U.S. patent application number 17/516542 was filed with the patent office on 2022-05-19 for disubstituted benzothienyl-pyrrolotriazines and their use as fgfr kinase inhibitors.
The applicant listed for this patent is Bayer Intellectual Property GmbH, Bayer Pharma Aktiengesellschaft. Invention is credited to Ulf BOMER, Dirk BROHM, Marie-Pierre COLLIN, Sylvia GRUNEWALD, Melanie HEROULT, Walter HUBSCH, Mario LOBELL, Klemens LUSTIG, Verena VOHRINGER.
Application Number | 20220153745 17/516542 |
Document ID | / |
Family ID | 1000006122071 |
Filed Date | 2022-05-19 |
United States Patent
Application |
20220153745 |
Kind Code |
A1 |
BROHM; Dirk ; et
al. |
May 19, 2022 |
DISUBSTITUTED BENZOTHIENYL-PYRROLOTRIAZINES AND THEIR USE AS FGFR
KINASE INHIBITORS
Abstract
This invention relates to novel substituted
5-(1-benzothiophen-2-yl) pyrrolo[2,1-f][1,2,4]triazin-4-amine
derivatives of formula (I) wherein R.sup.1 is hydrogen, chloro,
methyl or methoxy, R.sup.2 is hydrogen or methoxy, with the proviso
that at least one of R.sup.1 and R.sup.2 is other than hydrogen,
G.sup.1 represents chloro, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, 5-membered aza-heteroaryl, or the
group --CH.sub.2--OR.sup.3, --CH.sub.2--NR.sup.4R.sup.5 or
--C(=0)-NR.sup.4R.sup.6, and G.sup.2 represents chloro, cyano,
(C.sub.1-C.sub.4)-alkyl, or the group --CR.sup.8AR.sup.8B--OH,
--CH.sub.2--NR.sup.9R.sup.10, --C(=0)--NR.sup.11R.sup.12 or
--CH.sub.2--OR.sup.15, having protein tyrosine kinase inhibitory
activities, to processes for the preparation of such compounds, to
pharmaceutical compositions containing such compounds, and to the
use of such compounds or compositions for treating proliferative
disorders, in particular cancer and tumor diseases.
Inventors: |
BROHM; Dirk; (Mettmann,
DE) ; HEROULT; Melanie; (Berlin, DE) ; COLLIN;
Marie-Pierre; (Wuppertal, DE) ; HUBSCH; Walter;
(Wuppertal, DE) ; LOBELL; Mario; (Wuppertal,
DE) ; LUSTIG; Klemens; (Wuppertal, DE) ;
GRUNEWALD; Sylvia; (Berlin, DE) ; BOMER; Ulf;
(Glienicke, DE) ; VOHRINGER; Verena; (Wolfegg,
DE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Bayer Intellectual Property GmbH
Bayer Pharma Aktiengesellschaft |
Monheim
Berlin |
|
DE
DE |
|
|
Family ID: |
1000006122071 |
Appl. No.: |
17/516542 |
Filed: |
November 1, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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15900725 |
Feb 20, 2018 |
|
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17516542 |
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14365424 |
Jun 13, 2014 |
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PCT/EP2012/074977 |
Dec 10, 2012 |
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15900725 |
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Current U.S.
Class: |
1/1 ; 514/210.21;
514/228.5; 514/228.8; 514/233.2; 514/243; 544/112; 544/182;
544/183; 544/58.2; 544/63 |
Current CPC
Class: |
C07D 487/04 20130101;
C07D 409/04 20130101; A61P 35/00 20180101; A61K 31/541 20130101;
C07D 409/14 20130101; A61K 31/5377 20130101; A61K 31/5355 20130101;
A61K 31/53 20130101 |
International
Class: |
C07D 487/04 20060101
C07D487/04; C07D 409/14 20060101 C07D409/14; C07D 409/04 20060101
C07D409/04; A61P 35/00 20060101 A61P035/00; A61K 31/53 20060101
A61K031/53; A61K 31/5355 20060101 A61K031/5355; A61K 31/5377
20060101 A61K031/5377; A61K 31/541 20060101 A61K031/541 |
Foreign Application Data
Date |
Code |
Application Number |
Dec 15, 2011 |
EP |
11193841.1 |
Claims
1-12. (canceled)
13: A process for preparing a compound of formula (I-A):
##STR00256## or an enantiomer, a diastereomer, a hydrate, a
solvate, a salt, a hydrate of the salt, or a solvate of the salt
thereof, wherein R.sup.1 is hydrogen, chloro, methyl or methoxy,
R.sup.2 is hydrogen or methoxy, with the proviso that at least one
of R.sup.1 and R.sup.2 is other than hydrogen, R.sup.3 is
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or phenyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, (C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, and (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, and (iii)
said phenyl is optionally substituted with one or two substituents
independently selected from the group consisting of fluoro, chloro,
bromo, cyano, trifluoromethyl, trifluoromethoxy,
(C.sub.1-C.sub.4)-alkyl and (C.sub.1-C.sub.4)-alkoxy, R.sup.9 and
R.sup.10, respectively, are joined and, taken together with the
nitrogen atom to which they are attached, form a monocyclic,
saturated 4- to 7-membered heterocycloalkyl ring which may contain
a second ring heteroatom selected from the group consisting of
N(R.sup.13), O, S and S(O).sub.2, and which may be substituted on
ring carbon atoms with up to three substituents independently
selected from the group consisting of fluoro,
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein R.sup.13 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl, formyl or
(C.sub.1-C.sub.4)-alkylcarbonyl, the process comprising: first
reacting a 6-substituted 4-aminopyrrolo[2,1-f][1,2,4]triazine of
formula (II) ##STR00257## wherein R.sup.3 is as defined above, with
formaldehyde and an amine of formula (III) ##STR00258## wherein
R.sup.9 and R.sup.10 are as defined above, in the presence of an
acid to give a compound of formula (IV) ##STR00259## wherein
R.sup.3, R.sup.9 and R.sup.10 are as defined above, then
brominating the compound of formula (IV) to give a compound of
formula (V) ##STR00260## wherein R.sup.3, R.sup.9 and R.sup.10 are
as defined above, and subsequently coupling the compound of formula
(V) with a benzothiophen-2-yl boronate of formula (VI) ##STR00261##
wherein R.sup.1 and R.sup.2 are as defined above, and R.sup.14 is
hydrogen or (C.sub.1-C.sub.4)-alkyl, or both R.sup.14 residues are
linked together to form a --(CH.sub.2).sub.2--,
--C(CH.sub.3).sub.2--C(CH.sub.3).sub.2--, --(CH.sub.2).sub.3--,
--CH.sub.2--C(CH.sub.3).sub.2--CH.sub.2-- or
--C(.dbd.O)--CH.sub.2--N(CH.sub.3)--CH.sub.2--C(.dbd.O)-- bridge,
in the presence of a palladium catalyst and a base to yield the
target compound of formula (I-A) ##STR00262## wherein R.sup.1,
R.sup.2, R.sup.3, R.sup.9 and R.sup.10 are as defined above.
14: The process of claim 13, further comprising: (i) separating the
compound of formula (I-A) into its enantiomers or diastereomers to
give an enantiomer or diastereomer thereof, and/or (ii) converting
the compound of formula (I-A) into a hydrate, a solvate, a salt, a
hydrate of the salt, or a solvate of the salt thereof by treating
the compound of formula (I-A) with the corresponding solvent and/or
acid or base.
15: The process of claim 14, wherein the compound of formula (I-A)
is converted into a hydrate, a salt, or a hydrate of the salt
thereof.
16: The process of claim 15, wherein the salt is a hydrochloride
salt, and the hydrate of the salt thereof is a hydrate of a
hydrochloride salt.
17: A process for preparing a compound of formula (I-A):
##STR00263## or an enantiomer, a diastereomer, a hydrate, a
solvate, a salt, a hydrate of the salt, or a solvate of the salt
thereof, wherein R.sup.1 is hydrogen, chloro, methyl or methoxy,
R.sup.2 is hydrogen or methoxy, with the proviso that at least one
of R.sup.1 and R.sup.2 is other than hydrogen, R.sup.3 is
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or phenyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, (C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)alkylaminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, and (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, and (iii)
said phenyl is optionally substituted with one or two substituents
independently selected from the group consisting of fluoro, chloro,
bromo, cyano, trifluoromethyl, trifluoromethoxy,
(C.sub.1-C.sub.4)-alkyl and (C.sub.1-C.sub.4)-alkoxy, R.sup.9 and
R.sup.10, respectively, are joined and, taken together with the
nitrogen atom to which they are attached, form a monocyclic,
saturated 4- to 7-membered heterocycloalkyl ring which may contain
a second ring heteroatom selected from the group consisting of
N(R.sup.13), O, S and S(O).sub.2, and which may be substituted on
ring carbon atoms with up to three substituents independently
selected from the group consisting of fluoro,
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein R.sup.13 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl, formyl or
(C.sub.1-C.sub.4)-alkylcarbonyl, the process comprising: first
formylating a 6-substituted 4-aminopyrrolo[2,1-f][1,2,4]triazine of
formula (II) ##STR00264## wherein R.sup.3 is as defined above, with
N,N-dimethylformamide in the presence of phosphoryl chloride to
give an aldehyde of formula (VII) ##STR00265## wherein R.sup.3 is
as defined above, then brominating the compound of formula (VII) to
give a compound of formula (VIII) ##STR00266## wherein R.sup.3 is
as defined above, and subsequently coupling the compound of formula
(VIII) with a benzothiophen-2-yl boronate of formula (VI)
##STR00267## wherein R.sup.1 and R.sup.2 are as defined above, and
R.sup.14 is hydrogen or (C.sub.1-C.sub.4)-alkyl, or both R.sup.14
residues are linked together to form a --(CH.sub.2).sub.2--,
--C(CH.sub.3).sub.2--C(CH.sub.3).sub.2--, --(CH.sub.2).sub.3--,
--CH.sub.2--C(CH.sub.3).sub.2--CH.sub.2-- or
--C(.dbd.O)--CH.sub.2--N(CH.sub.3)--CH.sub.2--C(.dbd.O)-- bridge,
in the presence of a palladium catalyst and a base to give a
compound of formula (IX) ##STR00268## wherein R.sup.1, R.sup.2 and
R.sup.3 are as defined above, and then reacting the compound of
formula (IX) with an amine of formula (III) ##STR00269## wherein
R.sup.9 and R.sup.10 are as defined above, in the presence of an
acid and a reducing agent to yield the target compound of formula
(I-A) ##STR00270## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.9 and
R.sup.10 are as defined above.
