10.3 mL, 0.072
mol) in DMF (60 mL) was bubbled with N.sub.2 for 10 minutes.
Et.sub.3N (50 mL) was then added and the mixture was stirred under
nitrogen at 60.degree. C. for 5 hours. The reaction mixture was
cooled and the volatiles were removed in vacuo. The dark brown
residue was adsorbed onto silica and purified by column
chromatography (Biotage Isolera, 120 g SiO.sub.2 cartridge, 0-30%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound I15 as an orange oil (11.5 g, 78%). LCMS-D: rt 3.49 min;
m/z 246 [M+H].sup.+.
(b) 2-(2-Ethynylphenyl)propanamide (K7)
[0416] A 1.0 M solution of TBAF in THF (45.6 mL, 45.6 mmol) was
added to a solution of
2-(2-((trimethylsilyl)ethynyl)phenyl)propanamide I15 (9.33 g, 38.0
mmol) in THF (200 mL) and the mixture was stirred for 10 minutes.
The reaction mixture was diluted with sat. aq. NaHCO.sub.3 (200
mL), and the aqueous layer was extracted with DCM (3.times.150 mL).
The combined organics were adsorbed onto silica and purified by
silica gel column chromatography (Biotage Isolera, 120 g SiO.sub.2
cartridge, 0-80% EtOAc in petroleum benzine 40-60.degree. C.) to
yield the title compound K7 as a tan solid (5.11 g, 76%). LCMS-D:
rt 3.03 min; m/z 174 [M+H].sup.+.
Synthesis of Key Intermediate 8
tert-Butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyr-
azol-1-yl)piperidine-1-carboxylate (K8)
##STR00118##
[0418] A 1.0 M solution of ZnCl.sub.2 in Et.sub.2O (22.5 mL, 22.5
mmol) was added to a solution of
2,4-dichloro-5-(trifluoromethyl)pyrimidine (4.48 g, 20.7 mmol) in
t-BuOH (50 mL) and DCE (50 mL) and the mixture was stirred for 10
minutes. The mixture was diluted with t-BuOH (50 mL) and DCE (50
mL) before tert-butyl
4-(4-amino-1H-pyrazol-1-yl)piperidine-1-carboxylate (5.00 g, 18.8
mmol) and Et.sub.3N (3.14 mL, 22.5 mmol) were added. Stirring was
continued overnight and the volatiles were subsequently removed in
vacuo. The resultant residue was suspended in acetone (50 mL) then
water (500 mL) and sonicated for 15 minutes before the solid was
removed by vacuum filtration and the filter cake washed with water
(100 mL). The solid was suspended in acetone (25 mL), filtered and
the filter cake was washed with petroleum benzine 40-60.degree. C.
(200 mL) to give the title compound K8 as a light pink solid (6.57
g, 78%). LCMS-D: rt 3.618 min; m/z 445.1 [M-H].sup.-.
Example 1
Synthesis of
2-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1)
##STR00119##
[0419] (a) tert-Butyl
4-(4-((4-((2-(1-amino-1-oxobutan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl)-
pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A1)
[0420] A mixture of tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)piperidine-
-1-carboxylate (K3) (193 mg, 0.422 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (15 mg, 0.021 mmol),
t-Bu.sub.3PH.BF.sub.4 (6 mg, 0.021 mmol), CuI (4 mg, 0.021 mmol)
and 2-(2-ethynylphenyl)butanamide (K1) (87 mg, 0.47 mmol) in DMF
(3.0 mL) was degassed with nitrogen for 10 minutes. Et.sub.3N (1.0
mL) was added and the resulting mixture was heated under microwave
irradiation at 120.degree. C. for 20 minutes. The volatiles were
removed under reduced pressure and the residue was adsorbed onto
silica gel then purified by column chromatography (Biotage Isolera,
SiO.sub.2 cartridge, 0-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A1 as a yellow solid
(175 mg, 68%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 10.39
(s, 1H), 8.88-8.76 (m, 1H), 7.67 (d, J=8.5 Hz, 2H), 7.63-7.47 (m,
3H), 7.37 (td, J=7.4, 1.6 Hz, 1H), 7.27-7.17 (m, 3H), 7.01 (s, 1H),
4.07 (s, 2H), 3.89 (dd, J=8.6, 6.4 Hz, 1H), 2.80 (br, 2H), 2.64 (J
m, 1H), 1.98-1.90 (m, 1H), 1.74 (d, J=12.7 Hz, 2H), 1.70-1.60 (m,
1H), 1.55-1.44 (m, 2H), 1.42 (s, 9H), 0.87 (t, J=7.3 Hz, 3H).
LCMS-A: rt 6.726 min; m/z 606 [M-H].sup.-. CL (b) tert-Butyl
4-(4-((4-(2-(1-amino-1-oxobutan-2-yl)phenethyl)-5-(trifluoromethyl)pyrimi-
din-2-yl)amino)phenyl)piperidine-1-carboxylate (A2)
[0421] A suspension of 10% Pd/C (100 mg), tert-butyl
4-(4-((4-((2-(1-amino-1-oxobutan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl)-
pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A1) (167 mg,
0.275 mmol) and Et.sub.3N (1.0 mL) in DMF (10 mL) was stirred under
an atmosphere of hydrogen for 16 hours. The resulting mixture was
diluted with EtOAc (60 mL), filtered through Celite then the
solvent removed under reduced pressure to give the title compound
A2 as a yellow oil (412 mg, 98%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.15 (s, 1H), 8.67 (s, 1H), 7.67 (d, J=8.6
Hz, 2H), 7.49-7.42 (m, 1H), 7.31 (s, 1H), 7.22-7.14 (m, 5H), 6.87
(s, 1H), 4.06 (d, J=10.5 Hz, 2H), 3.61 (dd, J=8.8, 6.1 Hz, 1H),
3.29-2.94 (m, 4H), 2.88-2.74 (m, obscured by solvent), 2.70-2.59
(m, 1H), 2.04-1.92 (m, 1H), 1.74 (d, J=12.6 Hz, 2H), 1.61-1.43 (m,
3H), 1.41 (s, 9H), 0.83 (t, J=7.3 Hz, 3H). LCMS-A: rt 6.722 min;
m/z 612 [M+H].sup.+.
(c)
2-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimi-
din-4-yl)ethyl)phenyl)butanamide (1)
[0422] A solution of TFA (0.84 mL, 11 mmol) and tert-butyl
4-(4-((4-(2-(1-amino-1-oxobutan-2-yl)phenethyl)-5-(trifluoromethyl)pyrimi-
din-2-yl)amino)phenyl)piperidine-1-carboxylate (A2) (168 mg, 0.275
mmol) in DCM (20 mL) was stirred for 24 hours at room temperature
under a nitrogen atmosphere. The volatiles were removed in vacuo
and the residue was taken up in MeOH and loaded onto an SCX
cartridge (10 g). The column was eluted with 5 column volumes of
MeOH and then 5 column volumes of 5% v/v aqueous ammonia in MeOH to
elute the amine product. The ammoniacal filtrate was evaporated
under reduced pressure and the residue was taken up in DCM
(.about.2 mL). Cyclohexane (.about.10 mL) was added and the
resulting suspension sonicated for 10 minutes. The precipitate was
isolated by filtration to give the title compound 1 as an off-white
solid (100 mg, 71%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta.
10.14 (s, 1H), 8.67 (s, 1H), 7.66 (d, J=8.6 Hz, 2H), 7.51-7.43 (m,
1H), 7.31 (s, 1H), 7.22-7.10 (m, 5H), 6.87 (s, 1H), 3.61 (dd,
J=8.8, 6.1 Hz, 1H), 3.22-2.89 (m, 6H), 2.62-2.53 (m, 3H), 2.07-1.91
(m, 1H), 1.67 (d, J=12.0 Hz, 2H), 1.62-1.42 (m, 3H), 0.87-0.80 (m,
3H). LCMS-A: rt 4.973 min; m/z 512 [M+H].sup.+.
Example 1-1A and 1-2A
Separation of
2-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1)
##STR00120##
[0424] Racemic
2-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1) was separated using the chiral
separation Method C. The absolute configuration of each enantiomer
has not been determined empirically.
[0425] Enantiomer A of
2-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1-1A), Chiral Characterisation
Method C: rt 5.58 min, enantiomeric purity >99%.
[0426] Enantiomer B of
2-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)butanamide (1-2A), Chiral Characterisation
Method C: rt 3.82 min, enantiomeric purity 97.4%.
Example 2
Synthesis of
1-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (2)
##STR00121## ##STR00122##
[0427] (a) tert-Butyl
4-(4-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate
(A3)
[0428] A solution of tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)piperidine-
-1-carboxylate (K3) (262 mg, 0.574 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (20 mg, 0.029 mmol,)
t-Bu.sub.3PH.BF.sub.4 (8 mg, 0.03 mmol), CuI (5 mg, 0.03 mmol) and
methyl 1-(2-ethynylphenyl)cyclopropanecarboxylate (K2) (138 mg,
0.689 mmol) in DMF (5.0 mL) was degassed with nitrogen for 10
minutes. Et.sub.3N (1.0 mL) was added and the resulting mixture
heated under microwave irradiation for 20 minutes at 120.degree. C.
The resulting mixture was adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, SiO.sub.2 cartridge, 0-40%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound A3 as a yellow solid (308 mg, 86%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.37 (s, 1H), 8.79 (s, 1H), 7.67 (d, J=8.2
Hz, 2H), 7.60 (dd, J=7.6, 1.0 Hz, 1H), 7.54-7.38 (m, 3H), 7.21 (d,
J=8.6 Hz, 2H), 4.07 (d, J=11.6 Hz, 2H), 3.52 (s, 3H), 2.80 (s, 2H),
2.65 (J m, 1H), 1.74 (d, J=12.4 Hz, 2H), 1.64 (q, J=4.0 Hz, 2H),
1.49 (td, J=12.6, 4.1 Hz, 2H), 1.42 (s, 9H), 1.29 (q, J=4.2 Hz,
2H). LCMS-A: rt 7.083 min; m/z 619 [M-H].sup.-.
(b) tert-Butyl
4-(4-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A4)
[0429] A suspension of 10% Pd/C (200 mg), tert-butyl
4-(4-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A3)
(0.302 g, 0.487 mmol) and Et.sub.3N (1.0 mL) in DMF (10 mL) was
stirred under an atmosphere of hydrogen for 16 hours. The resulting
mixture was diluted with EtOAc (60 mL) and filtered through Celite.
The filtrate was evaporated under reduced pressure to give the
title compound A4 as a yellow oil (494 mg, 99%); .sup.1H NMR (400
MHz, d.sub.6-DMSO) .delta. 10.15 (s, 1H), 8.68 (s, 1H), 7.67 (d,
J=8.6 Hz, 2H), 7.33-7.26 (m, 3H), 7.22-7.17 (m, 3H), 4.07 (d,
J=11.6 Hz, 2H), 3.49 (s, 3H), 3.15-3.07 (m, 2H), 3.07-3.00 (m, 2H),
2.86-2.75 (m, peaks obscured by solvent), 2.70-2.58 (m, 1H), 1.74
(d, J=11.3 Hz, 2H), 1.58 (d, J=3.4 Hz, 2H), 1.48 (dt, J=12.6, 6.2
Hz, 2H), 1.42 (s, 9H), 1.21 (s, 2H). LCMS-A: rt 7.175 min; m/z 625
[M+H].sup.+.
(c)
1-(2-(2-(2-((4-(1-(tert-Butoxycarbonyl)piperidin-4-yl)phenyl)amino)-5--
(trifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxylic
acid (A5)
[0430] A solution of tert-butyl
4-(4-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A4) (0.304
g, 0.487 mmol) and LiOH.H.sub.2O (204 mg, 4.87 mmol) in THF (7 mL),
MeOH (7 mL) and H.sub.2O (1.5 mL) was stirred at room temperature
overnight. The resulting mixture was then heated to 40.degree. C.
and stirred for 3.5 days. The volatiles were removed under reduced
pressure and the residue was taken up in EtOAc (50 mL). Aqueous HCl
(2 M, 50 mL) was added cautiously and the layers were separated.
The aqueous phase was extracted with EtOAc (2.times.50 mL), and
then the combined organics were washed with brine and dried over
MgSO.sub.4. The solvent was removed in vacuo to give the title
compound A5 as a yellow oil (680 mg, 98%); LCMS-A: rt 6.793 min;
m/z 609 [M-H].sup.-.
(d) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)phenyl)piperidine-1-carboxylate (A6)
[0431] HOBt (99 mg, 0.73 mmol), EDCl.HCl (140 mg, 0.73 mmol) and
DIPEA (0.42 mL, 2.43 mmol) were added to a stirred solution of
1-(2-(2-(2-((4-(1-(tert-butoxycarbonyl)piperidin-4-yl)phenyl)amino)-5-(tr-
ifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxylic
acid (A5) (297 mg, 0.486 mmol) in dry THF (6 mL) and dry DMF (1 mL)
under an atmosphere of nitrogen. After 10 minutes ammonium
carbonate (234 mg, 2.43 mmol) was added in one portion and the
resulting solution was stirred at 40.degree. C. for 72 hours. The
volatiles were removed in vacuo and EtOAc (50 mL) and saturated
NaHCO.sub.3 (50 mL) were added to the residue. After separating the
organic layer, the aqueous phase was extracted with EtOAc
(2.times.50 mL). The organic extracts were combined, washed with
brine and dried over MgSO.sub.4. The volatiles were removed under
reduced pressure and the residue was purified by silica gel column
chromatography (Biotage Isolera, SiO.sub.2 cartridge, 0-60% EtOAc
in petroleum benzine 40-60.degree. C.) to give the title compound
A6 as a yellow solid (131 mg, 44%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.15 (s, 1H), 8.67 (s, 1H), 7.67 (d, J=8.5
Hz, 2H), 7.32 (dd, J=13.9, 5.0 Hz, 3H), 7.26-7.14 (m, 3H), 6.98 (s,
1H), 5.99 (s, 1H), 4.05 (dd, J=15.7, 8.8 Hz, 2H), 3.13 (dd, J=25.3,
9.9 Hz, 4H), 2.93-2.70 (m, 2H), 2.63 (t, J=12.2 Hz, 1H), 1.74 (d,
J=12.5 Hz, 2H), 1.41 (s, 13H), 0.97 (s, 2H). LCMS-A: rt 6.720 min;
m/z 610 [M+H].sup.+.
(e)
1-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimi-
din-4-yl)ethyl)phenyl)cyclopropanecarboxamide (2)
[0432] A solution of TFA (0.65 mL, 8.5 mmol) and tert-butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)phenyl)piperidine-1-carboxylate (A6) (0.13 mg, 0.21
mmol) in DCM (20 mL) was stirred for 24 hours at room temperature
under nitrogen. The volatiles were removed in vacuo and the residue
was taken up in MeOH and loaded onto an SCX cartridge (10 g). The
column was eluted with 5 column volumes of MeOH and then 5 column
volumes of 5% v/v aqueous ammonia in MeOH to elute the amine
product. The ammoniacal filtrate was evaporated under reduced
pressure and the resulting solid was dried under high vacuum to
give the title compound 2 as a white solid (100 mg, 93%); .sup.1H
NMR (400 MHz, d.sub.6-DMSO) .delta. 10.14 (s, 1H), 8.67 (s, 1H),
7.66 (d, J=8.6 Hz, 2H), 7.37-7.27 (m, 3H), 7.25-7.14 (m, 3H), 6.99
(s, 1H), 6.00 (s, 1H), 3.22-3.13 (m, 2H), 3.13-3.06 (m, 2H), 3.01
(d, J=11.9 Hz, 2H), 2.62-2.52 (m, peaks obscured by solvent), 1.67
(d, J=10.7 Hz, 2H), 1.57-1.40 (m, 4H), 0.97 (s, 2H). LCMS-A: rt
4.963 min; m/z 510 [M+H].sup.+.
Example 3
Synthesis of
1-(2-(2-(2-((4-(1-methylpiperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)p-
yrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (3)
##STR00123##
[0434] Formaldehyde (37 wt % in H.sub.2O; 8.8 .mu.L, 0.12 mmol) was
added to a solution of
1-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (2) (20 mg, 39 .mu.mol)
in MeOH (5.0 mL) under an atmosphere of nitrogen. The resulting
mixture was stirred for 10 minutes at room temperature then sodium
triacetoxyborohydride (33 mg, 0.16 mmol) was added in one portion
and stirring continued for 2.5 hours. The volatiles were removed in
vacuo and the residue was diluted with EtOAc (20 mL) and saturated
aqueous NaHCO.sub.3 (20 mL). The layers were separated and the
aqueous layer was extracted with EtOAc (2.times.20 mL). The
combined organic layers were washed with brine (20 mL), dried over
MgSO.sub.4 and the volatiles were removed under reduced pressure.
The residue was dissolved in DCM (1 mL) to which cyclohexane (5 mL)
was added to form a cloudy suspension. The solvent was removed in
vacuo and the resulting solid was dried under high vacuum to give
the title compound 3 as a white powder (18 mg, 88%); .sup.1H NMR
(400 MHz, d.sub.6-DMSO) .delta. 10.15 (s, 1H), 8.67 (s, 1H), 7.66
(d, J=8.5 Hz, 2H), 7.40-7.27 (m, 3H), 7.22 (dd, J=14.4, 8.0 Hz,
3H), 6.99 (s, 1H), 6.01 (s, 1H), 3.21-3.05 (m, 4H), 2.88 (d, J=10.9
Hz, 2H), 2.50-2.30 (m, 1H), 2.21 (s, 3H), 1.99 (s, 2H), 1.79-1.58
(m, 4H), 1.44 (s, 2H), 0.97 (s, 2H). LCMS-A: 5.028 min; m/z 524
[M+H].sup.+.
Example 4
Synthesis of
1-(2-(2-(2-((4-(azetidin-3-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (4)
##STR00124##
[0435] (a) tert-Butyl
3-(4-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)phenyl)azetidine-1-carboxylate (A7)
[0436] A solution of tert-butyl
3-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)azetidine--
1-carboxylate (K4) (260 mg, 0.606 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (21 mg, 0.030 mmol), PPh.sub.3 (16 mg,
0.061 mmol), CuI (12 mg, 0.061 mmol) and methyl
1-(2-ethynylphenyl)cyclopropanecarboxylate (K2) (146 mg, 0.728
mmol) in DMF (3.0 mL) was degassed with nitrogen for 10 minutes.
Et.sub.3N (1.0 mL) was added and the resulting mixture heated under
microwave irradiation at 120.degree. C. for 20 minutes. The
resulting mixture was adsorbed onto silica and purified by silica
gel column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-25% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A7 as an orange oil (359 mg, >95%);
.sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 10.45 (s, 1H), 8.82 (s,
1H), 7.75 (d, J=8.4 Hz, 2H), 7.61 (dd, J=7.5, 1.0 Hz, 1H),
7.55-7.38 (m, 3H), 7.32 (d, J=8.6 Hz, 2H), 4.32-4.17 (m, 2H),
3.89-3.74 (m, 3H), 3.52 (s, 3H), 1.64 (q, J=4.0 Hz, 2H), 1.40 (s,
9H), 1.29 (q, J=4.2 Hz, 2H). LCMS-A: rt 6.550 min; m/z 593
[M+H].sup.+.
(b) tert-Butyl
3-(4-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)phenyl)azetidine-1-carboxylate (A8)
[0437] A suspension of 10% Pd/C (200 mg), tert-butyl
3-(4-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)phenyl)azetidine-1-carboxylate (A7)
(353 mg, 0.596 mmol) and Et.sub.3N (1.5 mL) in DMF (10 mL) was
stirred with under a hydrogen atmosphere for 16 hours. The
resulting mixture was diluted with EtOAc (60 mL), filtered through
Celite and the volatiles were removed under reduced pressure. The
residue was adsorbed onto silica gel and purified by column
chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-40%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound A8 as yellow oil (353 mg, 99%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.22 (s, 1H), 8.69 (s, 1H), 7.74 (d, J=8.6
Hz, 2H), 7.32-7.26 (m, 5H), 7.22-7.16 (m, 1H), 4.24 (t, J=7.4 Hz,
2H), 3.87-3.71 (m, 3H), 3.49 (s, 3H), 3.17-2.98 (m, 4H), 1.58 (d,
J=3.3 Hz, 2H), 1.40 (s, 9H), 1.19 (s, 2H). LCMS-A: rt 6.650 min;
m/z 595 [M-H].sup.-.
(c) tert-Butyl
3-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)phenyl)azetidine-1-carboxylate (A9)
[0438] A solution of tert-butyl
3-(4-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)phenyl)azetidine-1-carboxylate (A8) (347 mg,
0.582 mmol) and LiOH.H.sub.2O (293 mg, 6.98 mmol) in THF (7.0 mL),
MeOH (7.0 mL) and H.sub.2O (2.0 mL) was stirred at room temperature
for 3 days. LiOH.H.sub.2O (147 mg, 3.49 mmol) was added and the
resulting mixture was heated at 35.degree. C. for 16 hours. EtOAc
(50 mL) and aqueous HCl (2 M, 50 mL) were added cautiously, then
the layers were separated and the aqueous phase was extracted with
EtOAc (2.times.50 mL). The organic layers were combined, washed
with brine, dried (MgSO.sub.4) then evaporated to dryness. The
residue was dissolved in THF (12 mL) and DMF (2.0 mL), to which
HOBt (157 mg, 1.16 mmol), EDCl.HCl (223 mg, 1.16 mmol) and DIPEA
(0.507 mL, 2.91 mmol) were added under an atmosphere of nitrogen.
After 10 minutes ammonium carbonate (280 mg, 2.91 mmol) was added
in one portion and the resulting mixture was stirred at room
temperature for 16 hours, then 40.degree. C. for a further 24
hours. The volatiles were removed in vacuo and EtOAc (20 mL) and
saturated NaHCO.sub.3 (20 mL) were added. The layers were separated
then the aqueous phase extracted with EtOAc (2.times.20 mL). The
organic extracts were combined, washed with brine, dried
(MgSO.sub.4) then evaporated to dryness. The residue was purified
by silica gel column chromatography (Biotage Isolera, 24 g
SiO.sub.2 cartridge, 10-50% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A9 as a white solid
(179 mg, 53%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 10.22
(s, 1H), 8.69 (s, 1H), 7.73 (d, J=8.6 Hz, 2H), 7.37-7.27 (m, 5H),
7.26-7.19 (m, 1H), 6.98 (s, 1H), 5.99 (s, 1H), 4.23 (t, J=7.7 Hz,
2H), 3.88-3.70 (m, 3H), 3.21-3.14 (m, 2H), 3.14-3.07 (m, 2H), 1.44
(d, J=3.5 Hz, 2H), 1.40 (s, 9H), 0.97 (s, 2H). LCMS-A: rt 6.168
min; m/z 582 [M+H].sup.+.
(d)
1-(2-(2-(2-((4-(azetidin-3-yl)phenyl)amino)-5-(trifluoromethyl)pyrimid-
in-4-yl)ethyl)phenyl)cyclopropanecarboxamide (4)
[0439] A solution of TFA (0.916 mL, 12.0 mmol) and tert-butyl
3-(4-((4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyri-
midin-2-yl)amino)phenyl)azetidine-1-carboxylate (A9) (174 mg, 0.299
mmol) in DCM (10 mL) was stirred for 16 hours at room temperature
under nitrogen. The volatiles were removed in vacuo and the residue
was taken up in MeOH and loaded onto an SCX cartridge (10 g). The
column was eluted with 5 column volumes of MeOH and then 5 column
volumes of 5% v/v aqueous ammonia in MeOH to elute the amine
product. The ammoniacal filtrate was evaporated under reduced
pressure and dried under high vacuum to give the title compound 4
as a yellow solid (129 mg, 90%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.19 (s, 1H), 8.68 (s, 1H), 7.77-7.67 (m,
2H), 7.38-7.26 (m, 5H), 7.26-7.18 (m, 1H), 6.99 (s, 1H), 6.00 (s,
2H), 3.86 (s, 3H), 3.71 (s, 2H), 3.22-3.13 (m, 2H), 3.13-3.05 (m,
2H), 1.44 (s, 2H), 0.96 (s, 2H). LCMS-A: rt 4.591 min; m/z 482
[M+H].sup.+.
Example 5
Synthesis of
1-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (5)
##STR00125##
[0440] (a) tert-Butyl
4-(5-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate
(A10)
[0441] A stirred suspension of tert-butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-2-yl)pipe-
ridine-1-carboxylate (K5) (152 mg, 0.332 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (12 mg, 0.017 mmol), PPh.sub.3 (9 mg,
0.03 mmol), CuI (6 mg, 0.03 mmol) and methyl
1-(2-ethynylphenyl)cyclopropanecarboxylate (K2) (80 mg, 0.40 mmol)
in DMF (4.0 mL) was degassed with nitrogen for 10 minutes.
Et.sub.3N (1.0 mL) was added and the resulting mixture heated under
microwave irradiation at 120.degree. C. for 20 minutes. The
volatiles were removed in vacuo and the residue was adsorbed onto
silica gel and purified by column chromatography (Biotage Isolera,
24 g SiO.sub.2 cartridge, 0-60% EtOAc in petroleum benzine
40-60.degree. C. then 10-45% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A10 as yellow oil (141
mg, 68%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 10.53 (s,
1H), 8.84 (s, 1H), 8.77 (d, J=2.5 Hz, 1H), 8.12 (d, J=6.5 Hz, 1H),
7.61 (dd, J=7.6, 1.0 Hz, 1H), 7.54-7.38 (m, 3H), 7.28 (d, J=8.5 Hz,
1H), 4.05 (br, 2H), 3.51 (s, 3H), 2.94-2.72 (m, 3H), 1.81 (d,
J=10.5 Hz, 2H), 1.63 (q, J=4.1 Hz, 2H), 1.61-1.49 (m, 2H), 1.41 (s,
9H), 1.29 (q, J=4.2 Hz, 2H). LCMS-A: rt 5.913 min; m/z 622
[M+H].sup.+.
(b) tert-Butyl
4-(5-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate
(A11)
[0442] A suspension of tert-butyl
4-(5-((4-((2-(1-(methoxycarbonyl)cyclopropyl)phenyl)ethynyl)-5-(trifluoro-
methyl)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate
(A10) (134 mg, 0.216 mmol), 10% Pd/C (100 mg) and Et.sub.3N (1.0
mL) in DMF (10 mL) was stirred under an atmosphere of hydrogen for
16 hours. The resulting mixture was diluted with EtOAc (60 mL) and
filtered through Celite. The filtrate was evaporated in vacuo and
the resulting residue was adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge,
10-50% EtOAc in petroleum benzine 40-60.degree. C.) to give the
title compound A11 as a colourless oil (117 mg, 87%); .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 8.65 (d, J=2.6 Hz, 1H), 8.57 (s, 1H),
8.15 (dd, J=8.5, 2.7 Hz, 1H), 7.45 (s, 1H), 7.34-7.27 (m, 3H),
7.24-7.15 (m, 2H), 4.26 (s, 2H), 3.60 (s, 3H), 3.22-3.07 (m, 4H),
2.92-2.76 (m, 3H), 1.92 (d, J=11.4 Hz, 2H), 1.79-1.65 (m, 4H), 1.48
(s, 9H), 1.22-1.18 (m, 2H). LCMS-A: rt 5.971 min; m/z 626
[M+H].sup.+.