18: The process of claim 17, further comprising: (i) separating the
compound of formula (I-A) into its enantiomers or diastereomers to
give an enantiomer or diastereomer thereof, and/or (ii) converting
the compound of formula (I-A) into a hydrate, a solvate, a salt, a
hydrate of the salt, or a solvate of the salt thereof by treating
the compound of formula (I-A) with the corresponding solvent and/or
acid or base.
19: The process of claim 18, wherein the compound of formula (I-A)
is converted into a hydrate, a salt, or a hydrate of the salt
thereof.
20: The process of claim 19, wherein the salt is a hydrochloride
salt, and the hydrate of the salt thereof is a hydrate of a
hydrochloride salt.
21: A process for preparing a compound of formula (I-A):
##STR00271## or an enantiomer, a diastereomer, a hydrate, a
solvate, a salt, a hydrate of the salt, or a solvate of the salt
thereof, wherein R.sup.1 is hydrogen, chloro, methyl or methoxy,
R.sup.2 is hydrogen or methoxy, with the proviso that at least one
of R.sup.1 and R.sup.2 is other than hydrogen, R.sup.3 is
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or phenyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, (C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)alkylaminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, and (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, and (iii)
said phenyl is optionally substituted with one or two substituents
independently selected from the group consisting of fluoro, chloro,
bromo, cyano, trifluoromethyl, trifluoromethoxy,
(C.sub.1-C.sub.4)-alkyl and (C.sub.1-C.sub.4)-alkoxy, R.sup.9 and
R.sup.10, respectively, are joined and, taken together with the
nitrogen atom to which they are attached, form a monocyclic,
saturated 4- to 7-membered heterocycloalkyl ring which may contain
a second ring heteroatom selected from the group consisting of
N(R.sup.13), O, S and S(O).sub.2, and which may be substituted on
ring carbon atoms with up to three substituents independently
selected from the group consisting of fluoro,
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein R.sup.13 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl, formyl or
(C.sub.1-C.sub.4)-alkylcarbonyl, the process comprising: first
coupling a 6-substituted
4-amino-5-bromopyrrolo[2,1-f][1,2,4]-triazine of formula (XII)
##STR00272## with a benzothiophen-2-yl boronate of formula (VI)
##STR00273## wherein R.sup.1 and R.sup.2 and are as defined above,
and R.sup.14 is hydrogen or (C.sub.1-C.sub.4)-alkyl, or both
R.sup.14 residues are linked together to form a
--(CH.sub.2).sub.2--, --C(CH.sub.3).sub.2--C(CH.sub.3).sub.2--,
--(CH.sub.2).sub.3--, --CH.sub.2--C(CH.sub.3).sub.2--CH.sub.2-- or
--C(.dbd.O)--CH.sub.2--N(CH.sub.3)--CH.sub.2--C(.dbd.O)-- bridge,
in the presence of a palladium catalyst and a base to give a
compound of formula (XIII) ##STR00274## wherein R.sup.1 and R.sup.2
are as defined above, and then reacting the compound of formula
(XIII) with formaldehyde and an amine of formula (III) ##STR00275##
wherein R.sup.9 and R.sup.10 are as defined above, in the presence
of an acid to yield a compound of formula (I-C) ##STR00276##
wherein R.sup.1, R.sup.2, R.sup.9 and R.sup.10 are as defined
above, and subsequently converting the compound of formula (I-C)
into the corresponding 6-(halomethyl) derivative of formula (XVI)
##STR00277## wherein R.sup.1, R.sup.2, R.sup.9 and R.sup.10 are as
defined above, and X is chloro, bromo or iodo, and treating the
compound of formula (XVI) with an alcohol of formula (XVII)
R.sup.3A--OH (XVII), wherein R.sup.3A is R.sup.3 as defined above,
in the presence of a base to yield the target compound of formula
(I-A) ##STR00278## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.9 and
R.sup.10 are as defined above.
22: The process of claim 21, further comprising: (i) separating the
compound of formula (I-A) into its enantiomers or diastereomers to
give an enantiomer or diastereomer thereof, and/or (ii) converting
the compound of formula (I-A) into a hydrate, a solvate, a salt, a
hydrate of the salt, or a solvate of the salt thereof by treating
the compound of formula (I-A) with the corresponding solvent and/or
acid or base.
23: The process of claim 22, wherein the compound of formula (I-A)
is converted into a hydrate, a salt, or a hydrate of the salt
thereof.
24: The process of claim 23, wherein the salt is a hydrochloride
salt, and the hydrate of the salt thereof is a hydrate of a
hydrochloride salt.
25: A compound of formula (I): ##STR00279## wherein R.sup.1 is
hydrogen, chloro, methyl or methoxy, R.sup.2 is hydrogen or
methoxy, with the proviso that at least one of R.sup.1 and R.sup.2
is other than hydrogen; G.sup.1 is chloro, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, 5-membered aza-heteroaryl, or the
group --CH.sub.2--OR.sup.3, --CH.sub.2--NR.sup.4R.sup.5 or
--C(.dbd.O)--NR.sup.4R.sup.6, wherein R.sup.3 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or phenyl,
(i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted with
hydroxy, (C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, and (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)alkyl, hydroxy and amino, and (iii)
said phenyl is optionally substituted with one or two substituents
independently selected from the group consisting of fluoro, chloro,
bromo, cyano, trifluoromethyl, trifluoromethoxy,
(C.sub.1-C.sub.4)-alkyl and (C.sub.1-C.sub.4)alkoxy; R.sup.4 is
hydrogen or (C.sub.1-C.sub.4)-alkyl; R.sup.5 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkylcarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or 4- to 6-membered heterocycloalkyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, (C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
(C.sub.3-C.sub.6)-cycloalkyl, and (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)alkyl, hydroxy and amino, and (iii) said 4- to
6-membered heterocycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy, oxo and amino; R.sup.6 is
hydrogen, (C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or
4- to 6-membered heterocycloalkyl, wherein (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
(C.sub.1-C.sub.4)alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
(C.sub.3-C.sub.6)-cycloalkyl, and (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)alkyl, hydroxy and amino, and (iii) said 4- to
6-membered heterocycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy, oxo and amino; or R.sup.4 and
R.sup.5, or R.sup.4 and R.sup.6, respectively, are joined and,
taken together with the nitrogen atom to which they are attached,
form a monocyclic, saturated 4- to 7-membered heterocycloalkyl ring
which may contain a second ring heteroatom selected from N(R.sup.7)
and O, and which may be substituted on ring carbon atoms with one
or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and
aminocarbonyl, and wherein R.sup.7 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, formyl or (C.sub.1-C.sub.4)-alkylcarbonyl;
and G.sup.2 is chloro, cyano, (C.sub.1-C.sub.4)-alkyl, or the group
--CR.sup.8AR.sup.8B--OH, --CH.sub.2--NR.sup.9R.sup.10,
--C(.dbd.O)--NR.sup.11R.sup.12 or --CH.sub.2--OR.sup.15, wherein
R.sup.8A and R.sup.8B are independently selected from the group
consisting of hydrogen, (C.sub.1-C.sub.4)-alkyl, cyclopropyl and
cyclobutyl; R.sup.9 is hydrogen or (C.sub.1-C.sub.4)-alkyl;
R.sup.10 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkylcarbonyl, (C.sub.3-C.sub.6)-cycloalkyl or 4-
to 6-membered heterocycloalkyl, wherein (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
amino, aminocarbonyl, mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
di-(C.sub.1-C.sub.4)alkylaminocarbonyl, and (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)alkyl, hydroxy and amino, and (iii) said 4- to
6-membered heterocycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy, oxo and amino; R.sup.11 is
hydrogen or (C.sub.1-C.sub.4)-alkyl; R.sup.12 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or 4- to
6-membered heterocycloalkyl, wherein (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
amino, aminocarbonyl, mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
di-(C.sub.1-C.sub.4)alkylaminocarbonyl, and (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)alkyl, hydroxy and amino, and (iii) said 4- to
6-membered heterocycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy, oxo and amino; or R.sup.9 and
R.sup.10, or R.sup.11 and R.sup.12, respectively, are joined and,
taken together with the nitrogen atom to which they are attached,
form a monocyclic, saturated 4- to 7-membered heterocycloalkyl ring
which may contain a second ring heteroatom selected from
N(R.sup.13), O, S and S(O).sub.2, and which may be substituted on
ring carbon atoms with up to three substituents independently
selected from the group consisting of fluoro,
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein R.sup.13 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl, formyl or
(C.sub.1-C.sub.4)alkylcarbonyl; and R.sup.15 is
(C.sub.1-C.sub.4)-alkyl, with the proviso that G.sup.1 is not
chloro when G.sup.2 is chloro or cyano, or a pharmaceutically
acceptable salt, a hydrate, or a solvate thereof.
26: The compound of claim 25, wherein: R.sup.1 is chloro, methyl or
methoxy; R.sup.2 is hydrogen or methoxy; G.sup.1 is chloro,
(C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkoxycarbonyl or
5-membered aza-heteroaryl selected from the group consisting of
pyrazolyl, imidazolyl, oxazolyl, isoxazolyl and oxadiazolyl, or is
the group --CH.sub.2--OR.sup.3 or --CH.sub.2--NR.sup.4R.sup.5,
wherein R.sup.3 is hydrogen, (C.sub.1-C.sub.4)-alkyl or
(C.sub.3-C.sub.6)-cycloalkyl, wherein said (C.sub.1-C.sub.4)-alkyl
is optionally substituted with hydroxy, (C.sub.1-C.sub.4)-alkoxy,
hydroxycarbonyl, (C.sub.1-C.sub.4)-alkoxycarbonyl, amino,
aminocarbonyl, (C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro
atoms; R.sup.4 is hydrogen or (C.sub.1-C.sub.4)-alkyl; R.sup.5 is
hydrogen, (C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkylcarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or 5- or 6-membered heterocycloalkyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, hydroxycarbonyl or (C.sub.3-C.sub.6)-cycloalkyl, and
(ii) said 5- or 6-membered heterocycloalkyl is optionally
substituted with oxo; or R.sup.4 and R.sup.5 are joined and, taken
together with the nitrogen atom to which they are attached, form a
monocyclic, saturated 4- to 6-membered heterocycloalkyl ring which
may contain a second ring heteroatom selected from N(R.sup.7) and
O, and which may be substituted on a ring carbon atom with oxo or
hydroxy, and wherein R.sup.7 is hydrogen or
(C.sub.1-C.sub.4)-alkyl; and G.sup.2 is chloro, cyano,
(C.sub.1-C.sub.4)-alkyl, or the group --CR.sup.8AR.sup.8B--OH,
--CH.sub.2--NR.sup.9R.sup.10, --C(.dbd.O)--NR.sup.11R.sup.12 or
--CH.sub.2--OR.sup.15, wherein R.sup.8A and R.sup.8B are
independently selected from the group consisting of hydrogen,
(C.sub.1-C.sub.4)-alkyl and cyclopropyl; R.sup.9 is hydrogen or
methyl; R.sup.10 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkylcarbonyl, (C.sub.3-C.sub.6)-cycloalkyl or 5-
or 6-membered heterocycloalkyl, wherein (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy or
aminocarbonyl, and (ii) said 5- or 6-membered heterocycloalkyl is
optionally substituted with oxo; R.sup.11 is hydrogen or methyl;
R.sup.12 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl or 5- or 6-membered heterocycloalkyl,
wherein (i) said (C.sub.1-C.sub.4)-alkyl is optionally substituted
with hydroxy, and (ii) said 5- or 6-membered heterocycloalkyl is
optionally substituted with oxo; or R.sup.9 and R.sup.10, or
R.sup.11 and R.sup.12, respectively, are joined and, taken together
with the nitrogen atom to which they are attached, form a
monocyclic, saturated 4- to 6-membered heterocycloalkyl ring which
may contain a second ring heteroatom selected from N(R.sup.13), O,
S and S(O).sub.2, and which may be substituted on ring carbon atoms
with up to three substituents independently selected from the group
consisting of fluoro, (C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino
and aminocarbonyl, and wherein R.sup.13 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, cyclopropyl, cyclobutyl, formyl or
(C.sub.1-C.sub.4)-alkylcarbonyl; and R.sup.15 is methyl or ethyl,
with the proviso that G.sup.1 is not chloro when G.sup.2 is chloro
or cyano, or a pharmaceutically acceptable salt, a hydrate, or a
solvate thereof.
27: The compound of claim 25, wherein: R.sup.1 is methyl; R.sup.2
is methoxy; G.sup.1 is methyl, oxazol-5-yl or the group
--CH.sub.2--OR.sup.3 or --CH.sub.2--NR.sup.4R.sup.5, wherein
R.sup.3 is hydrogen, (C.sub.1-C.sub.4)-alkyl, cyclopropyl or
cyclobutyl, wherein said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, methoxy, ethoxy, hydroxycarbonyl,
methoxycarbonyl, ethoxycarbonyl, amino, aminocarbonyl, cyclopropyl,
cyclobutyl or up to three fluoro atoms; R.sup.4 is hydrogen, methyl
or ethyl; R.sup.5 is hydrogen, (C.sub.1-C.sub.4)-alkyl, acetyl,
cyclopropyl, cyclobutyl or 2-oxopyrrolidin-3-yl, wherein said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
hydroxycarbonyl, cyclopropyl or cyclobutyl; or R.sup.4 and R.sup.5
are joined and, taken together with the nitrogen atom to which they
are attached, form a monocyclic, saturated 5- or 6-membered
heterocycloalkyl ring which may contain a second ring heteroatom
selected from NH and O, and which may be substituted on a ring
carbon atom with oxo or hydroxy; and G.sup.2 is methyl or the group
--CR.sup.8AR.sup.8B--OH, --CH.sub.2--NR.sup.9R.sup.10 or
--C(.dbd.O)--NR.sup.11R.sup.12, wherein R.sup.8A and R.sup.8B are
independently hydrogen or methyl; R.sup.9 is hydrogen; R.sup.10 is
hydrogen, (C.sub.1-C.sub.4)-alkyl, acetyl, cyclopropyl, cyclobutyl
or 2-oxopyrrolidin-3-yl; wherein said (C.sub.1-C.sub.4)-alkyl is
optionally substituted with hydroxy or aminocarbonyl; R.sup.11 is
hydrogen or methyl; R.sup.12 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
cyclopropyl, cyclobutyl or 2-oxopyrrolidin-3-yl, wherein said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy; or
R.sup.9 and R.sup.10, or R.sup.11 and R.sup.12, respectively, are
joined and, taken together with the nitrogen atom to which they are
attached, form a monocyclic, saturated 4- to 6-membered
heterocycloalkyl ring which may contain a second ring heteroatom
selected from N(R.sup.13), O and S(O).sub.2, and which may be
substituted on ring carbon atoms with up to three substituents
independently selected from the group consisting of fluoro, methyl,
oxo, hydroxy, amino and aminocarbonyl, and wherein R.sup.13 is
hydrogen, formyl or acetyl, or a pharmaceutically acceptable salt,
a hydrate, or a solvate thereof.
28: The compound of claim 25, wherein: R.sup.1 is methyl; R.sup.2
is methoxy; G.sup.1 is the group --CH.sub.2--OR.sup.3, wherein
R.sup.3 is (C.sub.1-C.sub.4)-alkyl optionally substituted with
hydroxy, amino or aminocarbonyl; and G.sup.2 is the group
--CH.sub.2--NR.sup.9R.sup.10 or --C(.dbd.O)--NR.sup.11R.sup.12,
wherein R.sup.9 is hydrogen, R.sup.10 is 2-oxopyrrolidin-3-yl, or
R.sup.9 and R.sup.10 are joined and, taken together with the
nitrogen atom to which they are attached, form a piperazin-1-yl,
3-oxopiperazin-1-yl or 4-acetylpiperazin-1-yl ring; R.sup.11 is
hydrogen; R.sup.12 is 2-oxopyrrolidin-3-yl; or R.sup.11 and
R.sup.12 are joined and, taken together with the nitrogen atom to
which they are attached, form a 3-hydroxyazetidin-1-yl,
4-hydroxypiperidin-1-yl or 3-oxopiperazin-1-yl ring, or a
pharmaceutically acceptable salt, a hydrate, or a solvate thereof.
Description
[0001] This invention relates to novel substituted
5-(1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
derivatives having protein tyrosine kinase inhibitory activities,
to processes for the preparation of such compounds, to
pharmaceutical compositions containing such compounds, and to the
use of such compounds or compositions for treating proliferative
disorders, in particular cancer and tumor diseases.
[0002] Cancer is a leading cause of death worldwide and accounted
for 7.6 million deaths (around 13% of all deaths) in 2008. Deaths
from cancer are projected to continue to rise worldwide to over 11
million in 2030 (WHO source, Fact Sheet No. 297, February
2011).
[0003] There are many ways how cancers can arise which is one of
the reasons why their therapy is difficult. One way that
transformation of cells can occur is following a genetic
alteration. The completion of the human genome project showed
genomic instability and heterogeneity of human cancer genes. Recent
strategies to identify these genetic alterations sped up the
process of cancer-gene discovery. Gene abnormality can, for
instance, lead to the overexpression of proteins, and hence to a
nonphysiological activation of these proteins. One family of
proteins from which a number of oncoproteins derive are tyrosine
kinases and in particular receptor tyrosine kinases (RTKs). In the
past two decades, numerous avenues of research have demonstrated
the importance of RTK-mediated signalling in adverse cell growth
leading to cancer. In recent years, promising results have been
achieved in the clinic with selective small-molecule inhibitors of
tyrosine kinases as a new class of anti-tumorigenic agents [Swinney
and Anthony, Nature Rev. Drug Disc. 10 (7), 507-519 (2011)].
[0004] Fibroblast growth factors (FGFs) and their receptors (FGFRs)
form part of a unique and diverse signalling system which plays a
key role in a variety of biological processes which encompass
various aspects of embryonic development and adult pathophysiology
[Itoh and Ornitz, J. Biochem. 149 (2), 121-130 (2011)]. In a
spatio-temporal manner, FGFs stimulate through FGFR binding a wide
range of cellular functions including migration, proliferation,
differentiation, and survival.
[0005] The FGF family comprises 18 secreted polypeptidic growth
factors that bind to four highly conserved receptor tyrosine
kinases (FGFR-1 to -4) expressed at the cell surface. In addition,
FGFR-5 can bind to FGFs but does not have a kinase domain, and
therefore is devoid of intracellular signalling. The specificity of
the ligand/receptor interaction is enhanced by a number of
transcriptional and translational processes which give rise to
multiple isoforms by alternative transcriptional initiation,
alternative splicing, and C-terminal truncations. Various heparan
sulfate proteoglycans (e.g. syndecans) can be part of the FGF/FGFR
complex and strongly influence the ability of FGFs to induce
signalling responses [Polanska et al., Developmental Dynamics 238
(2), 277-293 (2009)]. FGFRs are cell surface receptors consisting
of three extracellular immunoglobulin-like domains, a single-pass
transmembrane domain, and an intracellular dimerized tyrosine
kinase domain. Binding of FGF bring the intracellular kinases into
close proximity, enabling them to transphosphorylate each other.