(c) tert-Butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A12)
[0443] A solution of tert-butyl
4-(5-((4-(2-(1-(methoxycarbonyl)cyclopropyl)phenethyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A11)
(112 mg, 0.179 mmol) and LiOH.H.sub.2O (90 mg, 2.2 mmol) in THF
(7.0 mL), MeOH (7.0 mL) and H.sub.2O (2.0 mL) was stirred at
35.degree. C. for 24 hours. Additional LiOH.H.sub.2O (45 mg, 1.1
mmol) was added and stirring continued at 35.degree. C. for 16
hours. Further LiOH.H.sub.2O (45 mg, 1.07 mmol) was added and
stirring was continued at 35.degree. C. for 3 days. The volatiles
were removed in vacuo and the residue diluted with EtOAc (20 mL)
and aqueous HCl (2 M, 20 mL). The layers were separated and the
aqueous phase was extracted with EtOAc (2.times.50 mL). The
combined organics extracts were washed with brine and dried over
MgSO.sub.4. The volatiles were removed in vacuo and the residue
dissolved in anhydrous THF (9.0 mL) and anhydrous DMF (1.5 mL) then
HOBt (48 mg, 0.36 mmol), EDCl.HCl (68 mg, 0.36 mmol) and
diisopropylamine (0.310 mL, 2.21 mmol) were added under an
atmosphere of nitrogen. After 10 minutes of stirring ammonium
carbonate (86 mg, 0.89 mmol) was added in one portion and the
resulting mixture was stirred at room temperature for 16 hours. The
temperature was raised to 40.degree. C. and stirring was continued
for a further 24 hours. The volatiles were removed in vacuo before
EtOAc (20 mL) and saturated NaHCO.sub.3 (20 mL) were added to the
residue. After separating the layers, the aqueous phase was
extracted with EtOAc (2.times.20 mL) then the combined organic
extracts were washed with brine and dried over MgSO.sub.4. The
volatiles were removed under reduced pressure and the residue was
purified by silica gel column chromatography (Biotage Isolera, 24 g
SiO.sub.2 cartridge, 10-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A12 as a white solid
(77 mg, 71%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 10.32 (s,
1H), 8.76 (d, J=2.5 Hz, 1H), 8.72 (s, 1H), 8.14 (d, J=6.7 Hz, 1H),
7.37-7.26 (m, 4H), 7.26-7.18 (m, 1H), 6.98 (s, 1H), 6.02 (s, 1H),
4.05 (d, J=12.7 Hz, 2H), 3.21-3.07 (m, 4H), 2.87-2.75 (m, 3H), 1.81
(d, J=11.1 Hz, 2H), 1.56 (qd, J=13.0, 4.4 Hz, 2H), 1.47-1.38 (m,
11H), 0.97 (s, 2H). LCMS-A: rt 5.273 min; m/z 611 [M+H].sup.+.
(d)
1-(2-(2-(2-((6-(Piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)-
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (5)
[0444] A solution of TFA (0.386 mL, 5.04 mmol) and tert-butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)
pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A12)
(0.077 g, 0.13 mmol) in DCM (10.0 mL) was stirred at room
temperature for 3.5 h under N.sub.2. The volatiles were removed in
vacuo and the residue was dissolved in DCM (.about.1 mL).
Cyclohexane (.about.5 mL) was added until a white precipitate
formed. The resulting suspension was sonicated for 10 minutes and
the solid was isolated by filtration and dried to give the title
compound 5 as an off-white solid (0.042 g, 65%); .sup.1H NMR (400
MHz, d.sub.6-DMSO) .delta. 10.31 (s, 1H), 8.75 (d, J=2.4 Hz, 1H),
8.72 (s, 1H), 8.13 (d, J=7.3 Hz, 1H), 7.39-7.18 (m, 5H), 6.98 (s,
1H), 6.02 (s, 1H), 3.19-3.05 (m, 6H), 2.81-2.59 (m, 3H), 1.79 (d,
J=11.0 Hz, 2H), 1.63 (qd, J=12.5, 3.8 Hz, 2H), 1.47-1.41 (m, 2H),
1.02-0.92 (m, 2H). LCMS-A: rt 4.445 min; m/z 511 [M+H].sup.+.
Example 6
Synthesis of
1-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclop-
ropanecarboxamide (6)
##STR00126##
[0445] (a) 1-(2-((2,5-Dichloropyrimidin-4-yl)
ethynyl)phenyl)cyclopropanecarboxamide (A13)
[0446] PdCl.sub.2(PPh.sub.3).sub.2 (16 mg, 0.023 mmol) was added to
a stirred solution of 1-(2-ethynylphenyl)cyclopropanecarboxamide
(K6) (0.420 g, 2.26 mmol), 2,4,5-trichloropyrimidine (0.338 mL,
2.94 mmol), Et.sub.3N (1.26 mL, 9.07 mmol) and copper(I) iodide
(8.6 mg, 0.045 mmol) in 1,4-dioxane (4.5 mL). The resulting mixture
was stirred at 60.degree. C. for 2.5 hours under a nitrogen
atmosphere before being diluted with petroleum benzine
40-60.degree. C. (25 mL). The resultant precipitate was filtered,
dried, washed with water (50 mL) and dried to give the title
compound A13 as light coloured tan solid (0.570 g, 75%); .sup.1H
NMR (300 MHz, d.sub.6-DMSO) .delta. 9.01 (s, 1H), 7.72 (d, J=7.1
Hz, 1H), 7.41-7.57 (m, 3H), 6.97 (brs, 1H), 6.20 (brs, 1H), 1.53
(dq, J=4.0 Hz, 2H) 1.06 (dq, J=3.7 Hz, 2H). LCMS-B: rt 6.593 min;
m/z 332 [M+H].sup.+.
(b)
1-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxami-
de (A14)
[0447] A suspension of
1-(2-((2,5-dichloropyrimidin-4-yl)ethynyl)phenyl)cyclopropanecarboxamide
(A13) (0.536 g, 1.61 mmol) and platinum oxide (0.110 g, 0.484 mmol)
in DIPEA (16 mL) and MeOH (3 mL) was stirred under a hydrogen
atmosphere for 58 hours. The resulting mixture was filtered through
a pad of Celite and the filtrate concentrated in vacuo. The residue
was diluted with water and EtOAc and filtered through a pad of
Celite. The filtrate was extracted with EtOAc (2.times.20 mL) and
the combined organic fractions were washed with water (3.times.20
mL), brine, dried (magnesium sulfate), filtered and concentrated in
vacuo. The residue was adsorbed onto silica gel and purified by
column chromatography (CombiFlash Rf, 24 g SiO.sub.2 Cartridge,
30-60% EtOAc in cyclohexane) to give the title compound A14 as a
white solid (375 mg, 69%); .sup.1H NMR (300 MHz, CDCl.sub.3)
.delta. 8.52 (s, 1H), 7.43 (d, J=7.2 Hz, 1H), 7.28-7.31 (m, 3H),
5.37 (d, J=17 Hz, 2H), 3.27 (brs, 4H), 1.76 (brs, 2H), 1.16 (brs,
2H). LCMS-B: rt 5.30 min; m/z 337 [M+H].sup.+.
(c)
1-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyc-
lopropanecarboxamide (6)
[0448] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(A14) (100 mg, 297 .mu.mol), Cs.sub.2CO.sub.3 (291 mg, 892 .mu.mol)
and 3-aminopyridine (56.0 mg, 595 .mu.mol) in 1,4-dioxane (2.0 mL)
was sonicated for 10 minutes. To this Xantphos (6.88 mg, 11.9
.mu.mol) and Pd(OAc).sub.2 (1.33 mg, 5.95 .mu.mol) were added and
the resulting mixture heated under microwave irradiation for 20
minutes at 120.degree. C. The resulting mixture was adsorbed onto
silica gel and purified by column chromatography (Biotage Isolera,
24 g SiO.sub.2 cartridge, 0-50% MeOH in EtOAc) to yield a tan
solid. This was sonicated for 10 min in water (50 mL) and filtered
to give the title compound 6 as a green solid (29 mg, 25%); .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. 8.80-8.74 (d, J=2.5 Hz, 1H), 8.33
(s, 1H), 8.32-8.28 (dd, J=4.7, 1.3 Hz, 1H), 8.18-8.12 (m, 1H),
7.47-7.42 (d, J=7.2 Hz, 1H), 7.38-7.34 (m, 3H), 7.34-7.25 (m, 2H),
5.56 (s, 1H), 5.37 (s, 1H), 3.34-3.14 (m, 4H), 1.78 (s, 2H), 1.16
(s, 2H). LCMS-A: rt 4.502 min; m/z 394 [M+H].sup.+.
Example 7
Synthesis of
1-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclop-
ropanecarboxamide (7)
##STR00127##
[0449] (a) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)phenyl)piperidine-1-carboxylate (A15)
[0450] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(A14) (100 mg, 297 .mu.mol), Cs.sub.2CO.sub.3 (291 mg, 892 .mu.mol)
and tert-butyl 4-(4-aminophenyl)piperidine-1-carboxylate (164 mg,
595 .mu.mol) in 1,4-dioxane (2.0 mL) was sonicated for 10 min. To
this Xantphos (6.88 mg, 11.9 .mu.mol) and Pd(OAc).sub.2 (1.33 mg,
5.95 .mu.mol) were added and the resulting mixture heated under
microwave irradiation for 20 min at 120.degree. C. The resulting
mixture was adsorbed onto silica gel and purified by column
chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound A15 as a yellow oil (121 mg, 71%); .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 8.25-8.22 (s, 1H), 7.54-7.46 (m, 3H), 7.43-7.38
(dt, J=7.4, 1.0 Hz, 1H), 7.34-7.29 (m, 2H), 7.29-7.22 (m, 1H),
7.19-7.12 (m, 2H), 5.93-5.80 (s, 1H), 5.41-5.26 (s, 1H), 4.39-4.18
(s, 2H), 3.31-3.06 (m, 4H), 2.90-2.69 (t, J=11.8 Hz, 2H), 2.69-2.54
(tt, J=12.0, 3.4 Hz, 1H), 1.88-1.68 (t, J=15.8 Hz, 4H), 1.68-1.53
(qd, J=13.0, 4.4 Hz, 2H), 1.53-1.46 (s, 9H), 1.19-1.07 (s, 2H).
LCMS-A: rt 6.326 min; m/z 577 [M+H].sup.+.
(b)
1-(2-(2-(5-chloro-2-((4-(piperidin-4-yl)phenyl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (7)
[0451] TFA (1.0 mL) was added to a solution of tert-butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)phenyl)piperidine-1-carboxylate (A15) (121 mg, 210 .mu.mol) in
DCM (5 mL) and the resulting mixture was stirred at room
temperature for 16 hours. Saturated aqueous Na.sub.2CO.sub.3 was
added until the solution was basic then EtOAc (50 mL) and water (50
mL) were added. The organic layer was separated, washed with brine
(20 mL), dried over MgSO.sub.4 then the volatiles removed in vacuo.
The resulting residue was adsorbed onto silica gel and the
resulting material washed with MeOH (300 mL) then 1 M NH.sub.3 in
MeOH/EtOH (1:1) (300 mL). The volatiles from the ammoniacal washing
were removed in vacuo and the resultant gum sonicated in Et.sub.2O
(20 mL) to give a precipitate that was collected by filtration. Air
drying of the filter cake gave the product 7 as a white solid (55
mg, 55%); .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.28 (s, 1H),
7.52-7.46 (d, J=8.6 Hz, 2H), 7.46-7.41 (d, J=7.3 Hz, 1H), 7.37-7.32
(d, J=5.0 Hz, 2H), 7.31-7.25 (m, 2H, obscured), 7.23-7.18 (d, J=8.5
Hz, 2H), 7.15 (s, 1H), 5.33 (s, 2H), 3.34-3.12 (m, 6H), 2.84-2.71
(td, J=12.2, 2.3 Hz, 2H), 2.71-2.56 (tt, J=12.1, 3.7 Hz, 1H),
1.92-1.59 (m, 6H, obscured), 1.15 (s, 2H). LCMS-A: rt 4.527 min;
m/z 477 [M+H].sup.+.
Example 8
Synthesis of
1-(2-(2-(5-chloro-2-((4-(1-methylpiperidin-4-yl)phenyl)amino)pyrimidin-4--
yl)ethyl)phenyl)cyclopropanecarboxamide (8)
##STR00128##
[0453] Formaldehyde (37 wt % in H.sub.2O; 14 .mu.L, 0.19 mmol) was
added to a solution of
1-(2-(2-(5-chloro-2-((4-(piperidin-4-yl)phenyl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (7) (30 mg, 63 .mu.mol) in MeOH
(5.0 mL) under an atmosphere of nitrogen. The resulting mixture was
stirred for 10 minutes at room temperature before the addition of
sodium triacetoxyborohydride (53 mg, 0.25 mmol). Stirring was
continued for 2.5 hours at room temperature before the volatiles
were removed in vacuo and the resulting residue was partitioned
between EtOAc (20 mL) and saturated aqueous NaHCO.sub.3 (20 mL).
The layers were separated and the aqueous layer was extracted with
EtOAc (2.times.20 mL). The combined organic layers were washed with
brine (20 mL), dried over MgSO.sub.4 and the volatiles evaporated
in vacuo. The residue was dissolved in acetone (0.5 mL) and
petroleum benzine 40-60.degree. C. (25 mL) was added. The resulting
green precipitate was isolated by filtration and dried under high
vacuum to give the title compound 8 as a green solid (8 mg, 26%);
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.18 (s, 1H), 7.41 (d,
J=8.5 Hz, 2H), 7.34 (d, J=7.3 Hz, 1H), 7.30-7.23 (m, 3H), 7.12 (d,
J=8.5 Hz, 2H), 5.55 (s, 1H), 5.25 (s, 1H), 3.13 (J m, 6H),
2.52-2.41 (m, 1H), 2.38 (s, 3H), 2.23 (t, J=11.8 Hz, 2H), 1.89 (dd,
J=25.4, 12.3 Hz, 2H), 1.80 (d, J=12.3 Hz, 2H), 1.68 (s, 2H), 1.06
(s, 2H). LCMS-A: rt 4.611 min; m/z 491 [M+H].sup.+.
Example 9
Synthesis of
1-(2-(2-(5-chloro-2-((6-(trifluoromethyl)pyridin-3-yl)amino)pyrimidin-4-y-
l)ethyl)phenyl)cyclopropanecarboxamide (9)
##STR00129##
[0455] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(A14) (150 mg, 446 .mu.mol), Cs.sub.2CO.sub.3 (436 mg, 1.34 mmol)
and 5-amino-2-trifluoromethylpyridine (145 mg, 892 .mu.mol) in
1,4-dioxane (3 mL) was sonicated for 10 minutes. Xantphos (10.3 mg,
17.8 .mu.mol) and Pd(OAc).sub.2 (2.00 mg, 8.92 .mu.mol) were added
and the resulting mixture was heated under microwave irradiation
for 20 minutes at 120.degree. C. The resulting mixture was adsorbed
onto silica gel and purified by column chromatography (Biotage
Isolera, 40 g SiO.sub.2 cartridge, 0-100% EtOAc in petroleum
benzine 40-60.degree. C. then 0-40% MeOH in EtOAc). The purified
solid was dissolved in acetone (0.2 mL) and petroleum benzene
40-60.degree. C. (30 mL) added. The precipitate was collected by
vacuum filtration and the filter cake dried under high vacuum to
yield the title compound 9 as a tan solid (62 mg, 30%); .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 8.87 (d, J=2.6 Hz, 1H), 8.43-8.34 (m,
2H), 7.69 (d, J=8.7 Hz, 1H), 7.60 (s, 1H), 7.45 (d, J=8.2 Hz, 1H),
7.36 (dd, J=7.8, 3.1 Hz, 2H), 7.30 (d, J=4.9 Hz, 1H), 5.39 (d,
J=8.9 Hz, 2H), 3.35-3.17 (m, 4H), 1.78 (s, 2H), 1.16 (s, 2H).
LCMS-A: rt 4.816 min; m/z 462 [M+H].sup.+.
Example 10
Synthesis of
1-(2-(2-(5-(trifluoromethyl)-2-((6-(trifluoromethyl)pyridin-3-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (10)
##STR00130##
[0456] (a)
4-Chloro-5-(trifluoromethyl)-N-(6-(trifluoromethyl)pyridin-3-yl-
)pyrimidin-2-amine (A16)
[0457] A 1.0 M solution of zinc chloride in Et.sub.2O (18.5 mL,
18.5 mmol) was added to a solution of
2,6-dichloro-5-trifluoromethylpyrimidine (2.81 g, 13.0 mmol) in
t-BuOH/DCE (1:1, 50 mL) at 0.degree. C. over a period of 20 minutes
then the resulting mixture was stirred for 1 hour. A solution of
5-amino-2-trifluoromethylpyrimidine (2.00 g, 12.3 mmol) and DIPEA
(3.22 mL, 18.5 mmol) in t-BuOH/DCE (1:1, 30 mL) was added over 20
minutes then the resulting mixture was stirred at room temperature
for 4 days, before heating to 40.degree. C. and stirred for a
further 24 hours. The volatiles were removed under reduced pressure
to give a solid residue. Water (100 mL) was added and the
suspension sonicated for 30 minutes. The resulting mixture was
extracted with EtOAc (3.times.100 mL) and CHCl.sub.3 (100 mL). The
organic extracts were combined, passed through a phase separation
cartridge and the volatiles were evaporated under reduced pressure.
The residue was purified by silica gel column chromatography
(Biotage Isolera, 2.times.40 g SiO.sub.2, 0-25% EtOAc in petroleum
benzine 40-60.degree. C.) to give the title compound A16 as a white
solid (1.78 g, 42%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta.
11.21 (s, 1H), 9.02 (d, J=2.5 Hz, 1H), 8.96-8.89 (m, 1H), 8.40 (dd,
J=8.6, 2.2 Hz, 1H), 7.92 (d, J=8.6 Hz, 1H). LCMS-A: rt 5.931 min;
m/z 343 [M+H].sup.+.
(b)
1-(2-((5-(Trifluoromethyl)-2-((6-(trifluoromethyl)pyridin-3-yl)amino)p-
yrimidin-4-yl)ethynyl)phenyl)cyclopropanecarboxamide (A17)
[0458] A solution of 1-(2-ethynylphenyl)cyclopropanecarboxamide
(K6) (0.100 g, 0.540 mmol) in DMF (2.0 mL) was added to
4-chloro-5-(trifluoromethyl)-N-(6-(trifluoromethyl)pyridin-3-yl)pyrimidin-
-2-amine (A16) (0.185 g, 0.540 mmol), PdCl.sub.2(PPh.sub.3).sub.2
(0.019 g, 0.027 mmol), t-Bu.sub.3PH.BF.sub.4 (0.016 g, 0.054 mmol)
and CuI (0.010 g, 0.054 mmol) under nitrogen. The mixture was
degassed with nitrogen for 10 minutes then Et.sub.3N (1.0 mL) was
then added and the resulting mixture was heated under microwave
irradiation at 120.degree. C. for 15 minutes. The volatiles were
removed under reduced pressure, the residue was adsorbed onto
silica and purified by silica gel column chromatography (Biotage
Isolera, 24 g silica cartridge, 0-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A17 as a yellow solid
(0.157 g, 59%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. 11.05
(s, 1H), 9.05-8.93 (m, 2H), 8.64 (d, J=6.3 Hz, 1H), 7.93 (d, J=8.7
Hz, 1H), 7.62 (dd, J=7.6, 0.9 Hz, 1H), 7.58-7.39 (m, 3H), 7.00 (s,
1H), 6.17 (s, 1H), 1.54 (q, J=3.8 Hz, 2H), 1.10-1.02 (m, 2H).
LCMS-A: rt 5.851 min; m/z 492.2 [M+H].sup.+.
(c)
1-(2-(2-(5-(Trifluoromethyl)-2-((6-(trifluoromethyl)pyridin-3-yl)amino-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (10)
[0459] A suspension of 10% Pd/C (0.145 g) and
1-(2-((5-(trifluoromethyl)-2-((6-(trifluoromethyl)pyridin-3-yl)amino)pyri-
midin-4-yl)ethynyl)phenyl)cyclopropanecarboxamide (A17) (0.157 g,
0.320 mmol) in Et.sub.3N (1 mL) and DMF (10 mL) was stirred under
an atmosphere of hydrogen for 16 hours at 40.degree. C. The
resulting mixture was filtered through Celite and the filtrate
concentrated under reduced pressure. The residue was adsorbed onto
silica gel and purified by silica gel column chromatography
(Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-55% EtOAc in
petroleum benzine 40-60.degree. C.) to give the title compound 10
as a white solid (0.108 g, 68%); .sup.1H NMR (400 MHz,
d.sub.6-DMSO) .delta. 10.84 (s, 1H), 9.01 (d, J=2.2 Hz, 1H), 8.84
(s, 1H), 8.58 (dd, J=8.6, 2.1 Hz, 1H), 7.93 (d, J=8.7 Hz, 1H),
7.40-7.28 (m, 3H), 7.27-7.19 (m, 1H), 7.01 (s, 1H), 6.04 (s, 1H),
3.27-3.08 (m, 4H), 1.44 (d, J=3.4 Hz, 2H), 1.02-0.91 (m, 2H).
LCMS-A: rt 5.959 min; m/z 496 [M+H].sup.+.
Example 11
Synthesis of
1-(2-(2-(5-chloro-2-(oxazol-2-ylamino)pyrimidin-4-yl)ethyl)phenyl)cyclopr-
opanecarboxamide (11)
##STR00131##
[0460]
1-(2-(2-(5-Chloro-2-(oxazol-2-ylamino)pyrimidin-4-yl)ethyl)phenyl)c-
yclopropanecarboxamide (11)
[0461] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.150 g, 0.446 mmol), Cs.sub.2CO.sub.3 (436 mg, 1.34 mmol) and
2-aminoxazole (75.0 mg, 0.892 mmol) in 1,4-dioxane (3 mL) was
sonicated for 10 minutes. Xantphos (10 mg, 18 .mu.mol) and Pd(II)
acetate (2.0 mg, 8.9 .mu.mol) were added and the reaction was
irradiated in the microwave for 20 minutes at 120.degree. C. The
resulting mixture was adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge,
0-100% EtOAc in cyclohexane then 0-20% MeOH in EtOAc) to give a
solid. Acetone (0.5 mL) followed by cyclohexane (25 mL) was added
and the mixture was sonicated for 10 minutes. The resulting
precipitate was collected by filtration and the filter cake was
washed with cyclohexane (25 mL) and dried under high vacuum to give
the title compound 11 as a white solid (10 mg, 6%). LCMS-B: rt 4.64
min; m/z 384.0 [M+H].sup.+.
Example 12
Synthesis of
1-(2-(2-(5-chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)phenyl)cycl-
opropanecarboxamide (12)
##STR00132##
[0462]
1-(2-(2-(5-Chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)pheny-
l)cyclopropanecarboxamide (12)
[0463] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.100 g, 0.297 mmol), Cs.sub.2CO.sub.3 (0.291 g, 0.892 mmol)
and 5-aminopyrimidine (56.6 mg, 0.595 mmol) in 1,4-dioxane (3 mL)
was sonicated for 10 minutes. Xantphos (6.9 mg, 12 .mu.mol) and
Pd(II) acetate (1.3 mg, 5.9 .mu.mol) were added and the reaction
was irradiated in the microwave at 120.degree. C. for 20 minutes.
The resulting mixture was adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge,
0-100% EtOAc in petroleum benzine 40-60.degree. C. then 0-40% MeOH
in EtOAc) to give a solid which was suspended in 0.5 M aqueous
citric acid (50 mL) and sonicated for 10 minutes. The precipitate
was collected by filtration and the filter cake was washed with 2 M
aqueous NaOH (100 mL), cyclohexane (100 mL) and air dried to give
the title compound 12 as a yellow solid (25 mg, 21%). LCMS-B: rt
4.89 min; m/z 395.0 [M+H].sup.+.
Example 13
Synthesis of
1-(2-(2-(5-Chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (13)
##STR00133##
[0464]
1-(2-(2-(5-Chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)cyclopropanecarboxamide (13)
[0465] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (1.50 g, 4.46 mmol) in 1,4-dioxane (25 mL) containing
4-amino-1-methylpyrazole (0.520 g, 5.35 mmol) and p-toluenesulfonic
acid (0.085 g, 0.45 mmol) was heated under microwave irradiation at
120.degree. C. for 7 hours. The volatiles were removed in vacuo and
the residue was adsorbed onto silica gel and the product was
separated using silica column chromatography (Combiflash Rf, 40 g
SiO.sub.2 Cartridge, 60-100% EtOAc in cyclohexane and then 0-5%
MeOH in DCM) to give the title compound 13 as a light orange foam
(1.01 g, 56%). .sup.1H NMR (300 MHz, d6-DMSO) .delta. 9.64 (s, 1H),
8.37 (s, 1H), 7.87 (s, 1H), 7.46 (s, 1H), 7.22-7.36 (m, 4H), 7.00
(brs, 1H), 6.01 (brs, 1H), 3.81 (s, 3H), 3.09 (brs, 4H), 1.45-1.44
(m, 2H), 1.00-0.99 (m, 2H). LCMS-B: rt 6.318 min; m/z 397
[M+H].sup.+.
Example 14
Synthesis of
1-(2-(2-(2-((1-(azetidin-3-yl)-1H-pyrazol-4-yl)amino)-5-chloropyrimidin-4-
-yl)ethyl)phenyl)cyclopropanecarboxamide (14)
##STR00134##
[0466] (a) tert-Butyl
3-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)azetidine-1-carboxylate (A18)
[0467] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.100 g, 0.297 mmol), Cs.sub.2CO.sub.3 (0.291 g, 0.892 mmol)
and tert-butyl 3-(4-amino-1H-pyrazol-1-yl)azetidine-1-carboxylate
(0.142 g, 0.595 mmol) in dioxane (3 mL) was sonicated for 10
minutes. Xantphos (6.9 mg, 12 .mu.mol) and Pd(II) acetate (1.3 mg,
5.9 .mu.mol) were added to the suspension and the mixture was
irradiated in the microwave for 20 minutes at 120.degree. C. The
resulting mixture was adsorbed onto silica gel and purified by
column chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge,
0-100% EtOAc in petroleum benzine 40-60.degree. C. then 0-40% MeOH
in EtOAc) to give the title compound A18 as a yellow oil (110 mg,
69%). LCMS-A: rt 6.349 min; m/z 538.3 [M+H].sup.+.
(b)
1-(2-(2-(2-((1-(Azetidin-3-yl)-1H-pyrazol-4-yl)amino)-5-chloropyrimidi-
n-4-yl)ethyl)phenyl)cyclopropanecarboxamide (14)
[0468] TFA (1 mL) was added to a solution of tert-butyl
3-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)azetidine-1-carboxylate A18 (0.110 g, 0.204
mmol) in DCM (5 mL) and the reaction was stirred for 16 hours. The
volatiles were removed in vacuo and the resultant residue loaded
onto an SCX cartridge (5 g). The cartridge was washed with MeOH
(100 mL) and then with 5% NH.sub.4OH in MeOH (100 mL). The basic
fractions were combined and the solvent was removed in vacuo to
give the title compound 14 as a tan solid (61 mg, 68%). .sup.1H NMR
(400 MHz, d.sub.4-MeOD) .delta. 8.27 (s, 1H), 8.09 (s, 1H), 7.65
(s, 1H), 7.44-7.37 (m, 2H), 7.34 (m, 1H), 7.27 (m, 1H), 5.36-5.18
(m, 1H), 4.15 (m, 2H), 3.96 (m, 2H), 3.20 (m, 4H), 1.63 (m, 2H),
1.20-1.05 (m, 2H). LCMS-C: rt 4.30 min; m/z 438.0 [M+H].sup.+.
Example 15
Synthesis of
1-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)cyclopropanecarboxamide (15)
##STR00135##
[0469] (a) tert-Butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)pyridin-2-yl)piperidine-1-carboxylate (A19)
[0470] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.150 g, 0.446 mmol), Cs.sub.2CO.sub.3 (0.436 g, 1.34 mmol)
and tert-butyl 4-(5-aminopyridin-2-yl)piperidine-1-carboxylate I9
(247 mg, 0.892 mmol) in 1,4-dioxane (3 mL) was sonicated for 10
minutes. Xantphos (10 mg, 18 .mu.mol) and Pd(II) acetate (2.0 mg,
8.9 .mu.mol) were added and the mixture was irradiated in the
microwave at 120.degree. C. for 20 minutes. The mixture was
adsorbed onto silica gel and purified by column chromatography
(Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-100% EtOAc in
cyclohexane) to give the title compound A19 as a yellow oil (79 mg,
31%). LCMS-C: rt 4.92 min; m/z 577.0 [M+H].sup.+.
(b)
1-(2-(2-(5-Chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-
-yl)ethyl)phenyl)cyclopropanecarboxamide (15)
[0471] TFA (1 mL) was added to a solution of tert-butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)pyridin-2-yl)piperidine-1-carboxylate A19 (79.0 mg, 0.137 mmol)
in DCM (5 mL) and stirred for 16 hours. The volatiles were removed
in vacuo and the resultant residue loaded onto an SCX cartridge (5
g). The cartridge was washed with MeOH (100 mL) and then with 5%
NH.sub.4OH in MeOH (100 mL). The basic fractions were combined and
the solvent was removed in vacuo to give the title compound 15 as a
white solid (50 mg, 77%). .sup.1H NMR (400 MHz, d.sub.4-MeOD)
.delta. 8.74 (s, 1H), 8.36 (s, 1H), 8.21 (dd, J=8.6, 2.7 Hz, 1H),
7.45-7.22 (m, 5H), 3.30-3.17 (m, 6H), 2.95-2.80 (m, 3H), 1.99-1.96
(m, 2H), 1.86-1.76 (m, 2H), 1.64-1.63 (m, 2H), 1.19-1.07 (m, 2H).
LCMS-C: rt 4.17 min; m/z 477.1 [M+H].sup.+.
Example 16
Synthesis of
1-(2-(2-(5-chloro-2-(5-(1-methylpiperidin-3-yl)pyridin-3-ylamino)pyrimidi-
n-4-yl)ethyl)phenyl)cyclopropanecarboxamide (16)
##STR00136##
[0472]
1-(2-(2-(5-Chloro-2-(5-(1-methylpiperidin-3-yl)pyridin-3-ylamino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (16)
[0473] Formaldehyde (32.8 .mu.L, 0.440 mmol, 37 wt % in H.sub.2O)
was added to a suspension of
1-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)cyclopropanecarboxamide 15 (42.0 mg, 0.088 mmol) in
MeOH (1.10 mL) under an atmosphere of nitrogen. Sodium
triacetoxyborohydride (187 mg, 0.881 mmol) was then added in one
portion and the mixture was stirred at room temperature overnight.
The volatiles were removed in vacuo and the residue was diluted
with EtOAc (10 mL) and sat. aq. NaHCO.sub.3 (10 mL). The layers
were separated and the aqueous layer was extracted with EtOAc
(2.times.15 mL). The combined organic layers were washed with water
(20 mL) and brine (20 mL) and dried over Na.sub.2SO.sub.4. The
solvent was removed under reduced pressure to yield a white solid
which was purified by prep-LCMS to give the title compound 16 as a
white solid (5.90 mg, 14%). .sup.1H NMR (300 MHz, CDCl.sub.3)
.delta. 8.69 (brs, 1H), 8.28 (s, 1H), 8.10 (dd, J=8.47, 2.53 Hz,
1H), 7.55-7.62 (m, 1H), 7.43 (d, J=7.04 Hz, 1H), 7.32-7.36 (m, 2H),
7.20-7.32 (m, 1H), 5.94 (brs, 1H), 5.40 (brs, 1H), 3.32-3.42 (m,
4H), 3.22-3.32 (m, 2H), 3.12-3.21 (m, 2H), 2.80-2.92 (m, 1H),
2.59-2.73 (m, 3H), 2.05-2.33 (m, 4H), 1.72-1.79 (m, 2H), 1.10-1.17
(m, 2H). LCMS-C: rt 5.08 min; m/z 491.3 [M+H].sup.+.
Example 17
Synthesis of
1-(2-(2-(5-chloro-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (17)
##STR00137##
[0474] (a) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate (A20)
[0475] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.100 g, 0.297 mmol), Cs.sub.2CO.sub.3 (0.291 g, 0.892 mmol)
and tert-butyl 4-(4-amino-1H-pyrazol-1-yl)piperidine-1-carboxylate
(0.158 g, 0.595 mmol) in 1,4-dioxane (2 mL) was sonicated for 10
minutes. Xantphos (6.9 mg, 12 .mu.mol) and Pd(II) acetate (1.3 mg,
5.9 .mu.mol) were added and the mixture was irradiated in the
microwave at 120.degree. C. for 20 minutes. The mixture was
adsorbed onto silica gel and purified by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100% EtOAc in
cyclohexane then 0-10% MeOH in EtOAc) to give the title compound
A20 as a brown solid (21 mg, 12%). LCMS-C: rt 5.54 min; m/z 565.9
[M+H].sup.+.
(b)
1-(2-(2-(5-Chloro-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrimid-
in-4-yl)ethyl)phenyl)cyclopropanecarboxamide (17)
[0476] TFA (1 mL) was added to a solution of tert-butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate A20 (0.021 g, 0.037
mmol) in DCM (5 mL) and the mixture was stirred for 16 hours. The
volatiles were removed in vacuo and the resultant residue loaded
onto a SCX cartridge (5 g). The cartridge was washed with MeOH (100
mL) and then with 5% NH.sub.4OH in MeOH (100 mL). The basic
fractions were combined, the solvent was removed in vacuo and the
resultant residue purified by prep-LCMS to give the title compound
17 as a yellow solid (3.6 mg, 21%). LCMS-B: rt 4.711 min; m/z 466.2
[M+H].sup.+.
Example 18
Synthesis of
1-(2-(2-(5-chloro-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (18)
##STR00138##
[0477]
1-(2-(2-(5-Chloro-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)am-
ino)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(DGG.sub.--011.sub.--20.sub.--06) (CTX-0358024)
[0478] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.060 g, 0.21 mmol) in MeOH (1.0 mL) and water (0.1 mL)
containing 1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-amine (0.039 g,
0.21 mmol) was heated at 70.degree. C. for 48 hours. The mixture
was adsorbed onto silica gel and purified by silica column
chromatography (Combiflash Rf, 4 g SiO.sub.2 Cartridge, 0-50% MeOH
in DCM). Further purification by HPLC (eluting with 30-100%
gradient in acetonitrile containing 0.1% formic acid and water
containing 0.1% formic acid, flow rate 4.0 mL/min) gave the title
compound 18 as a light yellow solid (0.015 g, 17%). LCMS-C: rt 4.31
min; m/z 480 [M+H].sup.+.
Example 19
Synthesis of
1-(2-(2-(2-(1H-pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phenyl)cyc-
lopropanecarboxamide (19)
##STR00139##
[0479]
1-(2-(2-(2-(1H-Pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phen-
yl)cyclopropanecarboxamide (19)
[0480] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.070 g, 0.21 mmol) and tert-butyl
4-amino-1H-pyrazole-1-carboxylate (0.11 g, 0.63 mmol) was stirred
in MeOH:water (10:1 ratio, 4 mL) at 70.degree. C. for 2 hours and
then at 90.degree. C. for 16 hours. The volatiles were removed in
vacuo and the residue was purified by silica gel column
chromatography (Combiflash Rf, 0-15% MeOH in DCM) to give the title
compound 19 as a light yellow solid (0.015 g, 19%). .sup.1H NMR
(300 MHz, d.sub.6-DMSO) .delta. 0.93-1.02 (m, 2H), 1.38-1.49 (m,
2H), 3.04-3.16 (m, 4H), 6.03 (brs, 1H), 6.98 (brs, 1H), 7.18-7.39
(m, 4H), 7.48-7.72 (m, 1H), 7.79-8.00 (m, 1H), 8.37 (s, 1H), 9.62
(s, 1H). LCMS-C: rt 4.92 min; m/z 383 [M+H].sup.+.
Example 20
Synthesis of
1-(2-(2-(5-chloro-2-(1-(pyrrolidin-3-yl)-1H-pyrazol-4-ylamino)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (20)
##STR00140##
[0481] (a) tert-Butyl
3-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-ylamino-
)-1H-pyrazol-1-yl)pyrrolidine-1-carboxylate (A21)
[0482] A stirred solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.150 g, 0.446 mmol),
4-amino-1-(1-Boc-pyrrolidin-3-yl)-1H-pyrazole (0.225 g, 0.892 mmol)
in MeOH (10 mL) and water (1.0 mL) was heated at 70.degree. C. for
2 days. Additional 4-amino-1-(1-boc-pyrrolidin-3-yl)-1H-pyrazole
(0.125 g, 0.446 mmol) was added and the mixture was heated for a
further 16 hours at reflux. After cooling the solvent was removed
to afford a crude red oil which was purified by silica gel column
chromatography (0-100% EtOAc in cyclohexane) to give the title
compound A21 as a red oil (0.164 g, 67% yield). LCMS-C: rt 5.51
min; m/z 551.9 [M+H].sup.+.
(b)
1-(2-(2-(5-Chloro-2-(1-(pyrrolidin-3-yl)-1H-pyrazol-4-ylamino)pyrimidi-
n-4-yl)ethyl)phenyl)cyclopropanecarboxamide (20)
[0483] To a stirred solution of tert-butyl
3-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-ylamino-
)-1H-pyrazol-1-yl)pyrrolidine-1-carboxylate A21 (0.164 g, 0.297
mmol) in DCM (5 mL) was added 4 M HCl solution in 1,4-dioxane
(0.223 mL). The mixture was stirred for 5 hours before additional 4
M HCl in dioxane (3 eq.) was added and the mixture stirred for
another 16 hours. Another portion of 4 M HCl in 1,4-dioxane (2 mL)
was added and the mixture was stirred for 16 hours. The solvent was
removed and the residue was diluted with 1 M HCl aq. (.about.5 mL)
and washed with EtOAc (2.times.10 mL). The aqueous layer was
basified with 1 M NaOH and extracted with EtOAc (3.times.10 mL).
The combined organics were dried (MgSO.sub.4), filtered and
concentrated in vacuo to give a crude white paste that was purified
by flash chromatography (0-100% EtOAc in cyclohexane) to give the
title compound as a red oil. Further purification by HPLC
(Gradient: 20-100%, Acetonitrile 0.1% formic acid in water 0.1%
formic acid, 12 min) gave the title compound 20 as a clear gum
(0.012 g, 9% yield). .sup.1H NMR (300 MHz, MeOD) .delta. 1.14 (d,
J=3.30 Hz, 2H), 1.64 (d, J=3.30 Hz, 2H), 2.32-2.44 (m, 1H),
2.46-2.65 (m, 1H), 3.12-3.26 (m, 4H), 3.43-3.54 (m, 1H), 3.60-3.80
(m, 3H), 5.21-5.32 (m, 1H), 7.23-7.45 (m, 4H), 7.64 (s, 1H), 8.08
(m, 1H), 8.29 (m, 1H). LCMS-C: rt 4.29 min; m/z 452
[M+H].sup.+.
Example 21
Synthesis of
1-(2-(2-(5-chloro-2-(1-(1-methylpyrrolidin-3-yl)-1H-pyrazol-4-ylamino)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (21)
##STR00141##
[0484] (a) 1-Methylpyrrolidin-3-yl methanesulfonate (A22)
[0485] 3-Hydroxy-1-methylpyrroldine (0.543 mL, 4.94 mmol) was
dissolved in DCM (10 mL) and cooled to 0.degree. C. Et.sub.3N
(0.827 mL, 5.93 mmol), methanesulfonyl chloride (0.421 mL, 5.44
mmol) and DMAP (0.006 g, 0.05 mmol) were added and the mixture was
stirred for 16 hours at room temperature. The mixture was diluted
with CHCl.sub.3 (5 mL) and washed with sat. NaHCO.sub.3 (5 mL) and
water (2.times.5 mL). The organic layer was concentrated in vacuo
to give the title compound A22 as a yellow oil (0.717 g, 81%
yield). .sup.1H NMR (300 MHz, CDCl.sub.3) .delta. 1.85-2.24 (m,
6H), 2.38-2.74 (m, 3H), 2.76-2.91 (m, 3H), 4.95 (m, 1H).
(b) 1-(1-Methylpyrrolidin-3-yl)-4-nitro-1H-pyrazole (A23)
[0486] 4-Nitro-1H-pyrazole (0.452 g, 4.00 mmol) was dissolved in
DMF (8 mL) and cooled to 0.degree. C. NaH (0.192 g, 60% dispersion
in oil, 4.80 mmol) was added and the mixture was stirred for 10
minutes. 1-Methylpyrrolidin-3-yl methanesulfonate A22 (0.717 g,
4.00 mmol) was added and the mixture was stirred for 16 hours at
100.degree. C. The mixture was diluted with EtOAc (5 mL) and washed
with sat. NH.sub.4Cl (5 mL) and water (4.times.5 mL). The organics
were concentrated in vacuo affording a crude pale yellow oil which
was purified by flash chromatography (0-30% MeOH in DCM) to give
the title compound A23 as a pale yellow oil (0.575 g, 73% yield).
LCMS-B: rt 1.12 min; m/z 197 [M+H].sup.+.
(c) 1-(1-Methylpyrrolidin-3-yl)-1H-pyrazol-4-amine (A24)
[0487] A solution of
1-(1-methylpyrrolidin-3-yl)-4-nitro-1H-pyrazole A23 (0.575 g, 2.93
mmol) and 10% Pd/C (0.061 g) in EtOH (15 mL) was stirred under a
hydrogen atmosphere for 16 hours. The reaction mixture was filtered
through Celite and the solvent was removed in vacuo to give the
title compound A24 as a red oil (0.555 g, quantitative). .sup.1H
NMR (300 MHz, MeOD) .delta. 2.27-2.42 (m, 1H), 2.72-2.76 (m, 2H),
3.02-3.10 (m, 1H), 3.46-3.55 (m, 1H), 3.70-3.85 (m, 4H), 5.14-5.26
(m, 1H), 7.33 (s, 1H), 7.46 (m, 1H).
(d)
1-(2-(2-(5-Chloro-2-(1-(1-methylpyrrolidin-3-yl)-1H-pyrazol-4-ylamino)-
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (21)
[0488] A stirred solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.085 g, 0.25 mmol),
1-(1-methylpyrrolidin-3-yl)-1H-pyrazol-4-amine A24 (0.046 g, 0.278
mmol) and TsOH (0.003 g, 0.02 mmol) in 1,4-dioxane (4 mL) was
stirred at 120.degree. C. in the microwave for 60 minutes followed
by 150.degree. C. for 2 hours and then 160.degree. C. for 2 hours.
After cooling, the solvent was removed to afford a crude red oil
that was purified by flash chromatography (0-30% MeOH in DCM)
followed by HPLC (Gradient: 20-100%, Acetonitrile 0.1% formic acid
in water 0.1% formic acid, 12 min) to give the title compound 21 as
a clear gum (0.008 g, 7% yield). LCMS-C: rt 4.34 min; m/z 466
[M+H].sup.+.
Example 22
Synthesis of
1-(2-(2-(5-chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)phenyl)cycl-
opropanecarboxamide (22)
##STR00142##
[0489] (a)
1-(2-(2-(5-Chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl)cyclopropanecarboxamide (22)
[0490] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 0.080 g, 0.24 mmol), 4-aminopyridazine (0.045 g, 0.48 mmol),
Xantphos (0.0055 g, 0.010 mmol) and Cs.sub.2CO.sub.3 (0.23 g, 0.71
mmol) in 1,4-dioxane (4 mL) was bubbled with nitrogen for 10
minutes. Palladium(II) acetate (0.0011 g, 0.0049 mmol) was added
and the mixture was heated in the microwave at 120.degree. C. for
25 minutes. The volatiles were removed in vacuo and the residue was
purified by silica gel column chromatography (Combiflash Rf, 0-15%
MeOH in DCM) to give the title compound 22 as a light yellow solid
(0.010 g, 11%). LCMS-C: rt 4.34 min; m/z 395 [M+H].sup.+.
Example 23
Synthesis of
1-(2-(2-(5-chloro-2-(phenylamino)pyrimidin-4-yl)ethyl)phenyl)cyclopropane-
carboxamide (23)
##STR00143##
[0491] (a)
1-(2-(2-(5-Chloro-2-(phenylamino)pyrimidin-4-yl)ethyl)phenyl)cy-
clopropanecarboxamide (23)
[0492] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.080 g, 0.24 mmol) in 1,4-dioxane (1.0 mL) containing aniline
(0.043 mL, 0.48 mmol) and p-toluenesulfonic acid (4.5 mg, 0.024
mmol) was heated under microwave irradiation at 120.degree. C. for
4 hours. The reaction mixture was adsorbed onto silica gel and
purified by silica column chromatography (Combiflash Rf, 12 g
SiO.sub.2 Cartridge, 20-50% EtOAc in cyclohexane) to give the title
product 23 as a light yellow foam (0.044 g, 46%). LCMS-B: rt 7.718
min; m/z 393 [M+H].sup.+.
Example 24
Synthesis of tert-Butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate (24)
##STR00144##
[0493] tert-Butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate (24)
[0494] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.080 g, 0.24 mmol), tert-butyl
1-(4-aminophenyl)ethylcarbamate (0.067 g, 0.29 mmol), Xantphos
(0.0057 g, 0.010 mmol) and Cs.sub.2CO.sub.3 (0.23 g, 0.71 mmol) in
1,4-dioxane (4 mL) was bubbled with nitrogen for 10 minutes.
Palladium(II) acetate (0.0010 g, 0.0045 mmol) was added and the
mixture was heated at 120.degree. C. under microwave irradiation
for 25 minutes. The mixture was partitioned between water and
EtOAc. The layers were separated and the aqueous was extracted with
EtOAc (2 times). The combined organic extracts were washed with
brine, dried (Na.sub.2SO.sub.4) and the solvent evaporated in vacuo
to give the crude product. Purification by silica gel column
chromatography (Combiflash Rf, 0-100% EtOAc in cyclohexane) gave
the title compound 24 as a colourless oil (0.015 g, 12%). LCMS-C:
rt 5.79 min; m/z 536 [M+H].sup.+.
Example 24A
Synthesis of
1-(2-(2-(2-((4-(1-aminoethyl)phenyl)amino)-5-chloropyrimidin-4-yl)ethyl)p-
henyl)cyclopropanecarboxamide (24A)
##STR00145##
[0495]
1-(2-(2-(2-((4-(1-Aminoethyl)phenyl)amino)-5-chloropyrimidin-4-yl)e-
thyl)phenyl)cyclopropanecarboxamide (24A)
[0496] To a solution of tert-butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-chloropyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate 24 (0.012 g, 0.022 mmol) in DCM (1 mL)
was added trifluoroacetic acid (0.5 mL). The mixture was stirred at
ambient temperature for 2 hours before the volatiles were removed
in vacuo. The crude residue was loaded onto an SCX cartridge
conditioned with methanol. The cartridge was washed with methanol
and then 2 N ammonia in ethanol. The basic fractions were combined
and the solvent removed in vacuo to give the title compound 24A as
a colourless oil (0.009 g, 92%). LCMS-C: rt 4.41 min; m/z 436
[M+H].sup.+.
Example 25
Synthesis of
1-(2-(2-(5-Chloro-2-((3-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (25)
##STR00146##
[0497] (a) tert-Butyl 3-methyl-4-nitro-1H-pyrazole-1-carboxylate or
tert-butyl 5-methyl-4-nitro-1H-pyrazole-1-carboxylate (A25)
[0498] Di-tert-butyl dicarbonate (5.15 g, 23.6 mmol) and
4-dimethylaminopyridine (0.481 g, 3.93 mmol) were added to a
solution of 3-methyl-4-nitropyrazole (2.50 g, 19.7 mmol) in DCM
(100 mL) and the mixture was stirred at room temperature for 16
hours. The reaction mixture was washed with water (100 mL), brine
(100 mL), dried (phase separator) and concentrated under reduced
pressure. The residue was adsorbed onto SiO.sub.2 and purified by
column chromatography (Biotage Isolera, 2.times.40 g SiO.sub.2
cartridges, 0-50% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A25 as a white solid (2.54 g, 57%). LCMS-D:
rt 3.39 min; no product ion detected.
(b) tert-Butyl 4-amino-3-methyl-1H-pyrazole-1-carboxylate or
tert-butyl 4-amino-5-methyl-1H-pyrazole-1-carboxylate (A26)
[0499] A solution of tert-butyl
3-methyl-4-nitro-1H-pyrazole-1-carboxylate or tert-butyl
5-methyl-4-nitro-1H-pyrazole-1-carboxylate (isomer not determined)
A25 (1.50 g, 6.60 mmol) in EtOH (75 mL) was stirred over 10% Pd/C
(wetted with ca. 53% water, 0.150 g) under an atmosphere of H.sub.2
at room temperature for 16 hours. The mixture was diluted with
EtOAc (50 mL) and filtered through Celite. Evaporation of the
volatiles in vacuo gave the title compound A26 as a pink solid
(1.30 g, >95%). LCMS-D: rt 2.78 min; m/z 198 [M+H].sup.+.
(c)
1-(2-(2-(5-Chloro-2-((3-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (25)
[0500] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.120 g, 0.357 mmol), tert-butyl
4-amino-3-methyl-1H-pyrazole-1-carboxylate or tert-butyl
4-amino-5-methyl-1H-pyrazole-1-carboxylate (isomer not determined)
A26 (0.141 g, 0.714 mmol) and p-toluenesulfonic acid monohydrate
(0.007 g, 0.04 mmol) in 1,4-dioxane (2.0 mL) was stirred in a
microwave reactor at 120.degree. C. for 3 hours. The reaction
mixture was adsorbed onto SiO.sub.2 and purified by column
chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-10%
MeOH in DCM) to give the title compound 25 as a white solid (0.035
g, 25%). LCMS-A: rt 5.62 min; m/z 397 [M+H].sup.+.
Example 26
Synthesis of
1-(2-(2-(5-Chloro-2-((1,3-dimethyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)e-
thyl)phenyl)cyclopropanecarboxamide (26)
##STR00147##
[0501] (a)
1-(2-(2-(5-Chloro-2-((1,3-dimethyl-1H-pyrazol-4-yl)amino)pyrimi-
din-4-yl)ethyl)phenyl)cyclopropanecarboxamide (26)
[0502] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.100 g, 0.297 mmol), 1,3-dimethyl-1H-pyrazol-4-amine (0.070
g, 0.63 mmol) and p-toluenesulfonic acid monohydrate (0.006 g,
0.032 mmol) in 1,4-dioxane (1.0 mL) was stirred in the microwave at
120.degree. C. for 2 hours. Additional 1,4-dioxane (1.5 mL) was
added and the reaction mixture was stirred in the microwave at
120.degree. C. for a further 4 hours. The reaction mixture was
adsorbed onto SiO.sub.2 and purified by column chromatography
(Biotage Isolera, 24 g SiO.sub.2 cartridge, 15-100% EtOAc in
petroleum benzine 40-60.degree. C.). Fractions containing suspected
product were combined and adsorbed onto silica and purified by
column chromatography (Biotage Isolera, 12 g SiO.sub.2, 0-5% MeOH
in DCM) to give the title compound 26 as a pale yellow solid (0.030
g, 25%). LCMS-D: rt 3.24 min; m/z 411 [M+H].sup.+.
Example 27
Synthesis of
1-(2-(2-(5-chloro-2-((5-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (27)
##STR00148##
[0503] (a) Mixture of tert-butyl
3-amino-5-methyl-1H-pyrazole-1-carboxylate and tert-butyl
5-amino-3-methyl-1H-pyrazole-1-carboxylate (A27)
[0504] Di-tert-butyl dicarbonate (531 mg, 2.57 mmol) was added to a
mixture of 3-amino-5-methylpyrazole (250 mg, 2.57 mmol) and KOH
(289 mg, 5.15 mmol) in THF (25 mL). The reaction mixture was
stirred at room temperature overnight and the volatiles were
removed in vacuo. The resulting gum was diluted with EtOAc (100 mL)
and washed with water (100 mL). The organic layer was separated,
adsorbed onto silica gel and purified by silica gel column
chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge, 0-50%
EtOAc in petroleum benzine 40-60.degree. C.) to give the title
compound/s A27 as a white solid (154 mg, 30%). LCMS-A: rt 5.237
min.
[0505] Note: No assignment of Boc position on the pyrazole was
made.
(b)
1-(2-(2-(5-Chloro-2-((5-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (27)
[0506] A suspension of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (263 mg, 0.781 mmol), Cs.sub.2CO.sub.3 (509 mg, 1.56 mmol) and
a mixture of tert-butyl 3-amino-5-methyl-1H-pyrazole-1-carboxylate
and tert-butyl 5-amino-3-methyl-1H-pyrazole-1-carboxylate A27 (154
mg, 0.781 mmol) in 1,4-dioxane (5 mL) was sonicated for 10 minutes.
Xantphos (23 mg, 0.039 mmol) and Pd.sub.2(dba).sub.3 (36 mg, 0.039
mmol) were added and the mixture was irradiated in the microwave at
120.degree. C. for 20 minutes. The resulting mixture was adsorbed
onto silica gel and purified by column chromatography (Biotage
Isolera, 40 g SiO.sub.2 cartridge, 0-100% EtOAc in petroleum
benzine 40-60.degree. C.) to give a yellow oil which was dissolved
in DCM (5.0 mL). TFA (1.0 mL) was added and the mixture was stirred
at room temperature for 2 hours before the addition of sat. aq.
Na.sub.2CO.sub.3 (20 mL) and water (100 mL). The resultant
precipitate was collected by filtration, adsorbed onto silica gel
and purified by silica gel column chromatography (Biotage Isolera,
12 g SiO.sub.2 cartridge, 0-100% EtOAc in petroleum benzine
40-60.degree. C. then 0-30% MeOH in EtOAc) to give the title
compound 27 as a yellow solid (8 mg, 3%). LCMS-A: rt 5.412 min; m/z
397.2 [M+H].sup.+.
Example 28
Synthesis of
1-(2-(2-(5-Chloro-2-((1-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (28)
##STR00149##
[0507]
1-(2-(2-(5-Chloro-2-(1-methyl-1H-pyrazol-3-yl)amino)pyrimidin-4-yl)-
ethyl)phenyl)cyclopropanecarboxamide (28)
[0508] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.100 g, 0.297 mmol), 1-methyl-1H-pyrazole-3-amine (0.058 g,
0.595 mmol) and p-toluenesulfonic acid monohydrate (0.006 g, 0.030
mmol) in 1,4-dioxane (2.0 mL) was stirred in a microwave reactor at
120.degree. C. for 3 hours. The reaction mixture was adsorbed onto
SiO.sub.2 and purified by column chromatography (Biotage Isolera,
24 g SiO.sub.2 cartridge, 0-5% MeOH in DCM) to give a gummy solid.
The solid was suspended in a mixture of Et.sub.2O and petroleum
benzine 40-60.degree. C. (1:1) and sonicated for 10 minutes. The
precipitate was isolated by vacuum filtration and dried to give the
title compound 28 as a yellow solid (0.014 g, 12%). LCMS-D: rt 3.26
min; m/z 397 [M+H].sup.+.
Example 29
Synthesis of
1-(2-(2-(5-chloro-2-((1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-yl)amino)pyri-
midin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (29)
##STR00150##
[0509]
1-(2-(2-(5-Chloro-2-((1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-yl)amin-
o)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (29)
[0510] A mixture of 1-(2,2,2-trifluoroethyl)-1H-pyrazol-4-amine
(0.160 g, 0.967 mmol),
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.101 g, 0.300 mmol) and TsOH.H.sub.2O (0.010 g, 0.051 mmol)
in 1,4-dioxane (2.0 mL) was heated to 120.degree. C. for 3 hours in
the microwave. The mixture was concentrated under reduced pressure
and purified using silica gel column chromatography (0-100% EtOAc
in petroleum benzine 40-60.degree. C.) to give the title compound
29 (0.036 g, 26%). LCMS-D: rt 3.402 min; m/z 465.2 [M+H].sup.+.