Seven phosphorylation sites have been identified (e.g., in FGFR-1
Tyr463, Tyr583, Tyr585, Tyr653, Tyr654, Tyr730, and Tyr766).
[0006] Some of these phosphotyrosine groups act as docking sites
for downstream signalling molecules which themselves may also be
directly phosphorylated by FGFR, leading to the activation of
multiple signal transduction pathways. Thus, the MAPK signalling
cascade is implicated in cell growth and differentiation, the
PI3K/Akt signalling cascade is involved in cell survival and cell
fate determination, while the PI3K and PKC signalling cascades have
a function in the control of cell polarity. Several feedback
inhibitors of FGF signalling have now been identified and include
members of the Spry (Sprouty) and Sef (similar expression to FGF)
families. Additionally, in certain conditions, FGFR is released
from pre-Golgi membranes into the cytosol. The receptor and its
ligand, FGF-2, are co-transported into the nucleus by a mechanism
that involves importin, and are engaged in the CREB-binding protein
(CBP) complex, a common and essential transcriptional coactivator
that acts as a gene activation gating factor. Multiple correlations
between the immunohistochemical expression of FGF-2, FGFR-1 and
FGFR-2 and their cytoplasmic and nuclear tumor cell localizations
have been observed. For instance, in lung adenocarcinomas this
association is also found at the nuclear level, emphasizing an
active role of the complex at the nucleus [Korc and Friesel, Curr.
Cancer Drugs Targets 5, 639-651 (2009)].
[0007] FGFs are widely expressed in both developing and adult
tissues and play important roles in a variety of normal and
pathological processes, including tissue development, tissue
regeneration, angiogenesis, neoplastic transformation, cell
migration, cellular differentiation, and cell survival.
Additionally, FGFs as pro-angiogenic factors have also been
implicated in the emerging phenomenon of resistance to vascular
endothelial growth factor receptor-2 (VEGFR-2) inhibition [Bergers
and Hanahan, Nat. Rev. Cancer 8, 592-603 (2008)].
[0008] Recent oncogenomic profiles of signalling networks
demonstrated an important role for aberrant FGF signalling in the
emergence of some common human cancers [Wesche et al., Biochem. J.
437 (2), 199-213 (2011)]. Ligand-independent FGFR constitutive
signalling has been described in many human cancers, such as brain
cancer, head and neck cancer, gastric cancer and ovarian cancer.
FGFR-mutated forms as well as FGFR-intragenic translocations have
been identified in malignancies such as myeloproliferative
diseases. Interestingly, the same mutations discovered to be the
cause of many developmental disorders are also found in tumor cells
(e.g., the mutations found in achondroplasia and thanatophoric
dysplasia, which cause dimerization and thus constitutive
activation of FGFR-3, are also frequently found in bladder cancer).
A mutation that promotes dimerization is just one mechanism that
can increase ligand-independent signalling from FGFRs. Other
mutations located inside or outside of the kinase domain of FGFRs
can change the conformation of the domain giving rise to
permanently active kinases.
[0009] Amplification of the chromosomal region 8p11-12, the genomic
location of FGFR-1, is a common focal amplification in breast
cancer and occurs in approximately 10% of breast cancers,
predominantly in oestrogen receptor-positive cancers. FGFR-1
amplifications have also been reported in non-small cell lung
squamous carcinoma and are found at a low incidence in ovarian
cancer, bladder cancer and rhabdomyosarcoma. Similarly,
approximately 10% of gastric cancers show FGFR-2 amplification,
which is associated with poor prognosis, diffuse-type cancers.
Moreover, multiple single nucleotide polymorphisms (SNPs) located
in FGFR-1 to -4 were found to correlate with an increased risk of
developing selective cancers, or were reported to be associated
with poor prognosis (e.g., FGFR-4 G388R allele in breast cancer,
colon cancer and lung adenocarcinoma). The direct role of these
SNPs to promote cancer is still controversial.
[0010] In summary, a great number of in vitro and in vivo studies
have been performed that validate FGFR-1 to -4 as important cancer
targets, and comprehensive reviews have summarized these findings
[see, for example, Heinzle et al., Expert Opin. Ther. Targets 15
(7), 829-846 (2011); Wesche et al., Biochem. J. 437 (2), 199-213
(2011); Greulich and Pollock, Trends in Molecular Medicine 17 (5),
283-292 (2011); Haugsten et al., Mol. Cancer Res. 8 (11), 1439-1452
(2010)]. Several strategies have been followed to attenuate
aberrant FGFR-1 to -4 signalling in human tumors including blocking
antibodies and small-molecule inhibitors, amongst others. A number
of selective small-molecule FGFR inhibitors are currently in
clinical development, such as AZD-4547 (AstraZeneca) and BJG-398
(Novartis).
[0011] Notwithstanding the significant advancements that have
generally been achieved in cancer therapy in recent years, there is
a continuing need to identify new anti-cancer compounds with
improved properties, such as higher potency, greater selectivity,
reduced toxicity and/or better tolerability. Therefore, the
technical problem to be solved according to the present invention
may be seen in providing alternative compounds having inhibitory
activity on the FGFR kinases, thus offering new therapeutic options
for the treatment of FGFR-mediated diseases, in particular cancer
and other proliferative disorders.
[0012] Fused hetero-5,6-bicyclic kinase inhibitors bearing a 9- or
a 10-membered bicyclic heteroaryl substituent have been disclosed
in WO 2007/061737-A2 and WO 2005/097800-A1, respectively. These
compounds were stated to be useful for the treatment of cancer and
other diseases owing to their inhibitory action on the mTOR
(mammalian target of Rapamycin) and/or IGF-1R (type 1 insulin-like
growth factor receptor) kinases. Further hetero-5,6-bicyclic
template structures associated with the inhibition of kinases have
been described in, inter alia, WO 01/19828-A2, WO 2007/079164-A2
and WO 2010/051043-A1.
[0013] 4-Aminopyrrolo[2,1-f][1,2,4]triazine derivatives with
differing inhibition profiles against a number of protein kinases
have been disclosed in, inter alia, WO 00/71129-A1, WO
2007/056170-A2, WO 2007/061882-A2, WO 2007/064932-A2, WO
2009/136966-A1, and WO 2010/126960-A1.
[0014] In WO 2005/121147-A1, WO 2007/064883-A2 and WO
2007/064931-A2, 4-aminopyrrolo[2,1-f]-[1,2,4]triazine derivatives
containing a substituted diarylurea group in 5-position were
described as having FGFR-1 inhibiting activity. However, other
receptor tyrosine kinases, notably the VEGFR, PDGFR and Tie-2
kinases, are also significantly inhibited by this particular class
of compounds. As it was hypothesized that such multi-kinase
activity might lead to an augmentation of potential side effects
during treatment, it was the aim of the present invention to
identify new agents having an improved selectivity for the FGFR
kinases, thus providing new options for a more tolerable cancer
therapy.
[0015] Surprisingly, it has now been found that certain
4-aminopyrrolo[2,1-f][1,2,4]triazine derivatives bearing a
specifically substituted benzothiophen-2-yl residue in 5-position
exhibit potent and selective inhibition of FGFR kinases, notably of
the FGFR-1 and FGFR-3 kinases, which renders these compounds
particularly useful for the treatment of proliferative disorders,
such as cancer and tumor diseases.