Example 30
Synthesis of
1-(2-(2-(5-chloro-2-((1-isopropyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (30)
##STR00151##
[0511]
1-(2-(2-(5-Chloro-2-((1-isopropyl-1H-pyrazol-4-yl)amino)pyrimidin-4-
-yl)ethyl)phenyl)cyclopropanecarboxamide (30)
[0512] A mixture of 1-isopropylpyrazol-4-amine (0.083 g, 0.66
mmol),
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.110 g, 0.327 mmol) and TsOH.H.sub.2O (0.013 g, 0.066 mmol)
in 1,4-dioxane (2.0 mL) was heated in the microwave to 100.degree.
C. for 3 hours. The mixture was concentrated under reduced pressure
and purified by silica gel column chromatography (0-100% EtOAc in
petroleum benzine 40-60.degree. C. then 0-10% MeOH in EtOAc) to
give the title compound 30 (0.094 g, 68%). LCMS-D: rt 3.355 min;
m/z 425.3 [M+H].sup.+.
Example 31
Synthesis of
1-(2-(2-(5-chloro-2-((6-cyanopyridin-3-yl)amino)pyrimidin-4-yl)ethyl)phen-
yl)cyclopropanecarboxamide (31)
##STR00152##
[0513]
1-(2-(2-(5-Chloro-2-((6-cyanopyridin-3-yl)amino)pyrimidin-4-yl)ethy-
l)phenyl)cyclopropanecarboxamide (31)
[0514] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.300 g, 0.892 mmol) in THF (2.5 mL) containing
5-amino-2-pyridinecarbonitrile (0.213 g, 1.78 mmol),
Cs.sub.2CO.sub.3 (0.872 g, 2.67 mmol), Xantphos (0.021 g, 0.036
mmol) and palladium (II) acetate (4 mg, 0.018 mmol) was heated
under microwave irradiation for 30 minutes at 120.degree. C. The
reaction mixture was adsorbed onto silica gel and purified by
silica column chromatography (Combiflash Rf, 12 g SiO.sub.2
Cartridge, 0-10% MeOH in DCM) to give the title compound 31 as a
cream solid (0.262 g, 70%). LCMS-B: rt 7.13 min; m/z 419
[M+H].sup.+.
Example 32
Synthesis of
1-(2-(2-(2-((6-acetylpyridin-3-yl)amino)-5-chloropyrimidin-4-yl)ethyl)phe-
nyl)cyclopropanecarboxamide (32)
##STR00153##
[0515]
1-(2-(2-(2-((6-Acetylpyridin-3-yl)amino)-5-chloropyrimidin-4-yl)eth-
yl)phenyl)cyclopropanecarboxamide (32)
[0516] A solution of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.200 g, 0.595 mmol) in 1,4-dioxane (2.0 mL) containing
1-(5-aminopyridin-2-yl)ethanone (0.121 g, 0.892 mmol),
Cs.sub.2CO.sub.3 (0.581 g, 1.78 mmol), Xantphos (0.014 g, 0.024
mmol) and palladium (II) acetate (2.67 mg, 0.012 mmol) was heated
under microwave irradiation for 30 minutes at 120.degree. C. The
reaction mixture was adsorbed onto silica gel and purified by
silica column chromatography (Combiflash Rf, 4 g SiO.sub.2
Cartridge, 0-10% MeOH in DCM) to give title compound 32 as a light
cream solid (0.183 g, 70%). LCMS-B: rt 6.98 min; m/z 437
[M+H].sup.+.
Example 33
Synthesis of
1-(2-(2-(2-((6-(1-aminoethyl)pyridin-3-yl)amino)-5-chloropyrimidin-4-yl)e-
thyl)phenyl)cyclopropanecarboxamide (33)
##STR00154##
[0517]
1-(2-(2-(2-((6-(1-Aminoethyl)pyridin-3-yl)amino)-5-chloropyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (33)
[0518] Ammonium acetate (0.587 g, 7.61 mmol) was added to a
solution of
1-(2-(2-(2-((6-Acetylpyridin-3-yl)amino)-5-chloropyrimidin-4-yl)ethyl)phe-
nyl)cyclopropanecarboxamide 32 (0.166 g, 0.381 mmol) in MeOH (5 mL)
and THF (5 mL) and the mixture was stirred for 20 minutes under a
nitrogen atmosphere. Sodium cyanoborohydride (0.017 g, 0.27 mmol)
was added and the mixture was stirred for 5 hours. Additional
sodium cyanoborohydride (0.017 g, 0.27 mmol) was added and the
mixture was stirred for 22 hours at 35.degree. C. Water (10 mL) was
added to the mixture which was then acidified with 20% aqueous
hydrochloride (10 mL). The aqueous phase was washed with Et.sub.2O
(2.times.30 mL) and then basified with solid potassium hydroxide to
pH 10. The aqueous phase was extracted with DCM (3.times.30 mL) and
the combined organics were washed with brine, dried
(Na.sub.2SO.sub.4), adsorbed onto silica and purified by silica
column chromatography (Combiflash Rf, 12 g SiO.sub.2 Cartridge,
0-30% MeOH in DCM) to give the title compound 33 as a white solid
(0.101 g, 60%). LCMS-B: rt 4.32 min; m/z 437 [M+H].sup.+.
Example 34
Synthesis of
1-(2-(2-(5-chloro-2-((1-(difluoromethyl)-1H-pyrazol-4-yl)amino)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide (34)
##STR00155##
[0519]
1-(2-(2-(5-Chloro-2-((1-(difluoromethyl)-1H-pyrazol-4-yl)amino)pyri-
midin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (34)
[0520] A mixture of
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
A14 (0.076 g, 0.225 mmol), 1-(difluoromethyl)-1H-pyrazol-4-amine
A28 (0.030 g, 0.23 mmol) and p-toluenesulfonic acid monohydrate
(0.004 g, 0.023 mmol) in 1,4-dioxane (4.0 mL) was stirred in a
microwave reactor at 120.degree. C. for 2 hours. The volatiles were
evaporated in vacuo and the residue was purified by column
chromatography (Biotage Isolera, 12 g SiO.sub.2 cartridge, 0-5%
MeOH in DCM). Fractions containing suspected product were combined
and the solvent removed in vacuo to give a residue that was
purified further by column chromatography (Biotage Isolera, 12 g
SiO.sub.2 cartridge, 0-60% EtOAc in petroleum benzine 40-60.degree.
C.). Fractions containing suspected product were combined, the
solvent was removed in vacuo and the resulting solid purified by
prep-LCMS to give the title compound 34 as a white solid (0.005 g,
5%). LCMS-B: rt 3.41 min; m/z 433 [M+H].sup.+.
Example 35
Synthesis of
2-(2-(2-(2-(1H-pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phenyl)
propanamide (35)
##STR00156##
[0521] (a)
2-(2-((2,5-Dichloropyrimidin-4-yl)ethynyl)phenyl)propanamide
(A29)
[0522] To a solution of 2-(2-ethynylphenyl)propanamide K7 (2.5 g,
14 mmol) and 2,4,5-trichloropyrimidine (2.2 mL, 19 mmol) in
1,4-dioxane (27 mL) and Et.sub.3N (8.1 mL, 58 mmol) containing CuI
(0.055 g, 0.29 mmol) was added PdCl.sub.2(PPh.sub.3).sub.2 (0.10 g,
0.14 mmol). The reaction mixture was heated at 60.degree. C. for
2.5 hours under a nitrogen atmosphere. The solvent was removed in
vacuo and the residue was diluted with 20% Et.sub.2O in
cyclohexane. The precipitate was filtered, washed with water and
dried in vacuo to give the title compound A29 as a light brown
solid (4.52 g, 98%). LCMS-C: rt 5.10 min; m/z 320 [M+H].sup.+.
(b) 2-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)propanamide
(A30)
[0523] A solution of
2-(2-((2,5-dichloropyrimidin-4-yl)ethynyl)phenyl)propanamide A29
(1.0 g, 3.1 mmol) in DMF (55 mL) and MeOH (5 mL) was stirred with
platinum(II)oxide (0.21 g, 0.94 mmol) under an atmosphere of
hydrogen for 120 hours at ambient temperature. The reaction mixture
was diluted with EtOAc and filtered through a plug of Celite. The
filter cake was washed with EtOAc and the solvents were removed in
vacuo to give a crude residue that was purified by silica gel
column chromatography (Combiflash Rf, 0-90% EtOAc in cyclohexane),
giving the title compound A30 as an off white solid (0.65 g, 64%).
LCMS-C: rt 4.98 min; m/z 324 [M+H].sup.+.
(C)
2-(2-(2-(2-(1H-Pyrazol-4-ylamino)-5-chloropyrimidin-4-yl)ethyl)phenyl)-
propanamide (35)
[0524] A solution of
2-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)propanamide A30
(0.10 g, 0.31 mmol) and tert-butyl
4-amino-1H-pyrazole-1-carboxylate (0.17 g, 0.93 mmol) was heated in
MeOH:water (10:1, 5 mL) at 90.degree. C. for 18 hours. The solvents
were removed in vacuo and the crude residue was purified by silica
gel column chromatography (Combiflash Rf, 0-15% MeOH in DCM). The
purified fractions were combined, the solvent was removed in vacuo
and the solid was sonicated in cyclohexane and filtered. The
residue was dried in vacuo to give the title compound 35 as a light
yellow solid (0.060 g, 52%). .sup.1H NMR (300 MHz, d.sub.6-DMSO)
.delta. 1.34 (d, J=7.04 Hz, 3H), 2.92-3.11 (m, 4H), 3.86 (q, J=7.04
Hz, 1H), 6.85 (brs, 1H), 7.13-7.25 (m, 4H), 7.36-7.44 (m, 1H),
7.57-87 (m, 2H), 8.37 (s, 1H), 9.63 (s, 1H). LCMS-C: rt 4.72 min;
m/z 371 [M+H].sup.+.
Example 36
Synthesis of
2-(2-(2-(5-chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl) propanamide (36)
##STR00157##
[0525]
2-(2-(2-(5-Chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)e-
thyl)phenyl) propanamide (36)
[0526] A solution of
2-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)propanamide A30
(50 mg, 0.15 mmol), 4-amino-1-methylpyrazole (15 mg, 0.015 mmol)
and p-toluenesulfonic acid (3.0 mg, 0.015 mmol) in 1,4-dioxane (2
mL) was heated at 80.degree. C. under microwave irradiation for 3
hours, then at 120.degree. C. for 3 hours and finally at
140.degree. C. for 2 hours. The mixture was partitioned between
water and EtOAc, the layers were separated and the aqueous phase
was extracted with EtOAc (3 times). The combined organic extracts
were washed with brine, dried (Na.sub.2SO.sub.4) and the solvent
evaporated in vacuo to give the crude product. Purification by
silica gel column chromatography (Combiflash Rf, 0-10% MeOH in DCM)
gave the title compound 36 as a pale solid (0.020 g, 34%). .sup.1H
NMR (300 MHz, CDCl.sub.3) .delta. 1.57 (d, J=7.26 Hz, 3H),
3.04-3.15 (m, 4H), 3.91 (s, 3H), 4.00 (q, J=6.97 Hz, 1H), 5.59
(brs, 2H), 7.19-7.31 (m, 3H), 7.34 (brs, 1H), 7.38-7.41 (m, 1H),
7.49 (s, 1H), 7.71 (s, 1H), 8.25 (s, 1H). LCMS-B: rt 6.09 min; m/z
385 [M+H].sup.+.
Example 37
Synthesis of
2-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)propanamide (37)
##STR00158##
[0527] (a) tert-Butyl
4-(5-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-chloropyrimidin-2-yl)a-
mino)pyridin-2-yl)piperidine-1-carboxylate (A31)
[0528] A suspension of
2-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)propanamide A30
(50.0 mg, 0.154 mmol), Cs.sub.2CO.sub.3 (151 mg, 0.463 mmol) and
tert-butyl 4-(4-aminophenyl)piperidine-1-carboxylate 19 (85.6 mg,
0.308 mmol) in 1,4-dioxane (1.0 mL) was sonicated for 10 minutes,
followed by the addition of Xantphos (3.57 mg, 6.18 .mu.mol) and
palladium(II) acetate (0.692 mg, 3.08 .mu.mol). The reaction was
heated under microwave irradiation for 20 minutes at 120.degree. C.
then filtered through Celite. The filter cake was washed with
EtOAc, the solvent was removed under reduced pressure and the crude
material was purified by silica gel column chromatography
(Combiflash Rf, 0-90% EtOAc in cyclohexane) to afford the title
compound A31 (34 mg, 39%). LCMS-C: rt 6.33 min; m/z 565.2
[M+H].sup.+.
(b)
2-(2-(2-(5-Chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-
-yl)ethyl)phenyl)propanamide (37)
[0529] TFA (0.71 mL) was added to a solution of tert-butyl
4-(5-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-chloropyrimidin-2-yl)a-
mino)pyridin-2-yl)piperidine-1-carboxylate A31 (34 mg, 0.06 mmol)
in DCM (7 mL) and the mixture was stirred at room temperature
overnight. The volatiles were removed in vacuo before the addition
of 2.0 M NaOH solution (2 mL) and EtOAc. The phases were separated
and the aqueous phase was extracted with EtOAc. The combined
organic phases were washed with water, dried over Na.sub.2SO.sub.4
and the solvent removed under reduced pressure to give the title
compound 37 (23 mg, 82%). LCMS-C: rt 6.02 min; m/z 465.3
[M+H].sup.+.
Example 38
Synthesis of
2-(2-(2-(5-chloro-2-(6-(1-methylpiperidin-4-yl)pyridin-3-ylamino)pyrimidi-
n-4-yl)ethyl)phenyl)propanamide (38)
##STR00159##
[0530]
2-(2-(2-(5-Chloro-2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)-
pyrimidin-4-yl)ethyl)phenyl)propanamide (38)
[0531] Formaldehyde (31.2 .mu.L, 0.419 mmol, 37 wt % in H.sub.2O)
was added to a suspension of
2-(2-(2-(5-chloro-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)propanamide 37 (39.0 mg, 0.084 mmol) in MeOH (2.0 mL)
under an atmosphere of nitrogen. Sodium triacetoxyborohydride (178
mg, 0.839 mmol) was added in one portion and the reaction mixture
was stirred at room temperature for 2.5 hours. The volatiles were
removed in vacuo and the residue was diluted with EtOAc (10 mL) and
sat. aq. NaHCO.sub.3 (10 mL). The layers were separated and the
aqueous layer was extracted with EtOAc (2.times.10 mL). The
combined organic layers were washed with water (20 mL) and brine
(15 mL) and dried over Na.sub.2SO.sub.4. The solvent was removed
under reduced pressure and the resulting solid was suspended in DCM
(20 mL) and cyclohexane (30 mL). The solid was filtered to give the
title compound 38 (34 mg, 84%). LCMS-C: rt 4.19 min; m/z 479.2
[M+H].sup.+.
Example 39
Synthesis of
2-(2-(2-(5-chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (39)
##STR00160##
[0532]
2-(2-(2-(5-Chloro-2-(pyridin-3-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (39)
[0533] A mixture of
2-(2-(2-(5-chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl) propanamide A30 (0.080 g, 0.25 mmol), 3-aminopyridine (0.046
g, 0.49 mmol), Xantphos (0.0057 g, 0.010 mmol) and Cs.sub.2CO.sub.3
(0.24 g, 0.74 mmol) in 1,4-dioxane (3 mL) was bubbled with nitrogen
for 10 minutes. Palladium (II) acetate (0.0011 g, 0.0049 mmol) was
added and the mixture was heated at 120.degree. C. under microwave
irradiation for 28 minutes. The volatiles were removed in vacuo and
the residue was purified by silica gel column chromatography
(Combiflash Rf, 0-10% MeOH in DCM) to give the title compound 39 as
a pink solid (0.029 g, 31%). LCMS-C: rt 4.30 min; m/z 382
[M+H].sup.+.
Example 40
Synthesis of
2-(2-(2-(5-chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (40)
##STR00161##
[0534]
2-(2-(2-(5-Chloro-2-(pyrimidin-5-ylamino)pyrimidin-4-yl)ethyl)pheny-
l) propanamide (40)
[0535] A mixture of
2-(2-(2-(5-chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl) propanamide A30 (0.080 g, 0.25 mmol), 5-aminopyrimidine
(0.047 g, 0.49 mmol), Xantphos (0.0057 g, 0.010 mmol) and
Cs.sub.2CO.sub.3 (0.24 g, 0.74 mmol) in 1,4-dioxane (3 mL) was
bubbled with nitrogen for 10 minutes. Palladium (II) acetate
(0.0011 g, 0.0049 mmol) was added and the mixture was heated at
120.degree. C. under microwave irradiation for 20 minutes. The
volatiles were removed in vacuo and the residue was purified by
silica gel column chromatography (Combiflash Rf, 0-10% MeOH in DCM)
to give the title compound 40 as an off white solid (0.030 g, 32%).
LCMS-C: rt 4.85 min; m/z 383 [M+H].sup.+.
Example 41
Synthesis of
2-(2-(2-(5-chloro-2-(6-methylpyridin-3-ylamino)pyrimidin-4-yl)ethyl)pheny-
l) propanamide (41)
##STR00162##
[0536]
2-(2-(2-(5-chloro-2-(6-methylpyridin-3-ylamino)pyrimidin-4-yl)ethyl-
)phenyl) propanamide (41)
[0537] A mixture of
2-(2-(2-(5-chloro-2-(1-methyl-1H-pyrazol-4-ylamino)pyrimidin-4-yl)ethyl)p-
henyl) propanamide WAK-011-06-05 (0.080 g, 0.25 mmol),
5-amino-2-methylpyridine (0.053 g, 0.49 mmol), Xantphos (0.0057 g,
0.010 mmol) and Cs.sub.2CO.sub.3 (0.24 g, 0.74 mmol) in 1,4-dioxane
(3 mL) was bubbled with nitrogen for 10 minutes. Palladium (II)
acetate (0.0011 g, 0.0049 mmol) was added and the mixture was
heated at 120.degree. C. under microwave irradiation for 20
minutes. The volatiles were removed in vacuo and the residue was
purified by silica gel column chromatography (Combiflash Rf, 0-10%
MeOH in DCM) to give the title compound 41 as a light yellow solid
(0.032 g, 33%). LCMS-C: rt 4.24 min; m/z 396 [M+H].sup.+.
Example 42
Synthesis of
2-(2-(2-(5-chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)phenyl)
propanamide (42)
##STR00163##
[0538]
2-(2-(2-(5-Chloro-2-(pyridazin-4-ylamino)pyrimidin-4-yl)ethyl)pheny-
l)propanamide (42)
[0539] A mixture of
2-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)propanamide (A30)
2-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)propanamide (A30
(0.080 g, 0.25 mmol), 4-aminopyridazine (0.046 g, 0.49 mmol),
Xantphos (0.0057 g, 0.010 mmol) and Cs.sub.2CO.sub.3 (0.24 g, 0.74
mmol) in 1,4-dioxane (3 mL) was bubbled with nitrogen for 10
minutes. Palladium (II) acetate (0.0011 g, 0.0049 mmol) was added
and the mixture was heated at 120.degree. C. under microwave
irradiation for 28 minutes. The volatiles were removed in vacuo and
the residue was purified by silica gel column chromatography
(Combiflash Rf, 0-15% MeOH in DCM) to give the title compound 42 as
a light yellow solid (0.020 g, 21%). LCMS-C: rt 4.23 min; m/z 383
[M+H].sup.+.
Example 43
Synthesis of
2-(2-(2-(5-chloro-2-(1-(piperidin-4-yl)-1H-pyrazol-4-ylamino)pyrimidin-4--
yl)ethyl)phenyl)propanamide (43)
##STR00164##
[0540] (a) tert-Butyl
4-(4-(4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-chloropyrimidin-2-ylami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate (A32)
[0541] A stirred solution of
2-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)propanamide (A30)
(0.120 g, 0.370 mmol), tert-butyl
4-(4-amino-1H-pyrazol-1-yl)piperidine-1-carboxylate (0.177 g, 0.666
mmol) in n-butanol (3 mL) and glacial acetic acid (0.004 mL, 0.074
mmol) was stirred at 150.degree. C. in the microwave for 30
minutes. After cooling, the solvent was removed to afford a crude
purple oil which was purified by silica gel column chromatography
(Combiflash Rf, 0-30% MeOH in DCM) to give the title compound A32
as a red oil (0.090 g, 44% yield). LCMS-C: rt 5.44 min; m/z 554
[M+H].sup.+.
(b)
2-(2-(2-(5-Chloro-2-(1-(piperidin-4-yl)-1H-pyrazol-4-ylamino)pyrimidin-
-4-yl)ethyl)phenyl) propanamide (43)
[0542] To a stirred solution of tert-butyl
4-(4-(4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-chloropyrimidin-2-ylami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate A32 (0.090 g, 0.162
mmol) in DCM (5 mL) was added TFA (2.0 mL) and the mixture was
stirred at room temperature for 1.5 hours. The solvent was removed
and the residue was suspended in EtOAc (5 mL) and washed with 1M
HCl. The aqueous layer was basified with 1M NaOH and extracted with
EtOAc (2.times.5 mL). The combined organic layers were dried
(MgSO.sub.4), filtered and concentrated in vacuo to give a crude
purple oil which was purified by flash chromatography (Combiflash
Rf, 0-30% MeOH in DCM) to give the title compound 43 as a red oil
(0.019 g, 26% yield). LCMS-C: rt 4.24 min; m/z 454 [M+H].sup.+.
Example 44
Synthesis of
2-(2-(2-(5-chloro-2-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-ylamino)pyri-
midin-4-yl)ethyl)phenyl)propanamide (44)
##STR00165##
[0543]
2-(2-(2-(5-Chloro-2-(1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-ylamin-
o)pyrimidin-4-yl)ethyl)phenyl)propanamide (44)
[0544] A stirred solution of
1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-amine (0.117 g, 0.648
mmol), 2-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)propanamide
A30 (0.060 g, 0.185 mmol) in MeOH (10 mL) and water (1.0 mL) was
stirred at 70.degree. C. for 16 hours. Additional
1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-amine (0.052 g, 0.29 mmol)
was added and the reaction was stirred for 16 hours at 70.degree.
C. After cooling the solvent was removed to afford an oil which was
purified by HPLC (Gradient: 30-100%, acetonitrile 0.1% formic acid
in water 0.1% formic acid, 12 min) to give the title compound 44 as
a red gum (0.0018 g, 2% yield). LCMS-C: rt 4.28 min; m/z 468
[M+H].sup.+.
Example 45
Synthesis of
2-(2-(2-(5-Chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)-2-methylpropanamide (45)
##STR00166##
[0545] (a) 2-(2-Bromophenyl)-2-methylpropanenitrile (A33)
[0546] A suspension of t-BuOK (0.898 g, 8.00 mmol) in anhydrous THF
(3 mL) and anhydrous NMP (3 mL) was cooled to 0.degree. C. under a
N.sub.2 atmosphere. To this suspension,
2-(2-bromophenyl)acetonitrile (0.392 g, 2.00 mmol) was added and
the mixture was allowed to stir at 0.degree. C. for 10 minutes.
Iodomethane (0.50 mL, 8.0 mmol) was added carefully over a period
of 5 minutes and the resulting mixture was stirred for an
additional 1 hour between 0.degree. C. and 10.degree. C. Excess
t-BuOK was quenched by the addition of sat. aq. NaHCO.sub.3 and the
aqueous phase was extracted with EtOAc (3.times.40 mL). The
combined organics were washed with brine, dried (MgSO.sub.4) and
the solvent removed in vacuo to give a yellow residue. Purification
by column chromatography (Biotage Isolera, 24 g SiO.sub.2
cartridge, 0-15% EtOAc in petroleum benzine 40-60.degree. C.) gave
the title compound A33 as a colourless oil (0.377 g, 84%). LCMS-D:
rt 3.48 min; m/z 197/199 [M-CN].sup.+.
(b) 2-(2-Bromophenyl)-2-methylpropanamide (A34)
[0547] A mixture of 2-(2-bromophenyl)-2-methylpropanenitrile A33
(0.377 g, 1.68 mmol), NaOH (0.135 g, 3.37 mmol) and t-BuOH (4 mL)
was stirred in a sealed vessel at 100.degree. C. for 40 hours. The
reaction mixture was cooled, diluted with DCM and filtered through
Celite. The filtrate solvent was removed in vacuo and the residue
was purified by column chromatography (Biotage Isolera, 24 g
SiO.sub.2 cartridge, 0-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A34 as a colourless
oil (0.192 g, 48%). LCMS-D: rt 3.06 min; m/z 242/244
[M+H].sup.+.
(c) 2-Methyl-2-(2-((triethylsilyl)ethynyl)phenyl)propanamide
(A35)
[0548] A mixture of 2-(2-bromophenyl)-2-methylpropanamide A34
(0.196 g, 0.810 mmol), CuI (0.008 g, 0.040 mmol),
tri-tert-butylphosphonium tetrafluoroborate (0.012 g, 0.040 mmol),
PdCl.sub.2(PPh.sub.3).sub.2 (0.028 g, 0.040 mmol) and
(triethylsilyl)acetylene (0.174 mL, 0.971 mmol) in DMF (5 mL) was
bubbled with N.sub.2 for 10 minutes. Et.sub.3N (5 mL) was added and
the mixture was stirred under nitrogen at 65.degree. C. for 4
hours. The mixture was cooled and the volatiles were removed in
vacuo. The dark brown residue was adsorbed onto silica and purified
by column chromatography (Biotage Isolera, 24 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A35 as a brown solid (0.116 g, 48%).
LCMS-D: rt 3.84 min; m/z 302 [M+H].sup.+.
(d) 2-(2-Ethynylphenyl)-2-methylpropanamide (A36)
[0549] A solution of
2-methyl-2-(2-((triethylsilyl)ethynyl)phenyl)propanamide A35 (0.116
g, 0.385 mmol) in THF (5 mL) was stirred with TBAF (1.0 M in THF,
0.40 mL, 0.40 mmol) at 0.degree. C. for 2 minutes. Sat. aq.
NaHCO.sub.3 (20 mL) was added and the aqueous phase was extracted
with EtOAc (3.times.30 mL). The combined organics were washed with
brine, dried (MgSO.sub.4) and the solvent was removed in vacuo. The
resultant yellow oil was purified by column chromatography (Biotage
Isolera, 24 g SiO.sub.2 cartridge, 20-100% EtOAc in petroleum
benzine 40-60.degree. C.) to give the title compound A36 as a
yellow solid (0.060 g, 83%). LCMS-D: rt 3.02 min; m/z 188
[M+H].sup.+.
(e)
2-(2-((2,5-Dichloropyrimidin-4-yl)ethynyl)phenyl)-2-methylpropanamide
(A37)
[0550] A mixture of 2-(2-ethynylphenyl)-2-methylpropanamide A36
(0.060 g, 0.32 mmol), 2,4,5-trichloropyrimidine (48 .mu.L, 0.42
mmol) and Et.sub.3N (0.18 mL, 1.3 mmol) in 1,4-dioxane (5 mL) was
bubbled with N.sub.2 for 10 minutes. CuI (0.001 g, 0.006 mmol) and
PdCl.sub.2(PPh.sub.3).sub.2 (0.002 g, 0.003 mmol) were added and
the reaction mixture was stirred at room temperature under a
N.sub.2 atmosphere for 5.5 hours. The volatiles were removed in
vacuo and the residue was purified by column chromatography
(Biotage Isolera, 12 g SiO.sub.2 cartridge, 10-100% EtOAc in
petroleum benzine 40-60.degree. C.) to give the title compound A37
as an off-white solid (0.067 g, 63%). LCMS-D: rt 3.33 min; m/z 334
[M+H].sup.+.