[0016] Thus, in one aspect, the present invention relates to
6,7-disubstituted
5-(1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
derivatives of the general formula (I)
##STR00001##
wherein [0017] R.sup.1 is hydrogen, chloro, methyl or methoxy,
[0018] R.sup.2 is hydrogen or methoxy, [0019] with the proviso that
at least one of R.sup.1 and R.sup.2 is other than hydrogen, [0020]
G.sup.1 represents chloro, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, 5-membered aza-heteroaryl, or the
group --CH.sub.2--OR.sup.3, --CH.sub.2--NR.sup.4R.sup.5 or
--C(.dbd.O)--NR.sup.4R.sup.6, wherein [0021] R.sup.3 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or phenyl,
wherein [0022] (i) said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, (C.sub.1-C.sub.4)-alkoxy,
hydroxycarbonyl, (C.sub.1-C.sub.4)-alkoxycarbonyl, amino,
aminocarbonyl, mono(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, [0023]
and [0024] (ii) said (C.sub.3-C.sub.6)-cycloalkyl is optionally
substituted with one or two substituents independently selected
from the group consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and
amino, [0025] and [0026] (iii) said phenyl is optionally
substituted with one or two substituents independently selected
from the group consisting of fluoro, chloro, bromo, cyano,
trifluoromethyl, trifluoromethoxy, (C.sub.1-C.sub.4)-alkyl and
(C.sub.1-C.sub.4)-alkoxy, [0027] R.sup.4 is hydrogen or
(C.sub.1-C.sub.4)-alkyl, [0028] R.sup.5 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkylcarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or 4- to 6-membered heterocycloalkyl,
wherein [0029] (i) said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, (C.sub.1-C.sub.4)-alkoxy,
hydroxycarbonyl, (C.sub.1-C.sub.4)-alkoxycarbonyl, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
(C.sub.3-C.sub.6)-cycloalkyl, [0030] and [0031] (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, [0032] and [0033]
(iii) said 4- to 6-membered heterocycloalkyl is optionally
substituted with one or two substituents independently selected
from the group consisting of (C.sub.1-C.sub.4)alkyl, hydroxy, oxo
and amino, [0034] R.sup.6 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.3-C.sub.6)-cycloalkyl or 4- to 6-membered heterocycloalkyl,
wherein [0035] (i) said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, (C.sub.1-C.sub.4)-alkoxy,
hydroxycarbonyl, (C.sub.1-C.sub.4)-alkoxycarbonyl, amino,
aminocarbonyl, mono(C.sub.1-C.sub.4)-alkylaminocarbonyl,
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
(C.sub.3-C.sub.6)-cycloalkyl, [0036] and [0037] (ii) said
(C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with one or
two substituents independently selected from the group consisting
of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, [0038] and [0039]
(iii) said 4- to 6-membered heterocycloalkyl is optionally
substituted with one or two substituents independently selected
from the group consisting of (C.sub.1-C.sub.4)alkyl, hydroxy, oxo
and amino, [0040] or [0041] R.sup.4 and R.sup.5, or R.sup.4 and
R.sup.6, respectively, are joined and, taken together with the
nitrogen atom to which they are attached, form a monocyclic,
saturated 4- to 7-membered heterocycloalkyl ring which may contain
a second ring heteroatom selected from N(R.sup.7) and O, and which
may be substituted on ring carbon atoms with one or two
substituents independently selected from the group consisting of
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein [0042] R.sup.7 is hydrogen, (C.sub.1-C.sub.4)-alkyl, formyl
or (C.sub.1-C.sub.4)-alkylcarbonyl, and [0043] G.sup.2 represents
chloro, cyano, (C.sub.1-C.sub.4)-alkyl, or the group
--CR.sup.8AR.sup.8B--OH, --CH.sub.2--NR.sup.9R.sup.10,
--C(.dbd.O)--NR.sup.11R.sup.12 or --CH.sub.2--OR.sup.15, wherein
[0044] R.sup.8A and R.sup.8B are independently selected from the
group consisting of hydrogen, (C.sub.1-C.sub.4)-alkyl, cyclopropyl
and cyclobutyl, [0045] R.sup.9 is hydrogen or
(C.sub.1-C.sub.4)-alkyl, [0046] R.sup.10 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkylcarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or 4- to 6-membered heterocycloalkyl,
wherein [0047] (i) said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, amino, aminocarbonyl,
mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl, [0048] and [0049] (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, [0050]
and [0051] (iii) said 4- to 6-membered heterocycloalkyl is
optionally substituted with one or two substituents independently
selected from the group consisting of (C.sub.1-C.sub.4)alkyl,
hydroxy, oxo and amino, [0052] R.sup.11 is hydrogen or
(C.sub.1-C.sub.4)-alkyl, [0053] R.sup.12 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or 4- to
6-membered heterocycloalkyl, wherein [0054] (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
amino, aminocarbonyl, mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl or
di-(C.sub.1-C.sub.4)-alkylaminocarbonyl, [0055] and [0056] (ii)
said (C.sub.3-C.sub.6)-cycloalkyl is optionally substituted with
one or two substituents independently selected from the group
consisting of (C.sub.1-C.sub.4)-alkyl, hydroxy and amino, [0057]
and [0058] (iii) said 4- to 6-membered heterocycloalkyl is
optionally substituted with one or two substituents independently
selected from the group consisting of (C.sub.1-C.sub.4)alkyl,
hydroxy, oxo and amino, [0059] or [0060] R.sup.9 and R.sup.10, or
R.sup.11 and R.sup.12, respectively, are joined and, taken together
with the nitrogen atom to which they are attached, form a
monocyclic, saturated 4- to 7-membered heterocycloalkyl ring which
may contain a second ring heteroatom selected from N(R.sup.13), O,
S and S(O).sub.2, and which may be substituted on ring carbon atoms
with up to three substituents independently selected from the group
consisting of fluoro, (C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino
and aminocarbonyl, and wherein [0061] R.sup.13 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl, formyl or
(C.sub.1-C.sub.4)-alkylcarbonyl, [0062] and [0063] R.sup.15 is
(C.sub.1-C.sub.4)-alkyl, with the proviso that G.sup.1 is not
chloro when G.sup.2 is chloro or cyano.
[0064] The compounds according to this invention can also be
present in the form of their salts, solvates and/or solvates of the
salts.
[0065] Compounds according to the invention are the compounds of
the formula (I) and their salts, solvates and solvates of the
salts, the compounds included in the formula (I) of the formulae
(I-A), (I-B), (IC), (I-D) and (I-E) mentioned in the following and
their salts, solvates and solvates of the salts, and the compounds
included in the formula (I) and mentioned in the following as
process products and/or embodiment examples and their salts,
solvates and solvates of the salts, where the compounds included in
the formula (I) and mentioned in the following are not already
salts, solvates and solvates of the salts.
[0066] Salts for the purposes of the present invention are
preferably pharmaceutically acceptable salts of the compounds
according to the invention (for example, see S. M. Berge et al.,
"Pharmaceutical Salts", J. Pharm. Sci. 1977, 66, 1-19). Salts which
are not themselves suitable for pharmaceutical uses but can be
used, for example, for isolation or purification of the compounds
according to the invention are also included.
[0067] Pharmaceutically acceptable salts include acid addition
salts of mineral acids, carboxylic acids and sulfonic acids, for
example salts of hydrochloric acid, hydrobromic acid, sulfuric
acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid,
benzenesulfonic acid, toluenesulfonic acid, naphthalenedisulfonic
acid, formic acid, acetic acid, trifluoroacetic acid, propionic
acid, lactic acid, tartaric acid, malic acid, citric acid, fumaric
acid, maleic acid, and benzoic acid.
[0068] Pharmaceutically acceptable salts also include salts of
customary bases, such as for example and preferably alkali metal
salts (for example sodium and potassium salts), alkaline earth
metal salts (for example calcium and magnesium salts), and ammonium
salts derived from ammonia or organic amines, such as
illustratively and preferably ethylamine, diethylamine,
triethylamine, N,N-diisopropylethylamine, monoethanolamine,
diethanolamine, triethanolamine, dimethylaminoethanol,
diethylaminoethanol, procaine, dicyclohexylamine, dibenzylamine,
N-methylmorpholine, N-methylpiperidine, arginine, lysine, and
1,2-ethylenediamine.
[0069] Solvates in the context of the invention are designated as
those forms of the compounds according to the invention which form
a complex in the solid or liquid state by stoichiometric
coordination with solvent molecules. Hydrates are a specific form
of solvates, in which the coordination takes place with water.
Hydrates are preferred solvates in the context of the present
invention.
[0070] The compounds of this invention may, either by nature of
asymmetric centers or by restricted rotation, be present in the
form of isomers (enantiomers, diastereomers). Any isomer may be
present in which the asymmetric center is in the (R)-, (S)-, or
(R,S)-configuration.
[0071] It will also be appreciated that when two or more asymmetric
centers are present in the compounds of the invention, several
diastereomers and enantiomers of the exemplified structures will
often be possible, and that pure diastereomers and pure enantiomers
represent preferred embodiments. It is intended that pure
stereoisomers, pure diastereomers, pure enantiomers, and mixtures
thereof, are within the scope of the invention.
[0072] Geometric isomers by nature of substituents about a double
bond or a ring may be present in cis (=Z-) or trans (=E-) form, and
both isomeric forms are encompassed within the scope of this
invention.
[0073] All isomers, whether separated, pure, partially pure, or in
racemic mixture, of the compounds of this invention are encompassed
within the scope of this invention. The purification of said
isomers and the separation of said isomeric mixtures may be
accomplished by standard techniques known in the art. For example,
diastereomeric mixtures can be separated into the individual
isomers by chromatographic processes or crystallization, and
racemates can be separated into the respective enantiomers either
by chromatographic processes on chiral phases or by resolution.
[0074] In addition, all possible tautomeric forms of the compounds
described above are included according to the present
invention.
[0075] The present invention also encompasses all suitable isotopic
variants of the compounds according to the invention. An isotopic
variant of a compound according to the invention is understood to
mean a compound in which at least one atom within the compound
according to the invention has been exchanged for another atom of
the same atomic number, but with a different atomic mass than the
atomic mass which usually or predominantly occurs in nature.
Examples of isotopes which can be incorporated into a compound
according to the invention are those of hydrogen, carbon, nitrogen,
oxygen, fluorine, chlorine, bromine and iodine, such as .sup.2H
(deuterium), .sup.3H (tritium), .sup.13C, .sup.14C, .sup.15N,
.sup.17O, .sup.18O, .sup.18F, .sup.36Cl, .sup.82Br, .sup.123I,
.sup.124I, .sup.129I and .sup.131I. Particular isotopic variants of
a compound according to the invention, especially those in which
one or more radioactive isotopes have been incorporated, may be
beneficial, for example, for the examination of the mechanism of
action or of the active compound distribution in the body. Due to
comparatively easy preparability and detectability, especially
compounds labelled with .sup.3H or .sup.14C isotopes are suitable
for this purpose. In addition, the incorporation of isotopes, for
example of deuterium, can lead to particular therapeutic benefits
as a consequence of greater metabolic stability of the compound,
for example an extension of the half-life in the body or a
reduction in the active dose required. Such modifications of the
compounds according to the invention may therefore in some cases
also constitute a preferred embodiment of the present invention.
Isotopic variants of the compounds according to the invention can
be prepared by processes known to those skilled in the art, for
example by the methods described below and the methods described in
the working examples, by using corresponding isotopic modifications
of the particular reagents and/or starting compounds therein.
[0076] In the context of the present invention, the substituents
and residues have the following meaning, unless specified
otherwise:
[0077] (C.sub.1-C.sub.4)-Alkyl in the context of the invention
represents a straight-chain or branched alkyl radical having 1 to 4
carbon atoms. There may be mentioned by way of example and
preferably: methyl, ethyl, n-propyl, isopropyl, n-butyl, iso-butyl,
sec-butyl, and tert-butyl.
[0078] (C.sub.1-C.sub.4-Alkoxy in the context of the invention
represents a straight-chain or branched alkoxy radical having 1 to
4 carbon atoms. There may be mentioned by way of example and
preferably: methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy,
iso-butoxy, sec-butoxy, and tert-butoxy.
[0079] Mono-(C.sub.1-C.sub.4)-alkylamino in the context of the
invention represents an amino group with a straight-chain or
branched alkyl substituent which contains 1 to 4 carbon atoms.
There may be mentioned by way of example and preferably:
methylamino, ethylamino, n-propylamino, isopropylamino,
n-butylamino, and tert-butylamino.
[0080] Di-(C.sub.1-C.sub.4)-alkylamino in the context of the
invention represents an amino group with two identical or different
straight-chain or branched alkyl substituents which each contain 1
to 4 carbon atoms. There may be mentioned by way of example and
preferably: N,N-dimethylamino, N,N-diethylamino,
N-ethyl-N-methylamino, N-methyl-N-n-propylamino,
N-isopropyl-N-methylamino, N-isopropyl-N-n-propylamino,
N,N-diisopropylamino, N-n-butyl-N-methylamino, and
N-tert-butyl-N-methylamino.
[0081] (C.sub.1-C.sub.4)-Alkylcarbonyl in the context of the
invention represents a straight-chain or branched alkyl radical
having 1 to 4 carbon atoms which is bonded to the rest of the
molecule via a carbonyl group [--C(.dbd.O)--]. There may be
mentioned by way of example and preferably: acetyl, propionyl,
n-butyryl, iso-butyryl, n-pentanoyl, and pivaloyl.
[0082] (C.sub.1-C.sub.4-Alkoxycarbonyl in the context of the
invention represents a straight-chain or branched alkoxy radical
having 1 to 4 carbon atoms which is bonded to the rest of the
molecule via a carbonyl group [--C(.dbd.O)--]. There may be
mentioned by way of example and preferably: methoxycarbonyl,
ethoxycarbonyl, n-propoxycarbonyl, isopropoxycarbonyl,
n-butoxycarbonyl, and tert-butoxycarbonyl.
[0083] Mono-(C.sub.1-C.sub.4)-alkylaminocarbonyl in the context of
the invention represents an amino group which is bonded to the rest
of the molecule via a carbonyl group [--C(.dbd.O)--] and which has
a straight-chain or branched alkyl substituent having 1 to 4 carbon
atoms. There may be mentioned by way of example and preferably:
methylaminocarbonyl, ethylaminocarbonyl, n-propylaminocarbonyl,
isopropylaminocarbonyl, n-butylaminocarbonyl, and
tert-butylaminocarbonyl.
[0084] Di-(C.sub.1-C.sub.4)-alkylaminocarbonyl in the context of
the invention represents an amino group which is bonded to the rest
of the molecule via a carbonyl group [--C(.dbd.O)--] and which has
two identical or different straight-chain or branched alkyl
substituents having in each case 1 to 4 carbon atoms. There may be
mentioned by way of example and preferably:
N,N-dimethylaminocarbonyl, N,N-diethylaminocarbonyl,
N-ethyl-N-methylaminocarbonyl, N-methyl-N-n-propylaminocarbonyl,
N-isopropyl-N-methylaminocarbonyl, N,N-diisopropylaminocarbonyl,
N-n-butyl-N-methylaminocarbonyl, and
N-tert-butyl-N-methylaminocarbonyl.
[0085] (C.sub.3-C.sub.6)-Cycloalkyl in the context of the invention
represents a monocyclic, saturated carbocycle having 3 to 6 ring
carbon atoms. There may be mentioned by way of example:
cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. Preferred are
cyclopropyl and cyclobutyl.
[0086] 4- to 7-membered heterocycloalkyl and 4- to 6-membered
heterocycloalkyl in the context of the invention represent a
monocyclic, saturated heterocycle with 4 to 7 or, respectively, 4
to 6 ring atoms in total, which contains one or two identical or
different ring heteroatoms from the series N, O, S and S(O).sub.2,
and which can be bonded via a ring carbon atom or via a ring
nitrogen atom (if present). 4- to 6-membered heterocycloalkyl
containing one ring nitrogen atom and optionally one further ring
heteroatom from the series N, O or S(O).sub.2 is preferred. 5- or
6-membered heterocycloalkyl containing one ring nitrogen atom and
optionally one further ring heteroatom from the series N or O is
particularly preferred. There may be mentioned by way of example:
azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl,
imidazolidinyl, tetrahydrofuranyl, thiolanyl, 1,1-dioxidothiolanyl,
1,2-oxazolidinyl, 1,3-oxazolidinyl, 1,3-thiazolidinyl, piperidinyl,
piperazinyl, tetrahydropyranyl, tetrahydrothiopyranyl,
1,3-dioxanyl, 1,4-dioxanyl, 1,2-oxazinanyl, morpholinyl,
thiomorpholinyl, 1,1-dioxidothiomorpholinyl, azepanyl,
1,4-diazepanyl, and 1,4-oxazepanyl. Preferred are azetidinyl,
pyrrolidinyl, pyrazolidinyl, imidazolidinyl, 1,2-oxazolidinyl,
1,3-oxazolidinyl, piperidinyl, piperazinyl, 1,2-oxazinanyl,
morpholinyl, and thiomorpholinyl. Particularly preferred are
pyrrolidinyl, piperidinyl, piperazinyl, and morpholinyl.
[0087] 5-membered aza-heteroaryl in the context of the invention
represents an aromatic heterocyclic radical (heteroaromatic) having
5 ring atoms in total, which contains at least one ring nitrogen
atom and optionally one or two further ring heteroatoms from the
series N, O and/or S, and which is bonded via a ring carbon atom or
optionally via a ring nitrogen atom (when allowed by valency).
5-membered aza-heteroaryl containing one ring nitrogen atom and one
or two further ring heteroatoms from the series N and/or O is
preferred. There may be mentioned by way of example: pyrrolyl,
pyrazolyl, imidazolyl, oxazolyl, thiazolyl, isoxazolyl,
isothiazolyl, triazolyl, oxadiazolyl, and thiadiazolyl. Preferred
are pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, and
oxadiazolyl.
[0088] An oxo substituent in the context of the invention
represents an oxygen atom, which is bonded to a carbon atom via a
double bond.
[0089] In the context of the present invention, for all the
radicals which occur several times, the meaning thereof is
independent of each other. If radicals in the compounds according
to the invention are substituted, the radicals can be mono- or
polysubstituted, unless specified otherwise. Substitution by one or
by two or three identical or different substituents is preferred.
Substitution by one or by two identical or different substituents
is particularly preferred.
[0090] In a preferred embodiment, the present invention relates to
compounds of general formula (I), wherein [0091] R.sup.1 is chloro,
methyl or methoxy, [0092] R.sup.2 is hydrogen or methoxy, [0093]
G.sup.1 represents chloro, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl or 5-membered aza-heteroaryl
selected from the group consisting of pyrazolyl, imidazolyl,
oxazolyl, isoxazolyl and oxadiazolyl, or represents the group
--CH.sub.2--OR.sup.3 or --CH.sub.2--NR.sup.4R.sup.5, wherein [0094]
R.sup.3 is hydrogen, (C.sub.1-C.sub.4)-alkyl or
(C.sub.3-C.sub.6)-cycloalkyl, [0095] wherein said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
(C.sub.1-C.sub.4)-alkoxy, hydroxycarbonyl,
(C.sub.1-C.sub.4)-alkoxycarbonyl, amino, aminocarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or up to three fluoro atoms, [0096]
R.sup.4 is hydrogen or (C.sub.1-C.sub.4)-alkyl, [0097] R.sup.5 is
hydrogen, (C.sub.1-C.sub.4)-alkyl, (C.sub.1-C.sub.4)-alkylcarbonyl,
(C.sub.3-C.sub.6)-cycloalkyl or 5- or 6-membered heterocycloalkyl,
wherein [0098] (i) said (C.sub.1-C.sub.4)-alkyl is optionally
substituted with hydroxy, hydroxycarbonyl or
(C.sub.3-C.sub.6)-cycloalkyl, [0099] and [0100] (ii) said 5- or
6-membered heterocycloalkyl is optionally substituted with oxo,
[0101] or [0102] R.sup.4 and R.sup.5 are joined and, taken together
with the nitrogen atom to which they are attached, form a
monocyclic, saturated 4- to 6-membered heterocycloalkyl ring which
may contain a second ring heteroatom selected from N(R.sup.7) and
O, and which may be substituted on a ring carbon atom with oxo or
hydroxy, and wherein [0103] R.sup.7 is hydrogen or
(C.sub.1-C.sub.4)-alkyl, and [0104] G.sup.2 represents chloro,
cyano, (C.sub.1-C.sub.4)-alkyl, or the group
--CR.sup.8AR.sup.8B--OH, --CH.sub.2--NR.sup.9R.sup.10,
--C(.dbd.O)--NR.sup.11R.sup.12 or --CH.sub.2--OR.sup.15, wherein
[0105] R.sup.8A and R.sup.8B are independently selected from the
group consisting of hydrogen, (C.sub.1-C.sub.4)-alkyl and
cyclopropyl, [0106] R.sup.9 is hydrogen or methyl, [0107] R.sup.10
is hydrogen, (C.sub.1-C.sub.4)-alkyl,
(C.sub.1-C.sub.4)-alkylcarbonyl, (C.sub.3-C.sub.6)-cycloalkyl or 5-
or 6-membered heterocycloalkyl, wherein [0108] (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy or
aminocarbonyl, [0109] and [0110] (ii) said 5- or 6-membered
heterocycloalkyl is optionally substituted with oxo, [0111]
R.sup.11 is hydrogen or methyl, [0112] R.sup.12 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, (C.sub.3-C.sub.6)-cycloalkyl or 5- or
6-membered heterocycloalkyl, wherein [0113] (i) said
(C.sub.1-C.sub.4)-alkyl is optionally substituted with hydroxy,
[0114] and [0115] (ii) said 5- or 6-membered heterocycloalkyl is
optionally substituted with oxo, [0116] or [0117] R.sup.9 and
R.sup.10, or R.sup.11 and R.sup.12, respectively, are joined and,
taken together with the nitrogen atom to which they are attached,
form a monocyclic, saturated 4- to 6-membered heterocycloalkyl ring
which may contain a second ring heteroatom selected from
N(R.sup.13), O, S and S(O).sub.2, and which may be substituted on
ring carbon atoms with up to three substituents independently
selected from the group consisting of fluoro,
(C.sub.1-C.sub.4)-alkyl, oxo, hydroxy, amino and aminocarbonyl, and
wherein [0118] R.sup.3 is hydrogen, (C.sub.1-C.sub.4)-alkyl,
cyclopropyl, cyclobutyl, formyl or (C.sub.1-C.sub.4)-alkylcarbonyl,
[0119] and [0120] R.sup.15 is methyl or ethyl, with the proviso
that G.sup.1 is not chloro when G.sup.2 is chloro or cyano.