(f)
2-(2-(2-(2,5-Dichloropyrimidin-4-yl)ethyl)phenyl)-2-methylpropanamide
(A38)
[0551] A mixture of
2-(2-((2,5-dichloropyrimidin-4-yl)ethynyl)phenyl)-2-methylpropanamide
A37 (0.067 g, 0.20 mmol) and PtO.sub.2 (0.014 g, 0.060 mmol) in
MeOH (3.0 mL) and DMF (3.0 mL) was stirred in an atmosphere of
H.sub.2 for 64 hours. The mixture was diluted with EtOAc, filtered
through Celite and the filtrate solvent was removed in vacuo. The
residue was purified by column chromatography (Biotage Isolera, 12
g SiO.sub.2, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A38 as an off-white solid (0.042 g, 62%).
LCMS-D: rt 3.37 min; m/z 338 [M+H].sup.+.
(g)
2-(2-(2-(5-Chloro-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)-2-methylpropanamide (45)
[0552] A mixture of
2-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)-2-methylpropanamide
A38 (0.042 g, 0.12 mmol), 1-methyl-1H-pyrazol-4-amine (0.022 g,
0.23 mmol), and p-toluenesulfonic acid monohydrate (0.002 g, 0.012
mmol) in 1,4-dioxane (4.0 mL) was heated in the microwave at
120.degree. C. for 3 hours. After cooling, the volatiles were
removed in vacuo and the residue was adsorbed onto silica.
Purification by column chromatography (Biotage Isolera, 12 g
SiO.sub.2, 0-5% MeOH in DCM) gave the title compound 45 as an
off-white solid (0.009 g, 18%). LCMS-D: rt 3.29 min; m/z 399
[M+H].sup.+.
Example 46
Synthesis of
1-(2-(2-(2-((1H-Pyrazol-4-yl)amino)-5-methylpyrimidin-4-yl)ethyl)phenyl)c-
yclopropanecarboxamide (46)
##STR00167##
[0553] (a)
1-(2-((2-Chloro-5-methylpyrimidin-4-yl)ethynyl)phenyl)cycloprop-
anecarboxamide (A39)
[0554] A mixture of 1-(2-ethynylphenyl)cyclopropanecarboxamide K6
(2.29 g, 12.4 mmol), 2,4-dichloro-5-methylpyrimidine (2.62 g, 16.1
mmol), CuI (0.047 g, 0.25 mmol) and PdCl.sub.2(PPh.sub.3).sub.2
(0.087 g, 0.12 mmol) in dioxane (25 mL) was bubbled with N.sub.2
for 10 minutes. Et.sub.3N (10 mL) was added and the mixture was
stirred under nitrogen at 70.degree. C. for 1 hour. The reaction
mixture was cooled and adsorbed onto silica. Purification by column
chromatography (Biotage Isolera, 120 g SiO.sub.2, 0-10% MeOH in
CHCl.sub.3) gave the title compound A39 as a yellow oil (1.85 g,
48%). LCMS-D: rt 3.24 min; m/z 312 [M+H].sup.+.
(b)
1-(2-(2-(2-Chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropanecarb-
oxamide (A40)
[0555] A mixture of
1-(2-((2-chloro-5-methylpyrimidin-4-yl)ethynyl)phenyl)cyclopropanecarboxa-
mide A39 (1.85 g, 5.93 mmol) and PtO.sub.2 (0.404 g, 1.78 mmol) in
MeOH (10 mL) and DMF (40 mL) was stirred under an atmosphere of
H.sub.2 for 110 hours at room temperature. The mixture was filtered
through Celite and the volatiles were removed in vacuo. The residue
was purified by column chromatography (Biotage Isolera, 40 g
SiO.sub.2, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A40 as a white solid (1.65 g, 88%). LCMS-D:
rt 3.20 min; m/z 316 [M+H].sup.+.
(c)
1-(2-(2-(2-((1H-Pyrazol-4-yl)amino)-5-methylpyrimidin-4-yl)ethyl)pheny-
l)cyclopropanecarboxamide (46)
[0556] A mixture of
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxa-
mide A40 (0.147 g, 0.465 mmol), tert-butyl
4-amino-1H-pyrazole-1-carboxylate (0.102 g, 0.559 mmol),
Cs.sub.2CO.sub.3 (0.455 g, 1.40 mmol), Xantphos (0.011 g, 0.019
mmol) and Pd(OAc).sub.2 (0.002 g, 0.009 mmol) in 1,4-dioxane (5 mL)
was bubbled with N.sub.2 for 10 minutes before being heated in the
microwave for 20 minutes at 120.degree. C. The reaction mixture was
cooled to room temperature, adsorbed onto silica, and purified by
column chromatography (Biotage Isolera, 24 g SiO.sub.2 cartridge,
20-100% EtOAc in petroleum benzine 40-60.degree. C., then 0-20%
MeOH in EtOAc) to give a white solid. This solid was taken up in
DCM and precipitated by the addition of cyclohexane. The suspension
was sonicated for 10 minutes and the precipitate was isolated by
vacuum filtration and washed with Et.sub.2O to give the title
compound 46 as a white solid (0.021 g, 12%). LCMS-D: rt 2.98 min;
m/z 363 [M+H].sup.+.
Example 47
Synthesis of
1-(2-(2-(5-methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)cyclopropanecarboxamide (47)
##STR00168##
[0557] (a) Methyl
1-(2-((2-chloro-5-methylpyrimidin-4-yl)ethynyl)phenyl)cyclopropanecarboxy-
late (A41)
[0558] A mixture of methyl
1-(2-ethynylphenyl)cyclopropanecarboxylate K2 (1.34 g, 6.67 mmol),
2,4,-dichloro-5-methylpyrimidine (1.41 g, 8.67 mmol), CuI (0.063 g,
0.33 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.234 g, 0.333 mmol) and
tri-t-butylphosphonium tetrafluoroborate (0.097 g, 0.33 mmol) in
1,4-dioxane (15 mL) was bubbled with N.sub.2 for 10 minutes. DIPEA
(3.48 mL, 20.0 mmol) was added and the reaction mixture was stirred
under N.sub.2 at 80.degree. C. for 3 hours. The reaction mixture
was cooled and the volatiles were removed in vacuo to give a black
residue that was purified by column chromatography (Biotage
Isolera, 2.times.40 g SiO.sub.2 cartridges, 0-50% EtOAc in
petroleum benzine 40-60.degree. C.) to give the impure title
compound A41 as a yellow oil (estimated 80% purity, 1.352 g, 50%).
LCMS-A: rt 6.79 min; m/z 327 [M+H].sup.+.
(b) Methyl
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cycloprop-
anecarboxylate (A42)
[0559] A solution of methyl
1-(2-((2-chloro-5-methylpyrimidin-4-yl)ethynyl)phenyl)cyclopropanecarboxy-
late A41 (estimated 80% purity, 1.35 g, 3.31 mmol) in DMF (15 mL)
and MeOH (5 mL) was stirred with PtO.sub.2 (0.235 g, 1.03 mmol)
under an atmosphere of H.sub.2 for 16 hours at room temperature.
The reaction mixture was diluted with EtOAc (50 mL) and filtered
through Celite. The volatiles were removed in vacuo and the black
residue was purified by column chromatography (Biotage Isolera, 40
g SiO.sub.2 cartridge, 0-25% EtOAc in petroleum benzine
40-60.degree. C. then 40 g SiO.sub.2 cartridge, 0-5% MeOH in DCM)
to give the impure title compound A42 (estimated 78% purity, 0.922
g, 66%). LCMS-D: rt 3.58 min; m/z 331 [M+H].sup.+.
(c)
1-(2-(2-(5-Methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)cyclopropanecarboxamide (47)
[0560] A mixture of 1-methyl-1H-pyrazol-4-amine (0.294 g, 3.02
mmol), methyl
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropane-
carboxylate A42 (0.500 g, estimated 78% purity, 1.18 mmol),
Xantphos (0.035 g, 0.060 mmol) and Cs.sub.2CO.sub.3 (1.48 g, 4.53
mmol) in 1,4-dioxane (5 mL) was bubbled with N.sub.2 for 10
minutes. Palladium (II) acetate (0.007 g, 0.030 mmol) was added and
the mixture was stirred in the microwave at 120.degree. C. for 20
minutes. The volatiles were removed in vacuo and the black residue
was purified by chromatography (Biotage Isolera, 2.times.40 g
SiO.sub.2 cartridges, 50-100% EtOAc in petroleum benzine
40-60.degree. C. and then 0-10% MeOH in EtOAc) to give a yellow
oil. This oil was purified further by column chromatography (0-5%
MeOH in DCM) to give a yellow solid. A mixture of this solid and
LiOH.H.sub.2O (0.234 g, 5.57 mmol) in THF (7 mL), MeOH (7 mL) and
H.sub.2O (1.5 mL) was stirred at room temperature for 24 hours.
Another portion of LiOH.H.sub.2O (0.234 g, 5.57 mmol) was added to
the reaction mixture and stirring was continued for 72 hours at
room temperature and then 24 hours at 35.degree. C. The volatiles
were removed in vacuo before H.sub.2O (10 mL) and aqueous HCl (2 M,
10 mL) were added. The aqueous phase was extracted with EtOAc
(3.times.30 mL), the organics were combined, washed with brine,
dried (MgSO.sub.4) and the solvent removed under reduced pressure
to give a yellow oil. A mixture of this oil, HOBt (0.072 g, 0.535
mmol), EDCl.HCl (0.103 g, 0.535 mmol) and DIPEA (0.23 mL, 1.34
mmol) in THF (6 mL) and DMF (1 mL) was stirred at room temperature
for 10 minutes before ammonium carbonate (0.257 g, 2.68 mmol) was
added. The mixture was stirred for a further 24 hours at room
temperature and then 24 hours at 35.degree. C. Sat. aqueous
NaHCO.sub.3 (30 mL) was added and the mixture was extracted with
EtOAc (3.times.30 mL). The organics were combined, washed with
brine and the solvent was removed in vacuo to give a yellow oil
that was purified by column chromatography (Biotage Isolera, 40 g
SiO.sub.2, 50-100% EtOAc in petroleum benzine 40-60.degree. C. and
then 0-10% MeOH in EtOAc). The purified fractions were combined and
the solvent removed in vacuo to give a yellow solid. Further
purification by column chromatography (Biotage Isolera, 40 g
SiO.sub.2 cartridge, 0-5% MeOH in CHCl.sub.3) gave a yellow solid.
Prep-LCMS of this material gave the title compound 47 as a white
solid (0.005 g, 1.1%). LCMS-D: rt 3.03 min; m/z 377
[M+H].sup.+.
Example 48
Synthesis of
1-(2-(2-(5-methyl-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)py-
rimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (48)
##STR00169##
[0561] (a) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate (A43)
[0562] A mixture of
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxa-
mide A40 (0.250 g, 0.792 mmol), tert-butyl
4-(4-amino-1H-pyrazol-1-yl)piperidine-1-carboxylate (0.422 g, 1.58
mmol), Pd(OAc).sub.2 (0.004 g, 0.016 mmol), Xantphos (0.018 g,
0.032 mmol) and Cs.sub.2CO.sub.3 (0.774 g, 2.38 mmol) in dioxane
(15 mL) was bubbled with N.sub.2 for 10 minutes and then stirred in
the microwave at 120.degree. C. for 20 minutes. The volatiles were
removed in vacuo and the residue was adsorbed onto SiO.sub.2 and
purified by column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A43 as a brown solid (0.118 g, 27%).
LCMS-D: rt 3.40 min; m/z 546 [M+H].sup.+.
(b)
1-(2-(2-(5-Methyl-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrimid-
in-4-yl)ethyl)phenyl)cyclopropanecarboxamide (A44)
[0563] A solution of tert-butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)ami-
no)-1H-pyrazol-1-yl)piperidine-1-carboxylate A43 (0.113 g, 0.207
mmol) in DCM (10 mL) was treated with TFA (0.32 mL, 4.1 mmol) and
stirred for 20 hours at room temperature. The volatiles were
evaporated in vacuo and aq. NaOH (2 M, 40 mL) was added to the
residue. The aqueous phase was extracted with EtOAc (3.times.40 mL)
and the combined organics were washed with brine and dried over
MgSO.sub.4. The solvent was removed in vacuo and the solid was
suspended in DCM. Cyclohexane was added and the suspension was
sonicated for 10 minutes. The solid was isolated by vacuum
filtration to give the title compound A44 as an off-white solid
(0.066 g, 72%). LCMS-D: rt 2.88 min; m/z 446 [M+H].sup.+.
(c)
1-(2-(2-(5-Methyl-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (48)
[0564] To a solution of
1-(2-(2-(5-methyl-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrimidin--
4-yl)ethyl)phenyl)cyclopropanecarboxamide A44 (0.064 g, 0.14 mmol)
in MeOH (12 mL) was added formaldehyde solution (37% in water, 32
.mu.L, 0.43 mmol) and sodium triacetoxyborohydride (0.122 g, 0.575
mmol) under a nitrogen atmosphere. The mixture was stirred at room
temperature for 4 hours before concentrating under reduced
pressure. The mixture was diluted with sat. aq. NaHCO.sub.3 (20 mL)
and the aqueous phase was extracted with EtOAc (3.times.25 mL). The
combined organics were washed with brine, and dried (MgSO.sub.4)
before the solvent was removed in vacuo to give the title compound
48 as a white solid (0.035 g, 53%). LCMS-D: rt 2.89 min; m/z 460
[M+H].sup.+.
Example 49
Synthesis of
1-(2-(2-(5-Methyl-2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)pyrimi-
din-4-yl)ethyl)phenyl)cyclopropanecarboxamide (49)
##STR00170##
[0565] (a) tert-Butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)ami-
no)pyridin-2-yl)piperidine-1-carboxylate (A45)
[0566] A mixture of
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxa-
mide A40 (0.240 g, 0.761 mmol), tert-butyl
4-(5-aminopyridin-2-yl)piperidine-1-carboxylate (0.422 g, 1.52
mmol), Pd(OAc).sub.2 (0.003 g, 0.015 mmol), Xantphos (0.018 g,
0.030 mmol) and Cs.sub.2CO.sub.3 (0.744 g, 2.28 mmol) in dioxane (8
mL) was bubbled with N.sub.2 for 10 minutes and then stirred in the
microwave at 120.degree. C. for 20 minutes. The volatiles were
removed in vacuo and the residue was adsorbed onto SiO.sub.2 and
purified by column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C. then
0-20% MeOH in EtOAc) to give the title compound A45 as a white
solid (0.212 g, 50%). LCMS-D: rt 3.20 min; m/z 557 [M+H].sup.+.
(b)
1-(2-(2-(5-Methyl-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-
-yl)ethyl)phenyl)cyclopropanecarboxamide (A46)
[0567] A solution of tert-butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)ami-
no)pyridin-2-yl)piperidine-1-carboxylate A45 (0.200 g, 0.359 mmol)
in DCM (10 mL) was treated with TFA (0.41 mL, 5.4 mmol) and stirred
at room temperature for 4 hours. An extra aliquot of TFA (0.14 mL,
1.8 mmol) was added to the reaction mixture and stirring was
continued for 1 hour at room temperature. Sat. aq. NaHCO.sub.3
(.about.20 mL) was carefully added to the mixture followed by aq.
NaOH (2 M, .about.20 mL). DCM was removed in vacuo and the aqueous
mixture was extracted with EtOAc (3.times.30 mL). The organic
extracts were combined, washed with brine, dried (MgSO.sub.4) and
the solvent removed in vacuo. The residue was dissolved in DCM
before cyclohexane was added to form a white precipitate. The
suspension was sonicated for 10 minutes and filtered to give the
title compound A46 as a white solid (0.129 g, 79%). LCMS-D: rt 3.08
min; m/z 457 [M+H].sup.+.
(c)
1-(2-(2-(5-Methyl-2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (49)
[0568] A solution of
1-(2-(2-(5-methyl-2-((6-(piperidin-4-yl)pyridin-3-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)cyclopropanecarboxamide A46 (0.121 g, 0.265 mmol) in
MeOH (10 mL) was treated with formaldehyde solution (37% in water,
59 .mu.L, 0.80 mmol) and sodium triacetoxyborohydride (0.225 g,
1.06 mmol) and stirred at room temperature for 3 hours. The mixture
was concentrated in vacuo, aq. NaOH (2 M, .about.30 mL) was added
and the aqueous phase was extracted with EtOAc (3.times.30 mL). The
organics were combined, washed with brine, dried (MgSO.sub.4) and
the solvent evaporated under reduced pressure to give a colourless
oil. The oil was taken up in DCM and cyclohexane was added until a
white precipitate formed. The suspension was sonicated for 10
minutes and the solid was isolated by vacuum filtration and dried
under high vacuum for 4 hours to give the title compound 49 as a
white solid (0.051 g, 41%). LCMS-D: rt 3.13 min; m/z 471
[M+H].sup.+.
Example 50
Synthesis of
1-(2-(2-(2-((4-(1-Aminoethyl)phenyl)amino)-5-methylpyrimidin-4-yl)ethyl)p-
henyl)cyclopropanecarboxamide (50)
##STR00171##
[0569] (a) tert-Butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate (A47)
[0570] A mixture of
1-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxa-
mide A40 (0.300 g, 0.950 mmol), tert-butyl
(1-(4-aminophenyl)ethyl)carbamate (0.449 g, 1.90 mmol),
Pd(OAc).sub.2 (0.004 g, 0.019 mmol), xantphos (0.022 g, 0.038 mmol)
and Cs.sub.2CO.sub.3 (0.929 g, 2.85 mmol) in dioxane (15 mL) was
bubbled with N.sub.2 for 10 minutes and then stirred in the
microwave at 120.degree. C. for 20 minutes. The volatiles were
removed in vacuo and the residue was adsorbed onto SiO.sub.2 and
purified by column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A47 as a white solid (0.110 g, 22%).
LCMS-D: rt 3.52 min; m/z 516 [M+H].sup.+.
(b)
1-(2-(2-(2-((4-(1-Aminoethyl)phenyl)amino)-5-methylpyrimidin-4-yl)ethy-
l)phenyl)cyclopropanecarboxamide (50)
[0571] A solution of tert-butyl
(1-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-methylpyrimidin-2-yl)am-
ino)phenyl)ethyl)carbamate A47 (0.110 g, 0.213 mmol) in DCM (10 mL)
was treated with TFA (0.33 mL, 4.3 mmol) and stirred for 6 hours at
room temperature. The volatiles were evaporated in vacuo before aq.
NaOH (2 M, 30 mL) was added to the residue. The aqueous phase was
extracted with EtOAc (3.times.30 mL) and the combined organics were
washed with brine and dried over MgSO.sub.4. The solvent was
removed in vacuo to give an off-white solid that still contained
starting material. The solid was taken up in DCM (20 mL) and
treated with TFA (0.17 mL, 2.2 mmol). The mixture was stirred at
room temperature for 24 hours before the volatiles were removed in
vacuo. Aq. NaOH (2 M, 30 mL) was added and the aqueous phase was
extracted with EtOAc (3.times.30 mL). The organics were combined,
washed with brine, dried (MgSO.sub.4) and the solvent removed in
vacuo. The resultant white solid was dissolved in DCM and the
desired product was precipitated by the addition of cyclohexane.
The precipitate was isolated by vacuum filtration to give the title
compound 50 (0.037 g, 42%) as a white solid. LCMS-A: rt 2.98 min;
m/z 399 [M-NH.sub.2].sup.+.
Example 51
Synthesis of
2-(2-(2-(5-Methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)propanamide (51)
##STR00172##
[0572] (a)
2-(2-((2-Chloro-5-methylpyrimidin-4-yl))ethynyl)phenyl)propanam-
ide (A48)
[0573] A suspension of 2-(2-ethynylphenyl)propanamide K7 (2.00 g,
11.5 mmol), 2,4,-dichloro-5-methylpyrimidine (2.45 g, 15.0 mmol),
CuI (0.044 g, 0.33 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.162 g,
0.231 mmol) and tri-t-butylphosphonium tetrafluoroborate (0.067 g,
0.23 mmol) in DMF (15 mL) was bubbled with N.sub.2 for 10 minutes.
Et.sub.3N (10 mL) was added and the reaction mixture was stirred
under N.sub.2 at 60.degree. C. for 2.5 hours. The mixture was
cooled, diluted with Et.sub.2O and the precipitate was isolated by
filtration and washed with Et.sub.2O then water to give the title
compound A48 as a white solid (2.08 g, 60%). LCMS-A: rt 5.86 min;
m/z 300 [M+H].sup.+.
(b)
2-(2-(2-(2-Chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)propanamide
(A49)
[0574] A solution of
2-(2-((2-chloro-5-methylpyrimidin-4-yl)ethynyl)phenyl)propanamide
A48 (2.08 g, 6.95 mmol) in DMF (110 mL) and MeOH (10 mL) was
stirred with PtO.sub.2 (0.395 g, 1.74 mmol) under an atmosphere of
H.sub.2 for 16 hours at room temperature. An extra portion of
PtO.sub.2 (0.189 g, 0.832 mmol) was added and stirring was
continued under H.sub.2 for 72 hours at room temperature. The
mixture was diluted with EtOAc (200 mL) and filtered through
Celite. The volatiles were removed in vacuo and the residue was
purified by column chromatography (Biotage Isolera, 120 g SiO.sub.2
cartridge, 10-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A49 as a white solid (1.51 g, 72%). LCMS-D:
rt 3.15 min; m/z 304 [M+H].sup.+.
(c)
2-(2-(2-(5-Methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)et-
hyl)phenyl)propanamide (51)
[0575] A mixture of
2-(2-(2-(2-chloro-5-methylpyrimidin-4-yl)ethyl)phenyl)propanamide
A49 (0.250 g, 0.823 mmol), 1-methyl-1H-pyrazol-4-amine (0.096 g,
0.99 mmol), and p-toluenesulfonic acid monohydrate (0.016 g, 0.082
mmol) in 1,4-dioxane (5 mL) was irradiated in the microwave at
120.degree. C. for 3 hours. The volatiles were removed in vacuo and
the residue was adsorbed onto silica and purified by column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 50-100%
EtOAc in petroleum benzine 40-60.degree. C., then 0-10% MeOH in
EtOAc). The fractions containing suspected product were combined
and the solvent removed in vacuo to give a green solid. The solid
was adsorbed onto silica and purified by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-10% MeOH in
CHCl.sub.3) to give the title compound 51 as a white solid (0.178
g, 59%); .sup.1H NMR (400 MHz, d.sub.6-DMSO) .delta. ppm 9.16 (s,
1H), 8.11 (s, 1H), 7.85 (s, 1H), 7.46 (s, 1H), 7.41-7.35 (m, 1H),
7.24 (s, 1H), 7.20-7.10 (m, 3H), 6.87 (s, 1H), 3.86 (q, J=7.1, 7.0,
7.0 Hz, 1H), 3.78 (s, 3H), 3.23-3.11 (m, 1H), 3.08-2.97 (m, 1H),
2.96-2.78 (m, 2H), 2.05 (s, 3H), 1.33 (d, J=6.9 Hz, 3H). LCMS-D: rt
2.99 min; m/z 365 [M+H].sup.+.
Example 51-1A and 51-2A
Separation of
2-(2-(2-(5-Methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)propanamide (51)
##STR00173##
[0577]
2-(2-(2-(5-Methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl-
)ethyl)phenyl)propanamide (51) was separated at using the chiral
separation method A.
[0578] Enantiomer A of
2-(2-(2-(5-methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)propanamide (51-1A), Chiral Characterisation Method E: rt
12.11 min, enantiomeric purity >99%.
[0579] Enantiomer B of
2-(2-(2-(5-methyl-2-((1-methyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)ethyl-
)phenyl)propanamide (51-2A), Chiral Characterisation Method E: rt
14.72 min, enantiomeric purity >99%.
Example 52
Synthesis of
1-(2-(2-(2-((4-(piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclobutanecarboxamide (52)
##STR00174## ##STR00175##
[0580] (a) tert-Butyl
4-(4-((5-(trifluoromethyl)-4-((triethylsilyl)ethynyl)pyrimidin-2-yl)amino-
)phenyl) piperidine-1-carboxylate (A50)
[0581] A suspension of tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)piperidine-
-1-carboxylate K3 (5.10 g, 11.2 mmol), PdCl.sub.2(PPh.sub.3).sub.2
(392 mg, 0.558 mmol), PPh.sub.3 (146 mg, 0.558 mmol) and CuI (106
mg, 0.558 mmol) in THF (200 mL) and Et.sub.3N (7.78 mL, 55.8 mmol)
was sonicated for 10 minutes. TES acetylene (3.00 mL, 16.7 mmol)
was added and the mixture was stirred at 30.degree. C. overnight.
The crude mixture was adsorbed onto silica gel and purified by
silica gel column chromatography (Biotage Isolera, 0-50% EtOAc in
petroleum benzine 40-60.degree. C.) to give the title compound A50
as a yellow foam (4.64 g, 74%). LCMS-A: rt 7.737 min; m/z 561
[M+H].sup.+.
(b) tert-Butyl
4-(4-((4-ethynyl-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)piperidin-
e-1-carboxylate (A51)
[0582] A mixture of K.sub.2CO.sub.3 (1.72 g, 12.4 mmol) and
tert-butyl
4-(4-((5-(trifluoromethyl)-4-((triethylsilyl)ethynyl)pyrimidin-2-yl)amino-
)phenyl) piperidine-1-carboxylate A50 (4.64 g, 8.28 mmol) in MeOH
(250 mL) was stirred for 15 minutes at room temperature. The
mixture was diluted with water (250 mL) and extracted with DCM
(3.times.250 mL). The organics were combined and adsorbed onto
silica gel. Purification by silica gel column chromatography
(Biotage Isolera, 0-100% EtOAc in petroleum benzine 40-60.degree.
C.) gave the title compound A51 as a yellow solid (2.196 g, 59%).
LCMS-A: rt 6.615 min; m/z 445 [M-H].sup.-.
(c) 1-(2-Iodophenyl)cyclobutanecarboxylic acid (A52)
[0583] A solution of 1-phenyl-1-cyclobutanecarboxylic acid (2.00 g,
11.4 mmol) in DMF (10 mL) containing palladium (II) acetate (0.109
g, 0.567 mmol), iodine (2.16 g, 8.51 mmol) and
(diacetoxyiodo)benzene (2.47 g, 8.51 mmol) was stirred at
60.degree. C. for 18 hours in the absence of light. Additional
iodine (2.16 g, 8.51 mmol) and (diacetoxyiodo)benzene (2.47 g, 8.51
mmol) were added and stirring was continued at 60.degree. C. for 8
hours. A final addition of iodine (2.16 g, 8.51 mmol) and
(diacetoxyiodo)benzene (2.47 g, 8.51 mmol) was performed and
stirring was continued at 60.degree. C. for 16 hours. The reaction
mixture was partitioned between EtOAc and water and the aqueous
phase was extracted several times with EtOAc. The combined organic
fractions were washed with 10% sodium metabisulfate (3.times.30
mL), 10% citric acid (2.times.30 mL), water, brine, and then dried
(MgSO.sub.4), filtered and the solvent evaporated. The crude
product was dry loaded onto silica gel and the product was
separated using silica column chromatography (Biotage Isolera, 40 g
SiO.sub.2 Cartridge, 0-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A52 as a cream solid
(425 mg, 12%). LCMS-A: rt 5.711 min.
(d) Methyl 1-(2-iodophenyl)cyclobutanecarboxylate (A53)
[0584] A solution of 1-(2-iodophenyl)cyclobutanecarboxylic acid A52
(425 mg, 1.41 mmol) in MeOH (25 mL) and conc. H.sub.2SO.sub.4 (1.0
mL) was stirred at 60.degree. C. overnight. The volatiles were
removed in vacuo and the resulting residue diluted in EtOAc (50 mL)
and sat. soln. Na.sub.2CO.sub.3 (50 mL). The aqueous layer was
extracted with EtOAc (3.times.50 mL), the combined organic
fractions were dried (MgSO.sub.4) and the volatiles removed in
vacuo to give the title compound A53 as a clear oil (361 mg, 81%).