[0121] In a particularly preferred embodiment, the present
invention relates to compounds of general formula (I), wherein
[0122] R.sup.1 is methyl, [0123] R.sup.2 is methoxy, [0124] G.sup.1
represents methyl, oxazol-5-yl or the group --CH.sub.2--OR.sup.3 or
--CH.sub.2--NR.sup.4R.sup.5, wherein [0125] R.sup.3 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, cyclopropyl or cyclobutyl, [0126] wherein
said (C.sub.1-C.sub.4)-alkyl is optionally substituted with
hydroxy, methoxy, ethoxy, hydroxycarbonyl, methoxycarbonyl,
ethoxycarbonyl, amino, aminocarbonyl, cyclopropyl, cyclobutyl or up
to three fluoro atoms, [0127] R.sup.4 is hydrogen, methyl or ethyl,
[0128] R.sup.5 is hydrogen, (C.sub.1-C.sub.4)-alkyl, acetyl,
cyclopropyl, cyclobutyl or 2-oxopyrrolidin-3-yl, [0129] wherein
said (C.sub.1-C.sub.4)-alkyl is optionally substituted with
hydroxy, hydroxycarbonyl, cyclopropyl or cyclobutyl, or [0130]
R.sup.4 and R.sup.5 are joined and, taken together with the
nitrogen atom to which they are attached, form a monocyclic,
saturated 5- or 6-membered heterocycloalkyl ring which may contain
a second ring heteroatom selected from NH and O, and which may be
substituted on a ring carbon atom with oxo or hydroxy, and [0131]
G.sup.2 represents methyl or the group --CR.sup.8AR.sup.8B--OH,
--CH.sub.2--NR.sup.9R.sup.10 or --C(.dbd.O)--NR.sup.11R.sup.12,
wherein [0132] R.sup.8A and R.sup.8B are independently hydrogen or
methyl, [0133] R.sup.9 is hydrogen, [0134] R.sup.10 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, acetyl, cyclopropyl, cyclobutyl or
2-oxopyrrolidin-3-yl, [0135] wherein said (C.sub.1-C.sub.4)-alkyl
is optionally substituted with hydroxy or aminocarbonyl, [0136]
R.sup.11 is hydrogen or methyl, [0137] R.sup.12 is hydrogen,
(C.sub.1-C.sub.4)-alkyl, cyclopropyl, cyclobutyl or
2-oxopyrrolidin-3-yl, [0138] wherein said (C.sub.1-C.sub.4)-alkyl
is optionally substituted with hydroxy, [0139] or [0140] R.sup.9
and R.sup.10, or R.sup.11 and R.sup.12, respectively, are joined
and, taken together with the nitrogen atom to which they are
attached, form a monocyclic, saturated 4- to 6-membered
heterocycloalkyl ring which may contain a second ring heteroatom
selected from N(R.sup.13), O and S(O).sub.2, and which may be
substituted on ring carbon atoms with up to three substituents
independently selected from the group consisting of fluoro, methyl,
oxo, hydroxy, amino and aminocarbonyl, and wherein [0141] R.sup.13
is hydrogen, formyl or acetyl.
[0142] In a distinct embodiment, the present invention relates to
compounds of general formula (I), wherein [0143] R.sup.1 is methyl,
[0144] and [0145] R.sup.2 is methoxy.
[0146] In a further distinct embodiment, the present invention
relates to compounds of general formula (I), wherein [0147] G.sup.1
represents the group --CH.sub.2--OR.sup.3, wherein [0148] R.sup.3
is hydrogen or (C.sub.1-C.sub.4)-alkyl optionally substituted with
hydroxy, methoxy, amino, aminocarbonyl or up to three fluoro
atoms.
[0149] In another distinct embodiment, the present invention
relates to compounds of general formula (I), wherein [0150] G.sup.1
represents the group --CH.sub.2--NR.sup.4R.sup.5, wherein [0151]
R.sup.4 is hydrogen or methyl, [0152] R.sup.5 is
(C.sub.1-C.sub.4)-alkyl, acetyl, cyclopropyl, cyclobutyl or
2-oxopyrrolidin-3-yl, [0153] wherein said (C.sub.1-C.sub.4)-alkyl
is optionally substituted with hydroxy, [0154] or [0155] R.sup.4
and R.sup.5 are joined and, taken together with the nitrogen atom
to which they are attached, form a monocyclic, saturated 5- or
6-membered heterocycloalkyl ring which may contain a second ring
heteroatom selected from NH and O, and which may be substituted on
a ring carbon atom with oxo or hydroxy.
[0156] In another distinct embodiment, the present invention
relates to compounds of general formula (I), wherein [0157] G.sup.2
represents the group --CH.sub.2--NR.sup.9R.sup.10, wherein [0158]
R.sup.9 is hydrogen, [0159] R.sup.10 is acetyl or
2-oxopyrrolidin-3-yl, [0160] or [0161] R.sup.9 and R.sup.10 are
joined and, taken together with the nitrogen atom to which they are
attached, form a monocyclic, saturated 5- or 6-membered
heterocycloalkyl ring which may contain a second ring heteroatom
selected from N(R.sup.13) and O, and which may be substituted on
ring carbon atoms with up to two substituents independently
selected from the group consisting of methyl, oxo, hydroxy and
amino, and wherein [0162] R.sup.13 is hydrogen, formyl or
acetyl.
[0163] In yet another distinct embodiment, the present invention
relates to compounds of general formula (I), wherein [0164] G.sup.2
represents the group --C(.dbd.O)--NR.sup.11R.sup.12, wherein [0165]
R.sup.11 is hydrogen, [0166] R.sup.12 is (C.sub.1-C.sub.4)-alkyl or
2-oxopyrrolidin-3-yl, [0167] wherein said (C.sub.1-C.sub.4)-alkyl
is optionally substituted with hydroxy, [0168] or [0169] R.sup.11
and R.sup.12 are joined and, taken together with the nitrogen atom
to which they are attached, form a monocyclic, saturated 4- to
6-membered heterocycloalkyl ring which may contain a second ring
heteroatom selected from NH and O, and which may be substituted on
a ring carbon atom with oxo or hydroxy.
[0170] In an especially preferred embodiment, the present invention
relates to compounds of general formula (I), wherein [0171] R.sup.1
is methyl, [0172] R.sup.2 is methoxy, [0173] G.sup.1 represents the
group --CH.sub.2--OR.sup.3, wherein [0174] R.sup.3 is
(C.sub.1-C.sub.4)-alkyl optionally substituted with hydroxy, amino
or aminocarbonyl, and [0175] G.sup.2 represents the group
--CH.sub.2--NR.sup.9R.sup.10 or --C(.dbd.O)--NR.sup.11R.sup.12,
wherein [0176] R.sup.9 is hydrogen, [0177] R.sup.10 is
2-oxopyrrolidin-3-yl, [0178] or [0179] R.sup.9 and R.sup.10 are
joined and, taken together with the nitrogen atom to which they are
attached, form a piperazin-1-yl, 3-oxopiperazin-1-yl or
4-acetylpiperazin-1-yl ring, [0180] R.sup.11 is hydrogen, [0181]
R.sup.12 is 2-oxopyrrolidin-3-yl, [0182] or [0183] R.sup.11 and
R.sup.12 are joined and, taken together with the nitrogen atom to
which they are attached, form a 3-hydroxyazetidin-1-yl,
4-hydroxypiperidin-1-yl or 3-oxopiperazin-1-yl ring.
[0184] The definitions of residues indicated specifically in the
respective combinations or preferred combinations of residues are
also replaced as desired by definitions of residues of other
combinations, irrespective of the particular combinations indicated
for the residues. Combinations of two or more of the abovementioned
preferred ranges are particularly preferred.
[0185] The compounds of the general formula (I) can be prepared by
various synthetic routes which are primarily governed by the nature
of the particular G.sup.1 and G.sup.2 groups chosen (see
definitions above).