LCMS-A: rt 7.020 min; m/z 317 [M+H].sup.+.
(e) tert-Butyl
4-(4-((4-((2-(1-(methoxycarbonyl)cyclobutyl)phenyl)ethynyl)-5-(trifluorom-
ethyl)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate
(A54)
[0585] A suspension of tert-butyl
4-(4-((4-ethynyl-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)piperidin-
e-1-carboxylate A51 (169 mg, 0.380 mmol), methyl
1-(2-iodophenyl)cyclobutanecarboxylate A53 (100 mg, 0.316 mmol),
PPh.sub.3 (8 mg, 0.03 mmol) and CuI (3 mg, 0.02 mmol) in DMF (3 mL)
and Et.sub.3N (0.5 mL) was sonicated for 10 minutes before
PdCl.sub.2(PPh.sub.3).sub.2 (22 mg, 0.032 mmol) was added. The
reaction mixture was irradiated in the microwave at 120.degree. C.
for 20 minutes, adsorbed onto silica gel and purified using column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-30%
EtOAc in petroleum benzine 40-60.degree. C. (column run twice)) to
give the title compound A54 as a yellow oil (33 mg, 16%). LCMS-A:
rt 7.919 min; m/z 635 [M+H].sup.+.
(f) tert-Butyl
4-(4-((4-(2-(1-(methoxycarbonyl)cyclobutyl)phenethyl)-5-(trifluoromethyl)-
pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate (A55)
[0586] Pd/C 10% (40 mg) was added to a solution of tert-butyl
4-(4-((4-((2-(1-(methoxycarbonyl)cyclobutyl)phenyl)ethynyl)-5-(trifluorom-
ethyl)pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate A54 (33
mg, 0.052 mmol) in EtOAc (10 mL) and Et.sub.3N (0.5 mL). The
mixture was stirred under a hydrogen atmosphere overnight and then
filtered through Celite. The filter cake was washed with EtOAc (50
mL) and the volatiles were removed in vacuo to give the title
compound A55 as a yellow oil (31 mg, 93%). LCMS-A: rt 8.013 min;
m/z 639.3 [M+H].sup.+.
(g)
1-(2-(2-(2-((4-(1-(tert-Butoxycarbonyl)piperidin-4-yl)phenyl)amino)-5--
(trifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclobutanecarboxylic
acid (A56)
[0587] LiOH.H.sub.2O (41 mg, 0.97 mmol) was added to a solution of
tert-butyl
4-(4-((4-(2-(1-(methoxycarbonyl)cyclobutyl)phenethyl)-5-(trifluoromethyl)-
pyrimidin-2-yl)amino)phenyl)piperidine-1-carboxylate A55 (31 mg,
0.049 mmol) in THF (5 mL) and H.sub.2O (0.5 mL) and the mixture was
stirred at 40.degree. C. overnight. Additional LiOH.H.sub.2O (400
mg, 9.53 mmol) was added and the mixture was heated at reflux for
12 days. Upon cooling DCM (50 mL) and water (50 mL) were added and
the aqueous layer was extracted with EtOAc (2.times.50 mL). The
combined organics were dried (Na.sub.2SO.sub.4) and the volatiles
were removed in vacuo to give the title compound A56 as a yellow
oil (30 mg, 98%). LCMS-A: rt 7.586 min, m/z 625 [M+H].sup.+.
(h) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclobutyl)phenethyl)-5-(trifluoromethyl)pyrimidi-
n-2-yl)amino)phenyl)piperidine-1-carboxylate (A57)
[0588] Ammonium carbonate (92 mg 0.96 mmol) was added to a
suspension of
1-(2-(2-(2-((4-(1-(tert-butoxycarbonyl)piperidin-4-yl)phenyl)amino)-5-(tr-
ifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclobutanecarboxylic
acid A56 (30 mg, 0.048 mmol), HOBt (32 mg, 0.24 mmol) and EDCl.HCl
(46 mg, 0.24 mmol) in Et.sub.3N (0.2 mL) and DMF (10 mL) and the
resulting mixture was stirred at 40.degree. C. overnight. The
volatiles were removed in vacuo and the resulting residue was
dissolved in DCM (50 mL), washed with water (50 mL), separated and
adsorbed onto silica gel. Purification by column chromatography
(Biotage Isolera, 12 g SiO.sub.2 cartridge, 0-100% EtOAc in
peteroleum benzine 40-60.degree. C.) gave the title compound A57 as
a yellow solid (6 mg, 20%). LCMS-A: rt 8.604 min; m/z 624.3
[M+H].sup.+.
(i)
1-(2-(2-(2-((4-(Piperidin-4-yl)phenyl)amino)-5-(trifluoromethyl)pyrimi-
din-4-yl)ethyl)phenyl)cyclobutanecarboxamide (52)
[0589] A mixture of TFA (0.5 mL) and tert-butyl
4-(4-((4-(2-(1-carbamoylcyclobutyl)phenethyl)-5-(trifluoromethyl)pyrimidi-
n-2-yl)amino)phenyl)piperidine-1-carboxylate A57 (6.0 mg, 9.6
.mu.L) in DCM (5 mL) was stirred at room temperature overnight. The
volatiles were removed in vacuo before 2 M aq. NaOH (5 mL) was
added to form a precipitate. The solid was collected by filtration
and washed with cyclohexane (10 mL) to give the title compound 52
as a tan solid (1.5 mg, 30%). LCMS-A: rt 4.994 min; m/z 524.3
[M+H].sup.+.
Example 53
Synthesis of
1-(3-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (53)
##STR00176##
[0590] (a) 1-(3-Bromophenyl)cyclopropanecarboxamide (A58)
[0591] Ammonium carbonate (7.97 g 83.0 mmol) was added to a
suspension of 1-(3-bromophenyl)cyclopropylacetic acid (2.00 g, 8.30
mmol), HOBt (1.68 g, 12.4 mmol) and EDCl.HCl (2.39 g, 12.4 mmol) in
Et.sub.3N (5 mL) and DMF (25 mL) and the mixture was stirred at
room temperature overnight. EtOAc (150 mL) was added and the
organic layer was washed with sat. NaHCO.sub.3 solution (100 mL),
0.5 M citric acid solution (3.times.100 mL), water (100 mL), brine
(50 mL) and dried (Na.sub.2SO.sub.4). The volatiles were removed in
vacuo to yield the title compound A58 as a white solid (1.78 g,
89%). LCMS-A: rt 5.807 min; m/z 241 [M+H].sup.+.
(b) 1-(3-((Triisopropylsilyl)ethynyl)phenyl)cyclopropanecarboxamide
(A59)
[0592] A suspension of 1-(3-bromophenyl)cyclopropanecarboxamide A58
(1.00 g, 4.17 mmol), TIPS acetylene (3.74 mL, 16.7 mmol), PPh.sub.3
(55 mg, 0.21 mmol) and CuI (40 mg, 0.21 mmol) in Et.sub.3N (4 mL)
and DMF (15 mL) was sonicated for 10 minutes.
PdCl.sub.2(PPh.sub.3).sub.2 (146 mg, 0.208 mmol) was added and the
mixture was heated to 80.degree. C. for 16 hours. The crude mixture
was adsorbed onto silica gel and purified by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100% EtOAc in
petroleum benzine 40-60.degree. C.) to give the title compound A59
as a white solid (1.20 g, 84%). LCMS-A: rt 8.081 min; m/z 342.4
[M+H].sup.+.
(c) 1-(3-Ethynylphenyl)cyclopropanecarboxamide (A60)
[0593] A 1.0 M TBAF solution in THF (10.5 mL, 10.5 mmol) was added
to a solution of
1-(3-((triisopropylsilyl)ethynyl)phenyl)cyclopropanecarboxamide A59
(1.20 g, 3.50 mmol) in THF (50 mL) and stirred for 15 minutes. The
mixture was diluted with water (200 mL) and EtOAc (200 mL) before
the organic fraction was separated and washed with 0.5 M citric
acid solution (200 mL), water (200 mL), brine (50 mL) and dried
(Na.sub.2SO.sub.4). The volatiles were removed in vacuo and the
resulting solid was suspended in cyclohexane (50 mL) and sonicated
for 10 minutes. The solid was collected by filtration and dried to
give the title compound A60 as a tan solid (386 mg, 59%). LCMS-A:
rt 5.635 min; m/z 186.2 [M+H].sup.+.
(d)
1-(3-(2-(2-((1-(Piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromet-
hyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (53)
[0594] A suspension of tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)-
piperidine-1-carboxylate K8 (483 mg, 1.08 mmol),
1-(3-ethynylphenyl)cyclopropanecarboxamide A60 (200 mg, 1.08 mmol),
PPh.sub.3 (14 mg, 0.054 mmol) and CuI (10 mg, 0.054 mmol) in
Et.sub.3N (1 mL) and DMF (4 mL) was sonicated for 10 minutes,
PdCl.sub.2(PPh.sub.3).sub.2 (38 mg, 0.054 mmol) was added and the
reaction mixture was irradiated in the microwave at 120.degree. C.
for 40 minutes. The mixture was adsorbed onto silica gel and
purified by column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give a yellow oil. Further purification by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100% MeOH in EtOAc)
gave a yellow oil which was taken up in EtOAc (20 mL) and DMF (2
mL). Pd/C 10% (100 mg) was added to this solution and the mixture
was stirred overnight under an atmosphere of hydrogen at room
temperature. The mixture was filtered through Celite and the filter
cake was washed with EtOAc (50 mL). The washings were combined and
the volatiles were removed in vacuo to give a yellow oil that was
adsorbed onto silica gel and purified by silica gel column
chromatography (Biotope Isolera, 40 g SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C. then 0-20% MeOH in
EtOAc). Further purification by silica column chromatography
(Biotage Isolera, 25 g SiO.sub.2 cartridge, 0-100% EtOAc in
petroleum benzine 40-60.degree. C. then 0-20% MeOH in EtOAc) gave a
crude material that was dissolved in DCM (20 mL). TFA (1 mL) was
added to this solution and the mixture was stirred at room
temperature overnight. The volatiles were removed in vacuo and the
resulting solid was sonicated for 10 minutes in sat.
Na.sub.2CO.sub.3 (aq.) (25 mL). The precipitate was collected by
filtration and washed with water (25 mL), toluene (50 mL) and air
dried to give the title compound 53 as a yellow solid (15 mg, 32%).
LCMS-A: rt 4.784 min; m/z 500.3 [M+H].sup.+.
Example 54
Synthesis of
2-(2-(2-(2-((1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)eth-
yl)phenyl)propanamide (54)
##STR00177##
[0595] (a) tert-Butyl
4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyrazole-1-carbo-
xylate (A61)
[0596] A 1.0 M solution of zinc chloride in Et.sub.2O (4.0 mL, 4.0
mmol) was added to 2,4-dichloro-5-trifluoromethylpyrimidine (0.49
mL, 3.6 mmol) in a 1:1 mixture of DCE/t-BuOH (40 mL) at 0.degree.
C. under a nitrogen atmosphere. The mixture was stirred for 1 hour
before tert-butyl 4-amino-1H-pyrazole-1-carboxylate (0.606 g, 3.31
mmol) and triethylamine (0.51 mL, 3.7 mmol) in a 1:1 mixture of
DCE/t-BuOH (30 mL) was added. The mixture was allowed to warm to
room temperature and stirred for 20 hours before concentrating in
vacuo and adding gradually to 100 mL water. The resulting
precipitate was removed by vacuum filtration and washed with DCM.
The filtrate was purified using silica gel column chromatography
(0-30% EtOAc in DCM) to give the title compound A61 (0.170 g, 14%).
LCMS-D: rt 3.622 min; m/z 362.1 [M-H].sup.-.
(b) tert-Butyl
4-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl)py-
rimidin-2-yl)amino)-1H-pyrazole-1-carboxylate (A62)
[0597] A degassed mixture of tert-butyl
4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyrazole-1-carbo-
xylate A61 (0.373 g, 1.03 mmol), 2-(2-ethynylphenyl)propanamide K7
(0.452 g, 2.40 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.015 g, 0.02
mmol), t-Bu.sub.3PH.BF.sub.4 (0.011 g, 0.038 mmol) and CuI (0.007
g, 0.04 mmol) in 1,4-dioxane (6.0 mL) and DIPEA (0.90 mL, 5.2 mmol)
was heated in the microwave at 100.degree. C. for 30 minutes. The
mixture was concentrated under reduced pressure and purified using
silica gel column chromatography (0-100% EtOAc in petroleum benzine
40-60.degree. C.) to give the title compound A62 (0.309 g, 58%).
LCMS-D: rt 3.533 min; m/z 502.2 499.2 [M-H].sup.-.
(c)
2-(2-((2-((1H-Pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)et-
hynyl)phenyl)propanamide (A63)
[0598] A mixture of tert-butyl
4-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl)py-
rimidin-2-yl)amino)-1H-pyrazole-1-carboxylate A62 (0.31 g, 0.62
mmol) and TFA (0.90 mL, 12 mmol) in DCM (3.0 mL) was stirred for 2
hours at room temperature. The mixture was concentrated under
reduced pressure and diluted with water (50 mL) before extracting
with DCM (2.times.50 mL) and EtOAc (50 mL). The combined organic
residues were concentrated and purified using silica gel column
chromatography (50-100% EtOAc in petroleum benzine 40-60.degree. C.
then 0-10% MeOH in EtOAc) to give the title compound A63 (0.211 g,
85%). LCMS-D: rt 3.162 min; m/z 401.2 [M+H].sup.+.
(d)
2-(2-(2-(2-((1H-Pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)-
ethyl)phenyl)propanamide (54)
[0599] Et.sub.3N (0.5 mL) was added to
2-(2-((2-((1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)ethyn-
yl)phenyl)propanamide A63 (0.211 g, 0.527 mmol) and 10% Pd/C (0.155
g) in DMF (4 mL) and EtOAc (16 mL) and the mixture was stirred
under a hydrogen atmosphere for 16 hours. The mixture was filtered
through Celite, concentrated under reduced pressure and purified
using silica gel column chromatography (80-100% EtOAc in petroleum
benzine 40-60.degree. C. then 0-10% MeOH in EtOAc). The resulting
product was further triturated with a mixture of Et.sub.2O and
acetone and the precipitate was collected by vacuum filtration to
give the title compound 54 (0.055 g, 26%). LCMS-D: rt 3.178 min;
m/z 405.2 [M+H].sup.+.
Example 55
Synthesis of
2-(2-(2-(2-((1-methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)propanamide (55)
##STR00178##
[0600] (a)
4-Chloro-N-(1-methyl-1H-pyrazol-4-yl)-5-(trifluoromethyl)pyrimi-
din-2-amine (A64)
[0601] A 1.0 M solution of ZnCl.sub.2 in Et.sub.2O (5.07 mL, 5.07
mmol) was added to a solution of
2,4-dichloro-5-(trifluoromethyl)pyrimidine (1.00 g, 4.61 mmol) in
t-BuOH (25 mL) and DCE (25 mL) and the mixture was stirred for 10
minutes. The mixture was diluted with t-BuOH (25 mL) and DCE (25
mL) before 1-methyl-1H-pyrazol-4-amine (492 mg, 5.07 mmol) and
Et.sub.3N (1.93 mL, 13.83 mmol) were added and stirring was
continued at room temperature overnight. The volatiles were removed
in vacuo and the resultant residue suspended in water (250 mL) and
sonicated for 10 minutes. The solid was collected by vacuum
filtration and the filter cake washed with water and then air
dried. The solid was dissolved in DCM, adsorbed onto silica gel and
purified by column chromatography (Biotage Isolera, 40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A64 as a white solid (182 mg, 14%). LCMS-A:
rt 6.113 min; m/z 278.1 [M+H].sup.+.
(b)
2-(2-(2-(2-((1-Methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimi-
din-4-yl)ethyl)phenyl) propanamide (55)
[0602] A suspension of
4-chloro-N-(1-methyl-1H-pyrazol-4-yl)-5-(trifluoromethyl)pyrimidin-2-amin-
e A64 (182 mg, 0.656 mmol), 2-(2-ethynylphenyl)propanamide K7 (110
mg, 0.635 mmol), PPh.sub.3 (8 mg, 0.032 mmol) and CuI (6 mg, 0.032
mmol) in Et.sub.3N (1 mL) and DMF (3 mL) was sonicated for 10
minutes, PdCl.sub.2(PPh.sub.3).sub.2 (22 mg, 0.032 mmol) was added
and the reaction mixture was irradiated in the microwave at
120.degree. C. for 20 minutes. The resultant mixture was adsorbed
onto silica gel and purified by column chromatography (Biotage
Isolera, 40 g SiO.sub.2 cartridge, 0-100% EtOAc in petroleum
benzine 40-60.degree. C. then 0-100% MeOH in EtOAc) to give a
yellow solid. Further purification by silica gel column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100%
MeOH in EtOAc) gave a yellow solid (105 mg) that was taken up in
EtOAc (10 mL) and DMF (2 mL). 10% Pd/C (50 mg) was added and the
mixture was stirred at room temperature overnight under an
atmosphere of hydrogen. The mixture was diluted with EtOAc (50 mL),
filtered through Celite and the filter cake was washed with EtOAc
(50 mL). The washings were combined and the volatiles were removed
in vacuo. The resulting residue was adsorbed onto silica gel and
purified by column chromatography (Biotage Isolera, 12 g SiO.sub.2
cartridge, 0-100% MeOH in EtOAc) to give the title compound 55 as a
tan solid (31 mg, 29%). LCMS-A: rt 5.931 min; m/z 419.2
[M+H].sup.+.
Example 56
Synthesis of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide (56)
##STR00179##
[0603] (a) tert-Butyl
4-(4-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(A65)
[0604] A mixture of tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)-
piperidine-1-carboxylate K8 (2.054 g, 4.597 mmol),
2-(2-ethynylphenyl)propanamide K7 (1.027 g, 5.929 mmol), CuI (0.017
g, 0.089 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.062 g, 0.088 mmol)
and t-Bu.sub.3PH.BF.sub.4 (0.025 g, 0.087 mmol) in dioxane (20 mL)
and Et.sub.3N (2.5 mL, 18 mmol) under a nitrogen atmosphere was
heated in the microwave at 110.degree. C. for 40 minutes. The
mixture was concentrated under reduced pressure and purified using
silica gel column chromatography (50-100% EtOAc in petroleum
benzine 40-60.degree. C.) to give the title compound A65 (2.121 g,
79%). .sup.1H NMR (400 MHz, d6-DMSO) .delta. 10.48-10.33 (m, 1H),
8.82-8.73 (m, 1H), 8.09-7.97 (m, 1H), 7.64-7.55 (m, 2H), 7.56-7.44
(m, 2H), 7.41-7.34 (m, 1H), 7.29-7.22 (m, 1H), 7.01 (s, 1H),
4.39-4.29 (m, 1H), 4.18-3.99 (m, 3H), 2.99-2.80 (m, 2H), 2.02-1.94
(m, 2H), 1.83-1.70 (m, 2H), 1.41 (s, 9H), 1.39-1.31 (m, 3H).
(b) tert-Butyl
4-(4-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-(trifluoromethyl)pyrim-
idin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate (A66)
[0605] A solution of tert-butyl
4-(4-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate A65
(3.30 g, 5.66 mmol) in DMF (50 mL) and TEA (5 mL) was stirred with
10% Pd/C (wetted with ca. 53% water, 3.00 g) under an atmosphere of
H.sub.2 for 20 hours at 35.degree. C. The mixture was filtered
through Celite and the solvent was removed in vacuo. Purification
by column chromatography (Biotage Isolera, 120 g SiO.sub.2
cartridge, 20-100% EtOAc in petroleum benzine 40-60.degree. C.)
gave the title compound A66 as a yellow solid (3.32 g, 99%).
LCMS-D: rt 3.54 min; m/z 588 [M+H].sup.+.
(c)
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromet-
hyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (56)
[0606] A solution of tert-butyl
4-(4-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-(trifluoromethyl)pyrim-
idin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate A66 (3.41
g, 5.80 mmol) in DCM (50 mL) was cooled to 0.degree. C. and treated
with TFA (4.44 mL, 0.058 mol). The mixture was stirred at 0.degree.
C. for 1 hour and then at room temperature for 3 hours. An extra
aliquot of TFA (2.2 mL, 0.029 mol) was added to the mixture and
stirring was continued for 2 hours at room temperature. Sat. aq.
NaHCO.sub.3 (.about.50 mL) was carefully added, followed by aq.
NaOH (2 M, .about.50 mL). DCM was removed in vacuo and the
resultant aqueous mixture was diluted with H.sub.2O (.about.50 mL).
The white suspension was sonicated for 10 minutes and vacuum
filtered. The isolated solid was washed with water and air dried to
give the title compound 56 as a white solid (2.8 g, 99%). .sup.1H
NMR (400 MHz, d6-DMSO) .delta. 10.20-10.08 (m, 1H), 8.69-8.56 (m,
1H), 8.02-7.93 (m, 1H), 7.58 (s, 1H), 7.44-7.35 (m, 1H), 7.27-7.14
(m, 4H), 6.92-6.80 (m, 1H), 4.21-4.08 (m, 1H), 3.91-3.79 (m, 1H),
3.18-2.91 (m, 6H), 2.61-2.50 (m, 2H), 1.92-1.86 (m, 2H), 1.81-1.68
(m, 2H), 1.33 (d, J=6.9 Hz, 3H). LCMS-A: rt 4.79 min; m/z 488
[M+H].sup.+.
Example 56-1A and 56-2A
Separation of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide (56)
##STR00180##
[0608] Racemic
2-(2-(2-(2-((1-(Piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide was separated using the
chiral separation Method D.
[0609] Enantiomer A of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide (56-1A), Chiral
Characterisation Method F: rt 9.16 min, enantiomeric purity
>99%.
[0610] Enantiomer B of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide (56-2A), Chiral
Characterisation Method F: rt 6.31 min, enantiomeric purity
97.6%.
Example 57
Synthesis of
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57)
##STR00181##
[0611]
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(t-
rifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57)
[0612] To a solution of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide 56 (1.14 g, 2.33 mmol) in
MeOH (40 mL) was added formaldehyde solution (37% in water, 0.52
mL, 7.0 mmol) and sodium triacetoxyborohydride (1.98 g, 9.33 mmol)
under a nitrogen atmosphere. The mixture was stirred at room
temperature for 4 hours before concentrating under reduced
pressure. The mixture was diluted with sat. aq. NaHCO.sub.3 (75 mL)
and the aqueous phase was extracted with EtOAc (3.times.50 mL). The
combined organics were washed with brine and dried (MgSO.sub.4)
before the solvent was removed in vacuo to give the product as a
white solid (1.05 g, 90%). .sup.1H NMR (400 MHz, d.sub.6-DMSO)
.delta. 10.22-10.09 (m, 1H), 8.69-8.55 (m, 1H), 8.04-7.95 (m, 1H),
7.58 (s, 1H), 7.43-7.35 (m, 1H), 7.29-7.12 (m, 4H), 6.96-6.84 (m,
1H), 4.13-4.01 (m, 1H), 3.90-3.79 (m, 1H), 3.27-2.88 (m, 4H), 2.83
(d, J=10.0 Hz, 2H), 2.19 (s, 3H), 2.07-1.87 (m, 6H), 1.33 (d, J=6.8
Hz, 3H). LCMS-D: rt 3.03 min; m/z 502 [M+H].sup.+.
Example 57-1A and 57-2A
Separation of
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57)
##STR00182##
[0614]
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(t-
rifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57) was
separated using the chiral separation Method B.
[0615] Enantiomer A of
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57-1A), Chiral
Characterisation Method G: rt 14.23 min, enantiomeric purity
>99%.
[0616] Enantiomer B of
2-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (57-2A), Chiral
Characterisation Method G: rt 20.34 min, enantiomeric purity
>99%.
Example 58
Synthesis of
2-(2-(2-(2-((1-(1-acetylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (58)
##STR00183##
[0617]
2-(2-(2-(2-((1-(1-Acetylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(t-
rifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (58)
[0618] Acetic anhydride (15 .mu.L, 0.15 mmol) was added to a
solution of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide 56 (50 mg, 0.10 mmol) in
DCM (5 mL) and pyridine (12 .mu.L, 0.15 mmol) and the mixture was
stirred at room temperature overnight. The volatiles were removed
in vacuo and the resultant solid was suspended in water (25 mL) and
sonicated for 5 minutes. The solid was collected by filtration and
washed with sat. aq. Na.sub.2CO.sub.3 (20 mL), water (50 mL) then
petroleum benzine 40-60.degree. C. to give the title compound 58 as
a white solid (42 mg, 77%). LCMS-D: rt 3.176 min; m/z 530.4
[M+H].sup.+.
Example 59
Synthesis of
2-(2-(2-(2-((1-(1-ethylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluor-
omethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (59)
##STR00184##
[0619]
2-(2-(2-(2-((1-(1-Ethylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(tr-
ifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (59)
[0620] Acetaldehyde (17 .mu.L, 0.31 mmol) was added to a solution
of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide A67 (50 mg, 0.10 mmol) in
EtOH (20 mL). After 10 minutes, NaHB(OAc).sub.3 (86 mg, 0.41 mmol)
was added and the mixture was stirred for 3 hours. The reaction was
quenched with water (20 mL) and the volatiles were removed in
vacuo. The resultant solid was suspended in water (25 mL),
sonicated for 10 minutes, collected by filtration and then
dissolved in acetone (3 mL). Addition of petroleum benzine
40-60.degree. C. gave a precipitate that was collected by
filtration to give the title compound 59 as a tan solid (42 mg,
79%). LCMS-A: rt 4.808 min; m/z 516.3 [M+H].sup.+.
Example 60
Synthesis of
2-(2-(2-(2-((1-(1-isopropylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trif-
luoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (60)
##STR00185##
[0621]
2-(2-(2-(2-((1-(1-Isopropylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-
-(trifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (60)
[0622] A suspension of
2-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)propanamide A67 (80 mg, 0.16 mmol),
2-iodopropane (25 .mu.L, 0.25 mmol) and K.sub.2CO.sub.3 (68 mg,
0.49 mmol) in MeCN (5 mL) was irradiated in the microwave at
120.degree. C. for 40 minutes. Additional 2-iodopropane (25 .mu.L,
0.25 mmol) was added and the reaction mixture was irradiated in the
microwave at 120.degree. C. for another 40 minutes. Upon cooling,
the liquid was decanted from the solid and the volatiles were
removed in vacuo. The resultant solid was suspended in water (50
mL) and sonicated for 10 minutes before being filtered. The solid
was dissolved in acetone (3 mL) and precipitated by the addition of
petroleum benzine 40-60.degree. C. before being collected by
filtration and dried under high vacuum to give the title compound
60 as a tan solid (61 mg, 70%). LCMS-D: rt 3.060 min; m/z 530.4
[M+H].sup.+.
Example 61
Synthesis of
2-(2-(2-(2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-4--
yl)ethyl)phenyl)propanamide (61)
##STR00186##
[0623] (a)
4-Chloro-N-(6-methoxypyridin-3-yl)-5-(trifluoromethyl)pyrimidin-
-2-amine (A68)
[0624] A 1.0 M solution of ZnCl.sub.2 in Et.sub.2O (5.07 mL, 5.07
mmol) was added to a solution of
2,4-dichloro-5-(trifluoromethyl)pyrimidine (1.00 g, 4.61 mmol) in
t-BuOH (25 mL) and DCE (25 mL). The mixture was stirred for 10
minutes and then diluted with t-BuOH (25 mL) and DCE (25 mL) before
5-amino-2-methoxypyridine (629 mg, 5.07 mmol) and Et.sub.3N (1.93
mL, 13.8 mmol) were added. The mixture was stirred at room
temperature overnight and the volatiles were removed in vacuo. The
resultant residue was suspended in water (250 mL) and sonicated for
10 minutes before the solid was isolated by vacuum filtration. The
filter cake was washed with water and air dried and then dissolved
in DCM and adsorbed onto silica gel. Purification by column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C.) gave the title
compound A68 as a white solid (944 mg, 67%). LCMS-A: rt 6.62 min;
m/z 305 [M+H].sup.+.
(b)
2-(2-((2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-4-
-yl)ethynyl)phenyl) propanamide (A69)
[0625] A mixture of 2-(2-ethynylphenyl)propanamide K7 (0.120 g,
0.693 mmol),
4-chloro-N-(6-methoxypyridin-3-yl)-5-(trifluoromethyl)pyrimidin-2--
amine A68 (0.211 g, 0.693 mmol), CuI (0.013 g, 0.069 mmol),
t-Bu.sub.3PHBF.sub.4 (0.020 g, 0.069 mmol) and
PdCl.sub.2(PPh.sub.3).sub.2 (0.024 g, 0.035 mmol) in DMF (3 mL) was
bubbled with N.sub.2 for 5 minutes. Et.sub.3N (1 mL) was added and
the reaction mixture was stirred in the microwave at 120.degree. C.
for 15 minutes. The volatiles were removed in vacuo and the black
residue was adsorbed onto silica. Purification by column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C.) gave the title
compound A69 as a yellow solid (0.198 g, 65%). LCMS-D: rt 3.43 min;
m/z 442 [M+H].sup.+.
(c)
2-(2-(2-(2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl) propanamide (61)
[0626] A solution of
2-(2-((2-((6-methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl-
)ethynyl)phenyl)propanamide A69 (0.198 g, 0.449 mmol) in EtOAc (20
mL) and MeOH (10 mL) was stirred over 10% Pd/C (wetted with ca. 53%
water, 0.150 g) under an atmosphere of hydrogen for 16 hours. The
mixture was filtered through Celite and the solvent was removed in
vacuo. Purification by column chromatography (Biotage Isolera, 40 g
SiO.sub.2, 0-80% EtOAc in petroleum benzine 40-60.degree. C.) gave
the title compound 61 as a white solid (0.126 g, 63%). LCMS-D: rt
3.43 min; m/z 446 [M+H].sup.+.
Example 62
Synthesis of
2-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)propanamide (62)
##STR00187##
[0627] (a) tert-Butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-2-yl)pipe-
ridine-1-carboxylate (A70)
[0628] 2,4-Dichloro-5-(trifluoromethyl)pyrimidine (411 mg, 1.89
mmol) was stirred in 1:1 t-BuOH:DCE (100 mL) at room temperature. A
1.0 M ZnCl.sub.2 solution in Et.sub.2O (2.16 mL, 2.16 mmol) was
added cautiously and after addition the mixture was stirred at room
temperature for 20 minutes. tert-Butyl
4-(5-aminopyridin-2-yl)piperidine-1-carboxylate 19 (500 mg, 1.80
mmol) was added followed by NEt.sub.3 (0.30 mL, 2.16 mmol) and the
mixture was stirred at room temperature for 44 hours. The organic
solvents were evaporated to dryness and the crude tan solid was
suspended in water (250 mL) and sonicated for 10 minutes. The solid
was isolated by filtration, washed with water (2.times.100 mL) and
air-dried to give a cream solid. The solid was adsorbed onto silica
and purified by silica gel column chromatography (40 g SiO.sub.2
cartridge, 0-100% EtOAc in petroleum benzine 40-60.degree. C.) to
give the title compound A70 as an off white solid (346 mg, 42%).
LCMS-A: rt 5.949 min; m/z 458 [M+H].sup.+.
(b) tert-Butyl
4-(5-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate
(A71)
[0629] To a mixture of tert-butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-2-yl)pipe-
ridine-1-carboxylate A70 (0.397 g, 0.867 mmol),
2-(2-ethynylphenyl)propanamide K7 (0.195 g, 1.13 mmol), CuI (0.011
g, 0.057 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.039 g, 0.056 mmol)
and t-Bu.sub.3PH.BF.sub.4 (0.017 g, 0.058 mmol) under a nitrogen
atmosphere was added dioxane (12 mL) and DIPEA (0.60 mL, 3.4 mmol).
The mixture was stirred at room temperature for 96 hours before
being concentrated under reduced pressure and purifying by silica
gel column chromatography (Biotage Isolera, SiO.sub.2 cartridge,
0-100% EtOAc in petroleum benzine 40-60.degree. C. and 0-20% MeOH
in EtOAc) to give the title compound A71 (0.139 g, 27%). LCMS-D: rt
3.626 min; m/z 595.3 [M+H].sup.+.
(c) tert-Butyl
4-(5-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-(trifluoromethyl)pyrim-
idin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A72)
[0630] A mixture of tert-Butyl
4-(5-((4-((2-(1-amino-1-oxopropan-2-yl)phenyl)ethynyl)-5-(trifluoromethyl-
)pyrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate A71
(0.139 g, 0.234 mmol) and 10% Pd/C (0.156 g) in DMF (3.0 mL) and
Et.sub.3N (0.30 mL) was stirred under a hydrogen atmosphere for 112
hours. The mixture was filtered through Celite and concentrated
under reduced pressure before being purified using silica gel
column chromatography (Biotage Isolera, SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C., 0-20% MeOH in EtOAc)
to give the title compound A72 (0.060 g, 42%). LCMS-D: rt 3.568
min; m/z 599.4 [M+H].sup.+.
(d)
2-(2-(2-(2-((6-(Piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)-
pyrimidin-4-yl)ethyl)phenyl)propanamide (62)
[0631] TFA (0.50 mL, 6.5 mmol) was added to tert-butyl
4-(5-((4-(2-(1-amino-1-oxopropan-2-yl)phenethyl)-5-(trifluoromethyl)pyrim-
idin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate A72 (0.060 g,
0.099 mmol) in DCM (5 mL) and the mixture was stirred for 2 hours.
The mixture was concentrated under reduced pressure and quenched
with 25% aq. NaOH (20 mL). The aqueous phase was extracted with
EtOAc (3.times.20 mL) before the combined organics were dried
(phase separation cartridge) and concentrated under reduced
pressure. The suspension was sonicated with hexane and concentrated
under reduced pressure to give the title compound (0.066 g, 134%
residual solvent/water). LCMS-D: rt 3.025 min; m/z 499.3
[M+H].sup.+.
Example 63
Synthesis of
2-(2-(2-(2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (63)
##STR00188##
[0632]
2-(2-(2-(2-((6-(1-Methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifl-
uoromethyl)pyrimidin-4-yl)ethyl)phenyl)propanamide (63)
[0633] To a mixture of
2-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)propanamide 62 (0.064 g, 0.13 mmol) in
anhydrous MeOH (2.0 mL) was added 37% aqueous formaldehyde (0.04
mL, 0.5 mmol) and sodium triacetoxyborohydride (0.138 g, 0.649
mmol). The mixture was stirred for 3 hours at room temperature
under a nitrogen atmosphere and then quenched by the addition of
sat. aq. NaHCO.sub.3 (30 mL). The aqueous phase was extracted with
EtOAc (2.times.20 mL), the combined organics were dried using a
phase separation cartridge and then concentrated under reduced
pressure. The organic residues were purified by prep-LCMS to give
the title compound 63 (0.020 g, 30%) as the formic acid salt (2
equivs. determined by .sup.1H NMR). LCMS-A: rt 4.786 min; m/z 513.3
[M+H].sup.+.
Example 64
Synthesis of
1-(2-(2-(2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (64)
##STR00189##
[0634] (a)
1-(2-((2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyri-
midin-4-yl)ethynyl)phenyl)cyclopropanecarboxamide (A73)
[0635] A mixture of 1-(2-ethynylphenyl)cyclopropanecarboxamide K6
(0.120 g, 0.648 mmol),
4-chloro-N-(6-methoxypyridin-3-yl)-5-(trifluoromethyl)pyrimidin-2-amine
A68 (0.197 g, 0.648 mmol), CuI (0.012 g, 0.065 mmol),
t-Bu.sub.3PHBF.sub.4 (0.019 g, 0.065 mmol) and
PdCl.sub.2(PPh.sub.3).sub.2 (0.023 g, 0.032 mmol) in DMF (3 mL) was
bubbled with N.sub.2 for 5 minutes. Et.sub.3N (1 mL) was added and
the reaction mixture was stirred in the microwave at 120.degree. C.
for 15 minutes. The volatiles were removed in vacuo and the black
residue was adsorbed onto silica. Purification by column
chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-100%
EtOAc in petroleum benzine 40-60.degree. C.) gave the title
compound A73 as a yellow solid (0.229 g, 78%). LCMS-D: rt 3.47 min;
m/z 454 [M+H].sup.+.
(b)
1-(2-(2-(2-((6-Methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (64)
[0636] A solution of
1-(2-((2-((6-methoxypyridin-3-yl)amino)-5-(trifluoromethyl)pyrimidin-4-yl-
)ethynyl)phenyl)cyclopropanecarboxamide A73 (0.229 g, 0.505 mmol)
in EtOAc (20 mL) was stirred over 10% Pd/C (wetted with ca. 53%
water, 0.150 g) under an atmosphere of hydrogen for 16 hours. The
mixture was filtered through Celite and the solvent was removed in
vacuo. Purification by column chromatography (Biotage Isolera, 40 g
SiO.sub.2, 0-80% EtOAc in petroleum benzine 40-60.degree. C.) gave
an off-white solid contaminated with alkyne starting material. The
solid was dissolved in EtOAc (20 mL) and MeOH (10 mL) and stirred
over 10% Pd/C (wetted with ca. 53% water, 0.130 g) for 16 hours
under an atmosphere of hydrogen. The mixture was filtered through
Celite and the solvent was removed in vacuo to give the title
compound 64 as a white solid (0.130 g, 56%). LCMS-D: rt 3.51 min;
m/z 458 [M+H].sup.+.
Example 65
Synthesis of
1-(2-(2-(2-((1-Methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-
-4-yl)ethyl)phenyl)cyclopropanecarboxamide (65)
##STR00190##
[0637] (a)
4-Chloro-N-(1-methyl-1H-pyrazol-4-yl)-5-(trifluoromethyl)pyrimi-
din-2-amine (A74)
[0638] ZnCl.sub.2 in Et.sub.2O (1M, 23.2 mL, 23.2 mmol) was added
to 2,4-dichloro-5-trifluoromethylpyrimidine (5.2 mL, 39 mmol) in
DCE:t-BuOH (1:1, 150 mL) under a nitrogen atmosphere. The mixture
was stirred for 45 minutes before addition of
1-methyl-1H-pyrazol-4-amine (1.880 g, 19.36 mmol) in DCE/t-BuOH
(1:1, 150 mL) and DIPEA (3.2 mL, 18 mmol). The mixture was heated
to 50.degree. C. for 16 hours, cooled and filtered using vacuum
filtration. The solid was washed with MeOH and the filtrate
concentrated under reduced pressure before being poured into water.
The resulting precipitate was collected under vacuum filtration and
washed with water then petroleum benzine 40-60.degree. C. and
finally dissolved in Et.sub.2O and filtered. The Et.sub.2O filtrate
was concentrated under reduced pressure to give the crude product
which was further purified in batches by a combination of
trituration with Et.sub.2O and cyclohexane or silica gel column
chromatography (0-30% EtOAc in petroleum benzine 40-60.degree. C.)
then combined to give the title compound A74 (1.6 g, 30%). .sup.1H
NMR (400 MHz, d-DMSO) .delta. ppm 10.66-10.58 (m, 1H), 8.77-8.68
(m, 1H), 7.93-7.86 (m, 1H), 7.53 (s, 1H), 3.88-3.78 (m, 3H).
(b)
1-(2-((2-((1-Methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidi-
n-4-yl)ethynyl)phenyl)cyclopropanecarboxamide (A75)
[0639] To a degassed mixture of
4-chloro-N-(1-methyl-1H-pyrazol-4-yl)-5-(trifluoromethyl)pyrimidin-2-amin-
e A74 (0.178 g, 0.640 mmol),
1-(2-ethynylphenyl)cyclopropanecarboxamide K6 (0.172 g, 0.928
mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.005 g, 0.007 mmol),
t-Bu.sub.3PH.BF.sub.4 (0.005 g, 0.02 mmol) and copper(I) iodide
(0.003 g, 0.02 mmol) in DMF (8.0 mL) was added DIPEA (0.33 mL, 1.9
mmol). The mixture was heated in the microwave for 2.times.20
minutes at 100.degree. C. before being concentrated under reduced
pressure. Purification by silica gel column chromatography
(Isolera, 0-100% EtOAc petroleum benzine 40-60.degree. C. then
0-20% MeOH in EtOAc) gave the title compound A75 (0.120 g, 44%).
LCMS-D: rt 3.30 min; m/z 427.2 [M+H].sup.+.
(c)
1-(2-(2-(2-((1-Methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimi-
din-4-yl)ethyl)phenyl)cyclopropanecarboxamide (65)
[0640] A mixture of
1-(2-((2-((1-methyl-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl)pyrimidin-4-
-yl)ethynyl)phenyl)cyclopropanecarboxamide A75 (0.120 g, 0.281
mmol) and 10% Pd/C (0.135 g) in DMF (3 mL), EtOAc (5 mL) and
Et.sub.3N (0.5 mL) was stirred under a hydrogen atmosphere for 16
hours. The mixture was filtered through Celite and concentrated
under reduced pressure before purification by silica gel column
chromatography (0-15% MeOH in EtOAc). The product was triturated
with Et.sub.2O and the resulting precipitate was collected to give
the title compound 65 (0.090 g, 74%). LCMS-D: rt 3.316 min; m/z
431.2 [M+H].sup.+.
Examples 66
Synthesis of
1-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (66)
##STR00191##
[0641] (a) tert-Butyl
4-(4-((4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)p-
yrimidin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate
(A76)
[0642] A mixture of 1-(2-ethynylphenyl)cyclopropanecarboxamide K6
(0.250 g, 1.35 mmol), tert-butyl
4-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)-
piperidine-1-carboxylate K8 (0.603 g, 1.35 mmol), CuI (0.026 g,
0.135 mmol), t-Bu.sub.3PHBF.sub.4 (0.039 g, 0.135 mmol) and
PdCl.sub.2(PPh.sub.3).sub.2 (0.047 g, 0.067 mmol) in DMF (10 mL)
was bubbled with N.sub.2 for 5 minutes. Et.sub.3N (3 mL) was added
and the reaction mixture was stirred in the microwave at
120.degree. C. for 20 minutes. The volatiles were removed in vacuo
and the black residue was adsorbed onto silica. Purification by
column chromatography (Biotage Isolera, 40 g SiO.sub.2 cartridge,
0-100% EtOAc in petroleum benzine 40-60.degree. C.) gave a yellow
solid which was purified further by two iterations of column
chromatography (Biotage Isolera, 2.times.40 g SiO.sub.2 cartridges,
0-10% MeOH in DCM then 40 g SiO.sub.2 cartridge, 0-5% MeOH in DCM)
to give the title compound A76 as a yellow solid (0.405 g, 50%).
LCMS-D: rt 3.62 min; m/z 596 [M+H].sup.+.
(b) tert-Butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate (A77)
[0643] A mixture of tert-butyl
4-(4-((4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)p-
yrimidin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate A76
(0.405 g, 0.680 mmol) and 10% Pd/C (wetted with ca. 53% water,
0.200 g) in EtOAc (30 mL) and MeOH (15 mL) was stirred under an
atmosphere of H.sub.2 at room temperature for 16 hours. The mixture
was filtered through a plug of Celite and the filtrate was
concentrated in vacuo. Purification by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 0-10% MeOH in DCM) gave
a yellow oil which was further purified by column chromatography
(Biotage Isolera, 40 g SiO.sub.2 cartridge, 10-90% EtOAc in
petroleum benzine 40-60.degree. C.) to give the title compound A77
as a yellow solid (0.213 g, 52%). LCMS-D: rt 3.62 min; m/z 600
[M+H].sup.+.
(c)
1-(2-(2-(2-((1-(Piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromet-
hyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (66)
[0644] A solution of tert-butyl
4-(4-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate A77 (0.213
g, 0.355 mmol) in DCM (10 mL) was treated with TFA (1.09 mL, 14.2
mmol) and stirred for 16 hours at room temperature. The volatiles
were evaporated in vacuo before aq. HCl (2 M, 25 mL) was added to
the residue. The aqueous phase was extracted with EtOAc (3.times.25
mL) and the combined organics were washed with brine and dried over
MgSO.sub.4. The solvent was removed in vacuo and the crude product
was purified by column chromatography (Biotage Isolera, 12 g C18
cartridge, 0-100% MeOH in H.sub.2O) to give a yellow oil. This oil
was taken up in a minimum amount of DCM and the desired product was
precipitated by the addition of petroleum spirits. The solid was
isolated by filtration to give the title compound 66 as a yellow
solid (0.114 g, 64%). LCMS-D: rt 3.07 min; m/z 500 [M+H].sup.+.
Example 67
Synthesis of
1-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluo-
romethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(67)
##STR00192##
[0645] (a)
1-(2-(2-(2-((1-(1-Methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)--
5-(trifluoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(67)
[0646] A mixture of
1-(2-(2-(2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-5-(trifluoromethyl-
)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide 66 (0.081 g,
0.16 mmol) and formaldehyde (37 wt % in H.sub.2O; 36 .mu.L, 0.49
mmol) in MeOH (5 mL) was stirred at room temperature for 10
minutes. Sodium triacetoxyborohydride (0.137 g, 0.649 mmol) was
then added and stirring was continued for 3 hours at room
temperature. The volatiles were removed in vacuo and sat. aq.
NaHCO.sub.3 (20 mL) was added to the residue. The aqueous phase was
extracted with EtOAc (3.times.20 mL) and the combined organics were
washed with brine, dried (MgSO.sub.4) and the solvent was removed
in vacuo. The yellow residue was taken up in MeOH and loaded onto
an SCX cartridge (10 g). The column was eluted with 5 column
volumes of MeOH and then 5 column volumes of 2% v/v aqueous ammonia
in MeOH to release the amine product. The solvent was evaporated
under reduced pressure, the residue was taken up in DCM and a solid
was precipitated by the addition of petroleum benzine 40-60.degree.
C. The solid was isolated by filtration and air dried to give the
title compound 67 as a yellow solid (0.048 g, 58%). LCMS-A: rt 4.82
min; m/z 514 [M+H].sup.+.
Example 68
Synthesis of
1-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (68)
##STR00193##
[0647] (a) tert-Butyl
4-(5-((4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)p-
yrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A78)
[0648] To a mixture of tert-butyl
4-(5-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)pyridin-2-yl)pipe-
ridine-1-carboxylate A70 (0.208 g, 0.453 mmol),
1-(2-ethynylphenyl)cyclopropanecarboxamide K6 (0.079 g, 0.42 mmol),
CuI (0.003 g, 0.02 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.017 g,
0.024 mmol) and t-Bu.sub.3PH.BF.sub.4 (0.009 g, 0.03 mmol) under a
nitrogen atmosphere was added dioxane (6.0 mL) and DIPEA (0.23 mL,
1.3 mmol). The mixture was stirred at 85.degree. C. for 4 hours and
then concentrated under reduced pressure. Purification by silica
gel column chromatography (Biotage Isolera, 0-100% EtOAc in
Petroleum Benzine 40-60.degree. C., 0-20% MeOH in EtOAc) gave the
title compound A78 (0.118 g, 46%). LCMS-D: rt 3.645 min; m/z 607.4
[M+H].sup.+.
(b) tert-Butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate (A79)
[0649] DMF (3 mL) and Et.sub.3N (0.30 mL) were added to a mixture
of tert-butyl
4-(5-((4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)p-
yrimidin-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate A78
(0.118 g, 0.194 mmol) and 10% Pd/C (0.182 g). The mixture was
stirred under a hydrogen atmosphere for 20 hours, filtered through
Celite and the filtrate concentrated under reduced pressure.
Purification by silica gel column chromatography (0-100% EtOAc in
Petroleum Benzine 40-60.degree. C.) gave the title compound A79
(0.080 g, 67%). LCMS-A: rt 5.852 min; m/z 611.3 [M+H].sup.+.
(c)
1-(2-(2-(2-((6-(Piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)-
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (68)
[0650] TFA (0.50 mL, 6.5 mmol) was added to a mixture of tert-Butyl
4-(5-((4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimid-
in-2-yl)amino)pyridin-2-yl)piperidine-1-carboxylate A79 (0.080 g,
0.13 mmol) in DCM (5 mL) and the mixture stirred for 4 hours. The
mixture was concentrated under reduced pressure and quenched with
25% aqueous NaOH (20 mL) before extracting with EtOAc (3.times.20
mL). The combined organic extracts were concentrated under reduced
pressure and the above procedure repeated with DCM (1.5 mL) and TFA
(0.2 mL) for 3 hours. The mixture was concentrated under reduced
pressure and quenched with 25% aqueous NaOH (20 mL) before
extracting with EtOAc (3.times.20 mL). The combined organic
residues were dried (phase separation cartridge) and concentrated
under reduced pressure to give the title compound 68 (0.069 g,
quantitative). LCMS-A: rt 4.783 min; m/z 511.3 [M+H].sup.+.
Example 69
Synthesis of
1-(2-(2-(2-((6-(1-methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (69)
##STR00194##
[0651]
1-(2-(2-(2-((6-(1-Methylpiperidin-4-yl)pyridin-3-yl)amino)-5-(trifl-
uoromethyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
(69)
[0652] To a mixture of
1-(2-(2-(2-((6-(piperidin-4-yl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyr-
imidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide 68 (0.069 g, 0.14
mmol) in anhydrous MeOH (2.0 mL) was added 37% aqueous formaldehyde
(0.04 mL, 0.5 mmol) and sodium triacetoxyborohydride (0.147 g,
0.695 mmol). The mixture was stirred for 3 hours at room
temperature under a nitrogen atmosphere and then quenched with sat.
aq. NaHCO.sub.3 (30 mL). The aqueous phase was extracted with EtOAc
(2.times.20 mL) and the combined organics were dried using a phase
separation cartridge and concentrated in vacuo. Purification by
prep-LCMS gave the title compound 69 (0.020 g, 29%) as the formic
acid salt (2 equivs. determined by 1H NMR). LCMS-A: rt 4.802 min;
m/z 525.3 [M+H].sup.+.
Example 70
Synthesis of tert-Butyl
1-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)pyrimidi-
n-2-ylamino)phenyl)ethylcarbamate (70)
##STR00195##
[0653] (a) tert-Butyl
(1-(4-((4-chloro-5-(trifluoromethyl)pyrimidin-2-yl)amino)phenyl)ethyl)car-
bamate (A80)
[0654] 2,4-Dichloro-5-(trifluoromethyl)pyrimidine (1.93 g, 8.89
mmol) was stirred in a 1:1 t-BuOH:DCE mixture (200 mL) at room
temperature. A 1.0 M ZnCl.sub.2 solution in Et.sub.2O (10.2 mL,
10.2 mmol) was added cautiously and the mixture was stirred at room
temperature for 20 minutes. A 1:1 t-BuOH:DCE mixture (100 mL) was
added followed by tert-butyl (1-(4-aminophenyl)ethyl)carbamate
(2.00 g, 8.46 mmol) and Et.sub.3N (1.42 mL, 10.2 mmol). The mixture
was stirred at room temperature overnight before the volatiles were
evaporated in vacuo. EtOH (30 mL) was added, the suspension was
sonicated for 1 minute before the solid was filtered, washed with
EtOH (10 mL) and dried in vacuo to give the title compound A80 as a
cream solid (3.2 g, 91%). LCMS-C: rt 4.88 min.
(b) tert-Butyl
1-(4-(4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)py-
rimidin-2-ylamino)phenyl)ethylcarbamate (A81)
[0655] A solution of 1-(2-ethynylphenyl)cyclopropanecarboxamide K6
(0.15 g, 0.81 mmol) in DMF (3 mL) was added to a reaction vessel
containing tert-butyl
1-(4-(4-chloro-5-(trifluoromethyl)pyrimidin-2-ylamino)phenyl)ethylcarbama-
te A80 (0.34 g, 0.81 mmol), PdCl.sub.2(PPh.sub.3).sub.2 (0.028 g,
0.040 mmol) tri-tert-butylphosphonium tetrafluoroborate (0.023 g,
0.081 mmol) and copper(I)iodide (0.015 g, 0.081 mmol). The mixture
was bubbled with nitrogen for 10 minutes before Et.sub.3N (1.5 mL)
was added. The mixture was heated at 120.degree. C. under microwave
irradiation for 15 minutes. The volatiles were removed in vacuo and
the residue was purified by silica gel column chromatography
(Combiflash Rf, 0-100% EtOAc in cyclohexane) to give the title
compound A81 as a yellow oil (0.26 g, 56%). LCMS-C: rt 5.80 min;
m/z 566 [M+H].sup.+.
(c) tert-Butyl
1-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)
pyrimidin-2-ylamino)phenyl)ethylcarbamate (70)
[0656] A solution of tert-butyl
1-(4-(4-((2-(1-carbamoylcyclopropyl)phenyl)ethynyl)-5-(trifluoromethyl)py-
rimidin-2-ylamino)phenyl)ethylcarbamate (0.25 g, 0.44 mmol) in
EtOAc (12 mL) and DMF (2.0 mL) was stirred with Pd/C 10% (0.15 g)
under an atmosphere of hydrogen for 16 hours at ambient
temperature. The reaction mixture was diluted with EtOAc, filtered
through a plug of Celite and washed with EtOAc. The solvents were
removed in vacuo and the crude residue was purified by silica gel
column chromatography (Combiflash Rf 0-100% EtOAc in cyclohexane)
to give the title compound 70 as a colourless oil (0.14 g, 56%).
LCMS-C: rt 5.88 min; m/z 570 [M+H].sup.+.
Example 70A
Synthesis of
1-(2-(2-(2-(4-(1-aminoethyl)phenylamino)-5-(trifluoromethyl)pyrimidin-4-y-
l)ethyl)phenyl)cyclopropanecarboxamide (70A)
##STR00196##
[0657]
1-(2-(2-(2-(4-(1-Aminoethyl)phenylamino)-5-(trifluoromethyl)pyrimid-
in-4-yl)ethyl)phenyl)cyclopropanecarboxamide (70A)
[0658] To a solution of tert-butyl
1-(4-(4-(2-(1-carbamoylcyclopropyl)phenethyl)-5-(trifluoromethyl)
pyrimidin-2-ylamino)phenyl)ethylcarbamate 70 (0.11 g, 0.19 mmol) in
DCM (4 mL) was added trifluoroacetic acid (2 mL). The mixture was
stirred at ambient temperature for 16 hours before the volatiles
were removed in vacuo. The crude residue was purified using an SCX
conditioned with MeOH, product was eluted off with 2 M ammonia in
EtOH) to give the title compound 70A as a cream solid (0.073 g,
81%). LCMS-C: rt 4.48 min; m/z 470 [M+H].sup.+.
Example 71
Synthesis of
1-(2-(2-(2-(4-(1-acetamidoethyl)phenylamino)-5-(trifluoromethyl)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (71)
##STR00197##
[0659]
1-(2-(2-(2-(4-(1-Acetamidoethyl)phenylamino)-5-(trifluoromethyl)
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide (71)
[0660] To a solution of
1-(2-(2-(2-(4-(1-aminoethyl)phenylamino)-5-(trifluoromethyl)pyrimidin-4-y-
l)ethyl)phenyl)cyclopropanecarboxamide 70A (0.015 g, 0.032 mmol) in
DCM (1 mL) was added pyridine (0.0040 mL, 0.050 mmol) and acetic
anhydride (0.0050 mL, 0.053 mmol) and the reaction mixture was
stirred at ambient temperature overnight. The mixture was
partitioned between water and EtOAc, the layers were separated and
the water layer was extracted with EtOAc (2 times). The combined
organic extracts were washed with brine, dried (Na.sub.2SO.sub.4)
and evaporated in vacuo to give the title compound 71 as a light
brown solid (0.016 g, 98%). LCMS-C: rt 5.24 min; m/z 512
[M+H].sup.+.
[0661] In addition to the Examples provided above, and the
description of the synthesis of those compounds, it would be
appreciated by a person skilled in the art that the following
compounds could also be prepared:
Example 72
Synthesis of
1-(2-(2-(2-((6-(1-aminoethyl)pyridin-3-yl)amino)-5-(trifluoromethyl)pyrim-
idin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
##STR00198##
[0663] This compound is prepared according to the methods described
for Example 2.
Example 73
Synthesis of
1-(2-(2-(2-((6-(1-(methylamino)ethyl)pyridin-3-yl)amino)-5-(trifluorometh-
yl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
##STR00199##
[0665] This compound is prepared via a methyl ketone intermediate
corresponding to Example 32 followed by reductive amination with
methylamine as described in Scheme V.
Example 74
Synthesis of
1-(2-(2-(2-((6-(1-(azetidin-1-yl)ethyl)pyridin-3-yl)amino)-5-(trifluorome-
thyl)pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
##STR00200##
[0667] This compound is prepared via a methyl ketone intermediate
corresponding to Example 32 followed by reductive amination with
azetidine as described in Scheme V.
Example 75
Synthesis of
1-(2-(2-(2-((6-(1-morpholinoethyl)pyridin-3-yl)amino)-5-(trifluoromethyl)-
pyrimidin-4-yl)ethyl)phenyl)cyclopropanecarboxamide
##STR00201##
[0669] This compound is prepared via a methyl ketone intermediate
corresponding to Example 32 followed by reductive amination with
morpholine as described in Scheme V.
Example 76
Synthesis of [Insert Compound Name]
##STR00202##
[0671]
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarbox-
amide A14 and 1-ethanol-1H-pyrazole-4-amine were reacted as
described for Example 34 to give Example 76 as a light yellow solid
(0.064 g, 63%). LCMS-C: rt 4.78 min; m/z 427 [M+H].sup.+.
Example 77
Synthesis of [Insert Compound Name]
##STR00203##
[0673]
1-(2-(2-(2,5-dichloropyrimidin-4-yl)ethyl)phenyl)cyclopropanecarbox-
amide A14 and 1-methoxyethanol-1H-pyrazole-4-amine were reacted as
described for Example 34 to give Example 77 as a light yellow oil
(0.060 g, 57%). LCMS-B: rt 6.43 min; m/z 441 [M+H].sup.+.
Biological Assays
[0674] The activity of compounds of the invention can be profiled
using biochemical and cellular assays.
[0675] Primary potency at VEGFR3 can be assessed using an Alpha
Screen.TM. technology biochemical assay.
[0676] The ability of compounds of the invention to inhibit VEGFR3
within cells can be assessed with an ELISA type assay.
VEGFR3 Biochemical Assay
[0677] Compounds of the invention may be tested for in vitro
activity in the following assay: A biotin labelled peptide is used
as substrate (amino acid sequence:
Biotin-Glu-Gly-Pro-Trp-Leu-Glu-Glu-Glu-Glu-Glu-Ala-Tyr-Gly-Trp-Met-Asp-Ph-
e-NH.sub.2). VEGFR3 cytoplasmic domain (amino acids 798-1298) was
purchased as N-terminal GST-fusion protein ("the enzyme"). The 15
.mu.L assay reactions are run in Greiner brand white 384-well low
volume plates. All reactions contained 10 mM HEPES pH 7.4, 10 mM
MgCl.sub.2, 0.01% (v/v) Tween-20, 50 .mu.M Na.sub.3VO.sub.4, 0.01%
(w/v) albumin from chicken egg white, 1 mM Dithiothreitol, 111 nM
peptide substrate, 500 .mu.M ATP, and 3.8 ng/reaction enzyme, with
the enzyme being omitted from negative control reactions. Compounds
were added in a volume of 100 nL from dilution series prepared in
DMSO, positive and negative control reactions receiving the same
volume DMSO without compound. The plates were sealed with adhesive
seals and incubated for 90 minutes at 30 degree Celsius. The
reactions were stopped with the detection reagents added at the
same time as follows: Product formation was quantified as amplified
luminescence between PerkinElmer AlphaScreen.TM. beads, using
Streptavidin-coated donor and anti-phosphotyrosine (P-Tyr-100)
acceptor beads. To each reaction, 5 .mu.L containing 10 mM HEPES pH
7.4, 25 mM NaCl, 100 mM EDTA, 0.01% (v/v) Tween-20, and 6.25
.mu.g/mL of each bead type were added. Plates were incubated for 6
hours before being read on a PerkinElmer EnVision.TM. plate reader
in HTS Alphascreen.TM. mode. IC.sub.50 values were obtained by
calculating percent inhibition (%1) for each reaction relative to
controls on the same plate (% I=(I-CN)/(CP-CN) where CN/=CP are the
averages of the negative/positive reactions, respectively), then
fitting the % I data vs. compound concentration [I] to %
I=(A+((B-A)/(I+((C/[I]) D)))) where A is the lower asymptote, B is
the upper asymptote, C is the IC.sub.50 value, and D is the slope
factor.
[0678] The above assay was also run in a modified form in some
cases (indicated below with *). In these cases, VEGFR3 cytoplasmic
domain (amino acids 818-1177, lacking 949-1002 of UniProt accession
number P35916) was expressed and purified as N-terminal
Hexa-His-fusion protein ("the enzyme"), rather than using the
N-terminal GST-fusion protein. The assay conditions were the same
as above but with 1 .mu.M ATP and 8 ng/reaction of the enzyme. The
comparable performance of both assay versions was monitored using
benchmark compounds as described in the literature.
VEGFR3 Biochemical Assay Results
TABLE-US-00003 [0679] Compound IC.sub.50 (nM) 1 91 1-1A 103 1-2A 75
2 10 3 9 4 22 5 28 6 8 7 10 8 10 9 26 10 92 11 1302 12 39 13 4 14 6
15 11 16 1 17 9 18 3 19 3 20 7 21 7 22 20 23 11 24 2 24A 14 25 7 26
12 27 47 28 268 29 8 30 4 31 48 32 44 33 60 34 7 35 9 36 16 37 31
38 32 39 44 40 160 41 45 42 154 43 12 44 9 45 3 46 13 47 5 48 12 49
23 50 46 51 15 51-1A 25 51-2A 41 52 23 53 228 54 17 55 15 56 27
56-1A 31 56-2A 93 57 27 58 22 59 86 60 89 61 118 62 492 63 28 64 8
65 3 66 7 67 7 68 28 69 63 70 64 70A 22 71 10 76 4 77 3
VEGFR3 Phospho ELISA Assay
[0680] Compounds of the invention may be tested for in vitro
activity in the following assay:
[0681] Adult human dermal lymphatic microvascular endothelial cells
(HMVEC-dLyAD) (Cat# CC-2810, Lonza) were seeded into clear-bottom,
TC treated 12 well plates (Cat #665180, Greiner Bio-One) in EGM-2MV
(Cat# CC-3202, Lonza) at 180,000 cells/well (volume 1 mL), and the
plates incubated at 37.degree. C. and 5% CO.sub.2 for 6 hours. The
media was replaced with EBM-2 (Cat # CC-3156, Lonza)+0.1% BSA (Cat#
A8412, Sigma) and cells incubated for a further period (overnight
at 37.degree. C. and 5% CO.sub.2).
[0682] 96 well Maxisorp immuno plates (Cat #439454, Nunc) were
coated with 100 .mu.L of Total VEGFR3 capture antibody (Part
#841888, Human Total VEGFR3/FLT4 ELISA Kit, Cat # DYC3491, R&D
Systems), or Phospho VEGFR3 Capture antibody (Part #841885, Human
Phospho VEGFR3/FLT4 ELISA Kit, Cat# DYC2724, R&D Systems). The
plates were covered and incubated at room temperature
overnight.
[0683] The coating antibody was flicked out and the plates washed
three times with Wash Buffer (Phosphate buffered saline (137 mM
NaCl, 2.7 nM KCl, 8.1 nM Na.sub.2HPO.sub.4, 1.5 mL
KH.sub.2PO.sub.4, pH 7.2-7.4), 0.05% Tween 20). 300 .mu.L of
blocking buffer (5% v/v Tween 20, 5% w/v sucrose in PBS) was then
added to wells and plate incubated for 2 hours at room temperature.
Blocking solution is flicked out and plates washed three times and
tapped dry.
[0684] Compound dilution series were prepared in EBM-2 (Cat #
CC-3156, Lonza)+0.1% BSA (Cat# A8412, Sigma) with constant 0.1%
DMSO concentration. 439 .mu.L of sample or vehicle control was
added to the cell monolayers. Cells are treated for 1 hour at
37.degree. C. and 5% CO.sub.2. 250 ng/mL Recombinant human VEGFC
(Cat #2179-VC, R & D Systems) added to wells and plates
incubated for an additional 10 minutes at 37.degree. C. and 5%
CO.sub.2.
[0685] The media and compounds were removed and the cell monolayer
washed once in Dulbecco's Phosphate Buffered Saline (Cat
#21600-044, Invitrogen). 130 .mu.L of Lysis buffer added to wells
and cell lysate harvested and transferred to tubes and stored on
ice. Complete lysis buffer was prepared by adding 10 .mu.L Protease
Inhibitor Cocktail (Cat # P8340, Sigma-Aldrich), 10 .mu.L PMSF
(Phenylmethanesulfonyl fluoride, Cat # P7626, Sigma-Aldrich,
prepared as 500 mM DMSO stock) per 1 mL of Phosphosafe.TM.
Extraction Reagent (Cat #71296, Merck).
[0686] The harvested samples were then diluted 1:2 in IC Diluent
#18 (5% Tween 20/PBS) and 100 .mu.L transferred to the Total and
Phospho VEGFR3 coated, blocked and washed 96 well plates and
incubated for 2 hours at room temperature. The plates were then
washed three times in wash buffer as described above and tapped
dry. For detection of Total VEGFR3 100 .mu.L of Detection antibody
(Total VEGFR3 Detection Antibody Part#841888 in Total VEGFR3 kit)
diluted in IC Diluent #1 (1% w/v BSA (Cat # A7906,
Sigma-Aldrich)/PBS) was added to wells and the plate incubated for
2 hours at room temperature. The plate was then washed three times
in wash buffer and tapped dry. 100 .mu.L of streptavidin-HPR
diluted in IC diluent #1 Streptavidin-HRP, Part #890803 in Total
VEGFR3 kit) was added to wells and incubated at room temperature
for 20 minutes followed by washing as described above. 100 .mu.L
Substrate solution (3,3',5,5'-Tetramethylbenzidine (TMB) Liquid
Substrate System for ELISA, Cat # T0440, Sigma-Aldrich) was added
and the plate incubated for 20 minutes in the dark at room
temperature followed by the addition of 50 .mu.L stop solution (2 M
H.sub.2SO.sub.4).
[0687] Total VEGFR3 levels were quantified using a Multiskan Ascent
plate reader and Ascent software fitted with 450 nm filter.
[0688] For detection of Phospho VEGFR3, 100 .mu.L of Detection
antibody (Anti-Phospho-Tyrosine-HRP Detection Antibody, Part
#841403 in Phospho VEGFR3 kit) was diluted in IC Diluent #1 (1% w/v
BSA/PBS), added to the wells and the plate incubated for 2 hours at
room temperature. The plate was then washed three times in wash
buffer as described above and tapped dry. 100 .mu.L Substrate
solution (3,3',5,5'-Tetramethylbenzidine (TMB) Liquid Substrate
System for ELISA, Cat # T0440, Sigma-Aldrich) was added and the
plate incubated for 20 minutes in the dark at room temperature
followed by the addition of 50 .mu.L stop solution (2 M
H.sub.2SO.sub.4).
[0689] Phospho VEGFR3 levels were quantified using a Multiscan
ascent plate reader and ascent software fitted with 450 nm
filter.
[0690] IC.sub.50 values are determined by first calculating the
level of phospho VEGFR3 relative to Total VEGFR3 according to the
following formula:
SRP = SP ST ##EQU00001##
[0691] Where SRP is the Sample Relative Phospho level, SP is
Phospho VEGFR3 reading and ST is Total VEGFR3 reading.
[0692] Percent inhibition (% I) for each lysate relative to vehicle
control (VEGFC stimulated) is then calculated according to the
following formula:
% I = SRP Vehicle - SRP Test SRP Vehicle * 100 ##EQU00002##
[0693] Where SRP is the Sample Relative Phospho level as calculated
above.
[0694] % I is plotted against compound concentration and data
fitted using a Sigmoidal dose response with IC50 determined from
curve.
VEGFR3 Phospho ELISA Assay Results
TABLE-US-00004 [0695] Compound IC.sub.50 (nM) 1 173 1-1A 329 1-2A
315 2 26 3 27 4 38 5 51 6 17 7 38 8 19 9 75 12 267 13 9 14 30 15 51
16 4 17 62 18 5 19 11 20 38 21 14 24 25 35 64 36 76 44 52 45 23 51
526 56 68 57 27 57-1A 38 57-2A 19 64 394 68 51
VEGFR2 Phospho ELISA Assay
[0696] Compounds of the invention may be tested for in vitro
activity in the following assay:
[0697] Adult human umbilical vein endothelial cells (HUVEC) (Cat#
CC-2519, Lonza) were seeded into clear-bottom, TC treated 12 well
plates (Cat #665180, Greiner Bio-One) in EGM-2 (Cat# CC-3162,
Lonza) at 180,000 cells/well (volume 1 mL), and the plates
incubated at 37.degree. C. and 5% CO.sub.2 for 6 hours. The media
was replaced with EBM-2 (Cat # CC-3156, Lonza)+0.1% BSA (Cat#
A8412, Sigma) and cells incubated for a further period (overnight
at 37.degree. C. and 5% CO.sub.2).
[0698] 96 well Maxisorp immuno plates (Cat #439454, Nunc) were
coated with 100 .mu.L of Total VEGFR2 capture antibody (Part
#841434, Human Total VEGFR2/FLT4 ELISA Kit, Cat # DYC1780, R&D
Systems), or Phospho VEGFR2 Capture antibody (Part #841419, Human
Phospho VEGFR2/FLT4 ELISA Kit, Cat# DYC1766, R&D Systems). The
plates were covered and incubated at room temperature
overnight.
[0699] The coating antibody was flicked out and the plates washed
three times with Wash Buffer (Phosphate buffered saline (137 mM
NaCl, 2.7 nM KCl, 8.1 nM Na.sub.2HPO.sub.4, 1.5 mL
KH.sub.2PO.sub.4, pH 7.2-7.4), 0.05% Tween 20). 300 .mu.L of
Blocking buffer (1% v/v BSA (Cat# A8412, Sigma) in PBS) was then
added to wells and plate incubated for 2 hours at room temperature.
Blocking solution is flicked out and plates washed three times and
tapped dry.
[0700] Compound dilution series were prepared in EBM-2 (Cat #
CC-3156, Lonza)+0.1% BSA (Cat# A8412, Sigma) with constant 0.1%
DMSO concentration. 427.5 .mu.L of sample or vehicle control was
added to the cell monolayers. Cells are treated for 1 hour at
37.degree. C. and 5% CO.sub.2. 50 ng/mL Recombinant human VEGF (Cat
#293-VC, R & D Systems) added to wells and plates incubated for
an additional 10 minutes at 37.degree. C. and 5% CO.sub.2.
[0701] The media and compounds were removed and the cell monolayer
washed once in Dulbecco's Phosphate Buffered Saline (Cat
#21600-044, Invitrogen). 130 .mu.L of Lysis buffer added to wells
and cell lysate harvested and transferred to tubes and stored on
ice. Complete lysis buffer was prepared by adding 10 .mu.L Protease
Inhibitor Cocktail (Cat # P8340, Sigma-Aldrich), 10 .mu.L PMSF
(Phenylmethanesulfonyl fluoride, Cat # P7626, Sigma-Aldrich,
prepared as 500 mM DMSO stock) per 1 mL of Phosphosafe.TM.
Extraction Reagent (Cat #71296, Merck).
[0702] The harvested samples were then diluted 1:2 in IC Diluent
#12 (1% NP-40, 20 nM Tris (pH 8.0), 137 mM NaCl, 10% glycerol, 2 mM
EDTA, 1 mM activated sodium orthovanadate) and 100 .mu.L
transferred to the Total and Phospho VEGFR2 coated, blocked and
washed 96 well plates and incubated for 2 hours at room
temperature. The plates were then washed three times in wash buffer
as described above and tapped dry.
[0703] For detection of Total VEGFR2 100 .mu.L of Detection
antibody (Total VEGFR2 Detection Antibody Part#841435 in Total
VEGFR2 kit) diluted in IC Diluent #14 (20 mM Tris, 137 mM
CaCl.sub.2, 0.05% Tween20, 0.1% BSA) was added to wells and the
plate incubated for 2 hours at room temperature. The plate was then
washed three times in wash buffer and tapped dry. 100 .mu.L of
streptavidin-HPR diluted in IC diluent #14 Streptavidin-HRP, Part
#890803 in Total VEGFR2 kit) was added to wells and incubated at
room temperature for 20 minutes followed by washing as described
above. 100 .mu.L Substrate solution (3,3',5,5'-Tetramethylbenzidine
(TMB) Liquid Substrate System for ELISA, Cat # T0440,
Sigma-Aldrich) was added and the plate incubated for 20 minutes in
the dark at room temperature followed by the addition of 50 .mu.L
stop solution (2 M H.sub.2SO.sub.4).
[0704] Total VEGFR2 levels were quantified using a Multiskan Ascent
plate reader and Ascent software fitted with 450 nm filter.
[0705] For detection of Phospho VEGFR2, 100 .mu.L of Detection
antibody (Anti-Phospho-Tyrosine-HRP Detection Antibody, Part
#841403 in Phospho VEGFR2 kit) was diluted in IC Diluent 14 (20 mM
Tris, 137 mM CaCl.sub.2, 0.05% Tween20, 0.1% BSA), was added to the
wells and the plate incubated for 2 hours at room temperature. The
plate was then washed three times in wash buffer as described above
and tapped dry. 100 .mu.L Substrate solution
(3,3',5,5'-Tetramethylbenzidine (TMB) Liquid Substrate System for
ELISA, Cat # T0440, Sigma-Aldrich) was added and the plate
incubated for 20 minutes in the dark at room temperature followed
by the addition of 50 .mu.L stop solution (2 M
H.sub.2SO.sub.4).
[0706] Phospho VEGFR2 levels were quantified using a Multiscan
ascent plate reader and ascent software fitted with 450 nm
filter.
[0707] IC.sub.50 values are determined by first calculating the
level of phospho VEGFR2 relative to Total VEGFR2 according to the
following formula:
SRP = SP ST ##EQU00003##
where SRP is the Sample Relative Phospho level, SP is Phospho
VEGFR2 reading and ST is Total VEGFR2 reading.
[0708] Percent inhibition (% I) for each lysate relative to vehicle
control (VEGF-A stimulated) is then calculated according to the
following formula:
% I = SRP Vehicle - SRP Test SRP Vehicle * 100 ##EQU00004##
where SRP is the Sample Relative Phospho level as calculated
above.
[0709] % I is plotted against compound concentration and data
fitted using a Sigmoidal dose response with IC.sub.50 determined
from plotted curve.
VEGFR2 Phospho ELISA Assay Results
TABLE-US-00005 [0710] Compound IC.sub.50 (nM) 1 >10000 2 283 3
1789 4 2036 5 4083 6 6608 7 629 8 897 9 >10000 12 >10000 13
257 14 357 15 1996 16 46 17 393 18 21 19 248 20 1755 21 73 24A 209
35 3786 36 8171 44 138 45 370 56 2224 57 394 57-1A 518 57-2A 382 68
4083
P397Y-FAK Inhibition MSD Platform Cellular Biomarker Assay
[0711] Compounds of the invention may be tested for in vitro
activity in the following assay:
[0712] 96-well plates (cat#MA6000, Meso Scale Discovery) are coated
with 30 .mu.L/well of mouse monoclonal FAK antibody [63D5]
(cat#ab72140, Abcam) pre-diluted in PBS to a concentration of 1
mg/mL. The plates are sealed with adhesive film and incubated for
16 hours at 4.degree. C. The antibody is then flicked out of the
plates and 150 .mu.L of 3% [w/v] Blocker A (cat#R93AA-1, Meso Scale
Discovery) is added. The plates are resealed with adhesive film and
incubated at room temperature on a shaker set at medium speed for 2
hours. The plates are then washed three times with a solution
containing 50 mM Tris-HCl pH 7.5, 0.15 M NaCl and 0.02% Tween-20,
before cell lysate addition described below.
[0713] Cells are split 1:2 into T150 cell culture flasks 2 days
prior to compound treatment. On the day prior to compound
treatment, 200 .mu.L media containing 20,000 cells is seeded into
all wells of white, clear-bottom, TC treated, pclear, 96-well
microtitre plates (cat#655098, Greiner Bio-One), and the plates are
incubated at 37.degree. C. and 5% CO.sub.2 for 36 hours. 1
.mu.L/well of compound is then added from dilution series prepared
in DMSO. Negative control wells receive the same volume of DMSO
without compounds, and positive control wells receive 2 .mu.M of a
control compound in the same volume of DMSO. Cells are treated for
1 hour at 37.degree. C. and 5% CO.sub.2. The media/compounds are
then flicked off and 55 .mu.L/well of ice-cold complete lysis
buffer is added. Complete lysis buffer is prepared by adding 1
tablet PhosSTOP complete phosphatase inhibitor (cat#04906837001,
Roche) and 1 tablet Complete, Mini, EDTA-free, protease inhibitor
(cat#04693159001, Roche) per 10 mL of incomplete lysis buffer (150
mM NaCl, 20 mM Tris-HCl pH 7.5, 1 mM EDTA, 1 mM EGTA, 1% Triton-X
100). Plates are incubated on ice for 30 minutes, with 30 seconds
high speed plate shaking every 5 minutes. 40 .mu.L/well of cell
lysate is transferred to the coated, blocked and washed 96-well
microtitre plates described above. The 96-well plates are sealed
with adhesive film and incubated for 16 hours at 4.degree. C. The
plates are then washed three times with a solution containing 50 mM
Tris-HCl pH 7.5, 0.15 M NaCl and 0.02% Tween-20 and tapped dry. 25
.mu.L/well of detection solution (1% [w/v] Blocker A (cat#R93AA-1,
Meso Scale Discovery) in 50 mM Tris-HCl pH 7.5, 0.15 M NaCl and
0.02% Tween-20, with 1:600 rabbit polyclonal FAK phospho Y397
antibody (cat#ab39967, Abcam), 1:1000 anti-rabbit sulfo-tag
antibody (cat#R32AB-1 Meso Scale Discovery) and 1:40 reconstituted
Blocker D-M (cat#D609-0100, Rockland Immunochemicals for Research))
is added, and the plates resealed with adhesive film and incubated
for 1 hour at room temperature on a plate shaker set to medium
speed. Plates are then washed three times with a solution
containing 50 mM Tris-HCl pH 7.5, 0.15 M NaCl and 0.02% Tween-20
and tapped dry. 150 .mu.L/well of Read Buffer T+Surfactant
(cat#R92TC-1, Meso Scale Discovery) is then added, and pFAK-397
levels quantified using a Meso Scale Discovery SECTOR Imager 6000
instrument.
[0714] IC.sub.50 values are determined by first calculating percent
inhibition (% I) for each lysate relative to controls on the same
plate (% I=(S-CP)/(CN-CP)) where S is the sample result, CN is the
average result of DMSO only treated negative controls, and CP is
the average result of 2 .mu.M treated positive controls. % I is
plotted against compound concentration [I] and the data fitted
using the following equation, % I=(A+((B-A)/(1+((C/[I]) D)))),
where A is the lower asymptote, B is the upper asymptote, C is the
IC50 value, and D is the slope factor.
P397Y-FAK Inhibition MSD Platform Cellular Biomarker Assay Results
for MDA-231-LNA Cells
TABLE-US-00006 [0715] Compound IC.sub.50 (nM) 1 1178 2 80 3 672 4
>2000 16 1159 54 88 55 115 56 517
B16F10 Melanoma Mouse Model
In-Life Study
[0716] Female BALB/c nu/nu mice at 6 to 8 weeks are inoculated with
4.times.10.sup.5 murine B16F10 melanoma cells (ATCC CRL-6475) in
Matrigel.RTM. suspension sub-dermally in the ear. 24 hours
following inoculation treatment commences via oral gavage twice
daily for 14 days. Animals are monitored daily for health, weight
changes and the appearance and number of satellite lesions tracking
down the ear.
[0717] On day 15 mice are sacrificed and ears removed and fixed in
4% paraformaldehyde. Ears are washed twice in PBS prior to being
photographed then stored in PBS for whole mount
immunohistochemistry analysis. Draining lymph nodes (superficial
cervicals) are removed, examined for the presence of metastatic
lesions and photographed prior to freezing in OTC medium.
[0718] Primary lesion size is determined through measurement of
length and width. Lesion volume is calculated using the following
equation (V=W.times.L.sup.2)/2).
Whole Mount Immunohistochemistry for Lymphatic and Blood
Vessels
[0719] Cartilage is removed from the edge of the paraformaldehyde
fixed ear prior to the ear being separated into the dorsal (with
primary lesion) and ventral sections. Dorsal ear sections are
premeablised in 0.3% Triton-x100 in PBS for 1 hour at 4.degree. C.
followed by blocking overnight at 4.degree. C. in 1% BSA/0.3%
Triton-x100/PBS on rotating wheel. Ears are then incubated for 24
hours in the primary antibody (Mouse LYVE-1 Biotinylated affinity
6402 purified pAB, R&D Systems, Cat # BAF2125 or Rat Anti-mouse
CD31 (PECAM) Clone 390, eBioscience, Cat #14-0311) at 4.degree. C.
on rotating wheel followed by 6.times.1 hour washes in 0.3%
Triton-x100/PBS at 4.degree. C.
[0720] Ear are then incubated for 24 hours in the secondary
detection reagent (Streptavidin Cy3 Conjugate, Sigma-Aldrich Cat #
S-6402 or Alexa Fluor 488 Goat Anti-Rat Igl (H+L) Antibody,
Invitrogen Molecular Probes Cat # A11006) at 4.degree. C. on
rotating wheel followed by 6.times.1 hour washes in 0.3%
Triton-x100/PBS at 4.degree. C.
[0721] Ear sections are refixed in 4% paraformaldehyde for 20
minutes then washed twice in PBS prior to mounting in whole mount
slide with Prolong Gold antifade reagent with Dapi (Invitrogen
Molecular Probes Cat# P36935).
[0722] Representative images of ears are taken on a Olympus BX51
microscope with DP72 CCD camera and associated software.
Caki-1 Tumor Model
[0723] Female BALB/c nu/nu mice at 6 to 8 weeks injected
subcutaneously (s.c.) with the human renal cancer cell line Caki-1
(ATCC HTB-46). Cells are resuspended in Dulbecco's PBS
(Sigma-Aldrich) and 5.times.10.sup.6 cells are injected s.c near
the third mammary fat pad. Tumors are grown to an average size of
150 mm.sup.3 prior to commencement of treatment. Treatment can
consist of a repeat oral gavage at varying doses dose. Tumour
growth and animal health is monitored over the course of the study.
Tumor growth is represented as mean tumor volume in mm.sup.3.
Animals are euthanized and tumors excised for either histologic
examination including development of lymphatic vessels and blood
vessels within the tumor and target engagement (phosphorylated
VEGFR3) using immunohistochemistry or alternatively using tumor
lysates to quantitate in situ inhibition of the target
(phosphorylated VEGFR3).
* * * * *