[0186] Thus, in another embodiment, the present invention relates
to a process for preparing the compounds of the general formula
(I), characterized in that
[A] a 6-substituted 4-aminopyrrolo[2,1-f][1,2,4]triazine of formula
(II)
##STR00002## [0187] wherein R.sup.3 has the meaning described
above, [0188] is at first reacted with formaldehyde and an amine of
formula (III)
[0188] ##STR00003## [0189] wherein R.sup.9 and R.sup.10 have the
meanings described above, [0190] in the presence of an acid to give
a compound of formula (IV)
[0190] ##STR00004## [0191] wherein R.sup.3, R.sup.9 and R.sup.10
have the meanings described above, [0192] then brominated to a
compound of formula (V)
[0192] ##STR00005## [0193] wherein R.sup.3, R.sup.9 and R.sup.10
have the meanings described above, [0194] and subsequently coupled
with a benzothiophen-2-yl boronate of formula (VI)
[0194] ##STR00006## [0195] wherein R.sup.1 and R.sup.2 have the
meanings described above, [0196] and [0197] R.sup.14 represents
hydrogen or (C.sub.1-C.sub.4)-alkyl, or both R.sup.14 residues are
linked together to form a --(CH.sub.2).sub.2--,
--C(CH.sub.3).sub.2--C(CH.sub.3).sub.2--, --(CH.sub.2).sub.3--,
--CH.sub.2--C(CH.sub.3).sub.2--CH.sub.2-- or
--C(.dbd.O)--CH.sub.2--N(CH.sub.3)--CH.sub.2--C(.dbd.O)-- bridge,
[0198] in the presence of a palladium catalyst and a base to yield
the target compound of formula (I-A)
[0198] ##STR00007## [0199] wherein R.sup.1, R.sup.2, R.sup.3,
R.sup.9 and R.sup.10 have the meanings described above, or [B] a
6-substituted 4-aminopyrrolo[2,1-f][1,2,4]triazine of formula
(II)
[0199] ##STR00008## [0200] wherein R.sup.3 has the meaning
described above, [0201] is at first formylated with
N,N-dimethylformamide in the presence of phosphoryl chloride to an
aldehyde of formula (VII)
[0201] ##STR00009## [0202] wherein R.sup.3 has the meaning
described above, [0203] then brominated to a compound of formula
(VIII)
[0203] ##STR00010## [0204] wherein R.sup.3 has the meaning
described above, [0205] and subsequently coupled with a
benzothiophen-2-yl boronate of formula (VI)
[0205] ##STR00011## [0206] wherein R.sup.1, R.sup.2 and R.sup.14
have the meanings described above, [0207] in the presence of a
palladium catalyst and a base to give a compound of formula
(IX)
[0207] ##STR00012## [0208] wherein R.sup.1, R.sup.2 and R.sup.3
have the meanings described above, [0209] which then is either
[B-1] reacted with an amine of formula (III)
[0209] ##STR00013## [0210] wherein R.sup.9 and R.sup.10 have the
meanings described above, [0211] in the presence of an acid and a
reducing agent to yield the target compound of formula (I-A)
[0211] ##STR00014## [0212] wherein R.sup.1, R.sup.2, R.sup.3,
R.sup.9 and R.sup.10 have the meanings described above, or [B-2]
oxidized to a carboxylic acid of formula (X)
[0212] ##STR00015## [0213] wherein R.sup.1, R.sup.2 and R.sup.3
have the meanings described above, [0214] and finally coupled with
an amine of formula (XI)
[0214] ##STR00016## [0215] wherein R.sup.11 and R.sup.12 have the
meanings described above, [0216] in the presence of a condensing
agent to yield the target compound of formula (I-B)
[0216] ##STR00017## [0217] wherein R.sup.1, R.sup.2, R.sup.3,
R.sup.11 and R.sup.12 have the meanings described above, or [C] a
6-substituted 4-amino-5-bromopyrrolo[2,1-f][1,2,4]triazine of
formula (XII)
[0217] ##STR00018## [0218] is at first coupled with a
benzothiophen-2-yl boronate of formula (VI)
[0218] ##STR00019## [0219] wherein R.sup.1, R.sup.2 and R.sup.14
have the meanings described above, [0220] in the presence of a
palladium catalyst and a base to give a compound of formula
(XIII)
[0220] ##STR00020## [0221] wherein R.sup.1 and R.sup.2 have the
meanings described above, [0222] and then reacted with formaldehyde
and an amine of formula (III)
[0222] ##STR00021## [0223] wherein R.sup.9 and R.sup.10 have the
meanings described above, [0224] in the presence of an acid to
yield the compound of formula (I-C)
[0224] ##STR00022## [0225] wherein R.sup.1, R.sup.2, R.sup.9 and
R.sup.10 have the meanings described above, [0226] which
subsequently is either [C-1] oxidized to an aldehyde of formula
(XIV)
[0226] ##STR00023## [0227] wherein R.sup.1, R.sup.2, R.sup.9 and
R.sup.10 have the meanings described above, [0228] and treated with
an amine of formula (XV)
[0228] ##STR00024## [0229] wherein R.sup.4 and R.sup.5 have the
meanings described above, [0230] in the presence of an acid and a
reducing agent to yield the target compound of formula (I-D)
[0230] ##STR00025## [0231] wherein R.sup.1, R.sup.2, R.sup.4,
R.sup.5, R.sup.9 and R.sup.10 have the meanings described above, or
[C-2] converted into the corresponding 6-(halomethyl) derivative of
formula (XVI)
[0231] ##STR00026## [0232] wherein R.sup.1, R.sup.2, R.sup.9 and
R.sup.10 have the meanings described above, [0233] and [0234] X is
chloro, bromo or iodo, [0235] and treated with an alcohol of
formula (XVII)
[0235] R.sup.3A--OH (XVII), [0236] wherein R.sup.3A has the meaning
of R.sup.3 as described above except for hydrogen, [0237] in the
presence of a base to yield the target compound of formula
(I-E)
[0237] ##STR00027## [0238] wherein R.sup.1, R.sup.2, R.sup.3A,
R.sup.9 and R.sup.10 have the meanings described above, optionally
followed, where appropriate, by (i) separating the compounds of
formula (I) thus obtained into their respective enantiomers and/or
diastereomers, preferably using chromatographic methods, and/or
(ii) converting the compounds of formula (I) into their respective
hydrates, solvates, salts and/or hydrates or solvates of the salts
by treatment with the corresponding solvents and/or acids or
bases.
[0239] The compounds of the formulae (I-A), (I-B), (I-C), (I-D) and
(I-E), which can be prepared by the processes described above, each
represent a particular subset of the compounds of the general
formula (I).
[0240] Process steps [A] (II).fwdarw.(IV) and [C]
(XIII).fwdarw.(I-C), representing Mannich-type aminomethylation
reactions, are carried out in the usual way by treating the
respective starting compound with a mixture of aqueous formaldehyde
and amine component (III) in the presence of an acid catalyst such
as formic acid or acetic acid. Preferably, acetic acid is used both
as catalyst and solvent. The reaction is usually performed at a
temperature ranging from +20.degree. C. to +80.degree. C.
[0241] As the brominating agent for process steps [A]
(IV).fwdarw.(V) and [B] (VII).fwdarw.(VIII), preferably
N-bromosuccinimide (NBS), 1,3-dibromo-5,5-dimethylhydantoin (DBDMH)
or elemental bromine are used. The reactions are generally carried
out in an inert solvent, such as dichloromethane, chloroform,
tetrahydrofuran, acetonitrile or N,N-dimethylformamide (DMF),
within a temperature range from -78.degree. C. to +20.degree.
C.
[0242] The coupling reactions [A] (V)+(VI).fwdarw.(I-A), [B]
(VIII)+(VI).fwdarw.(IX) and [C] (XII)+(VI).fwdarw.(XIII)
["Suzuki-Miyaura coupling" ] are generally carried out in an inert
solvent with the aid of a palladium catalyst and an aqueous base.
Palladium catalysts suitable for this purpose include, for example,
palladium(II) acetate, palladium(II) chloride,
bis(triphenylphosphine)palladium(II) chloride,
bis(acetonitrile)palladium(II) chloride,
[1,1'-bis(diphenylphosphino)ferrocene]palladium(II) chloride,
tetrakis(triphenylphosphine)palladium(0),
bis(dibenzylideneacetone)palladium(0), and
tris(dibenzylideneacetone)dipalladium(0), optionally in combination
with other phosphine ligands such as, for example,
2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (X-Phos),
2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (S-Phos),
4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (Xantphos), or
4-(di-tert-butylphosphino)-N,N-dimethylaniline. Also, palladium
pre-catalysts from which the catalytically active species is
generated under the reaction conditions, such as
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine, can be used [see, for example, S. Kotha
et al., Tetrahedron 58, 9633-9695 (2002); T. E. Barder et al., J.
Am. Chem. Soc. 127 (13), 4685-4696 (2005); S. L. Buchwald et al.,
J. Am. Chem. Soc. 132 (40), 14073-14075 (2010), and further
references cited therein].
[0243] Suitable bases for these coupling reactions are in
particular alkali carbonates, such as sodium, potassium or caesium
carbonate, alkali phosphates, such as sodium or potassium
phosphate, or alkali fluorides, such as potassium or caesium
fluoride. Usually, these bases are employed as aqueous solutions.
The reactions are carried out in organic solvents that are inert
under the reaction conditions. Preferably, water-miscible organic
solvents, such as 1,2-dimethoxyethane, tetrahydrofuran,
1,4-dioxane, acetonitrile, N,N-dimethylformamide (DMF) or
dimethylsulfoxide (DMSO), are employed but other inert solvents,
such as dichloromethane or toluene, may also be used.
[0244] Process step [B] (II).fwdarw.(VII) ["Vilsmeier-Haack
formylation" ] is carried out in the usual manner by treating the
pyrrolotriazine (II) in N,N-dimethylformamide (DMF) solvent with
phosphoryl chloride. The reaction is usually performed at a
temperature from 0.degree. C. to +80.degree. C.
[0245] Reducing agents suitable for the reductive amination
reactions [B-1] (IX)+(III).fwdarw.(I-A) and
[C-1](XIV)+(XV).fwdarw.(I-D) are customary alkali borohydrides,
such as lithium borohydride, sodium borohydride, potassium
borohydride, sodium cyanoborohydride or sodium
triacetoxyborohydride. The transformations are generally carried
out in the presence of an acid, preferably acetic acid, in an
alcohol or ether solvent, such as methanol, ethanol, isopropanol,
tetrahydrofuran or 1,4-dioxane, within a temperature range from
0.degree. C. to +80.degree. C., depending on the reactivity of the
amine components (III) and (XV), respectively, and/or the
particular borohydride used.
[0246] For the oxidation reaction in process step [B-2]
(IX).fwdarw.(X), oxidation with sodium chlorite in the presence of
a hypochlorite scavenger such as 2-methyl-2-butene represents the
method of choice [cf H. W. Pinnick et al., Tetrahedron 37,
2091-2096 (1981); A. Raach and O. Reiser, J. Prakt. Chem. 342 (6),
605-608 (2000), and references cited therein]. The reaction is
usually carried out in a tetrahydrofuran/water mixture at a
temperature between 0.degree. C. and ambient temperature.
[0247] Condensing agents suitable for process step [B-2]
(X)+(XI).fwdarw.(I-B) [amide formation] include, for example,
carbodiimides such as N,N'-diethyl-, N,N'-dipropyl-,
N,N'-diisopropyl-, N,N'-dicyclohexylcarbodiimide (DCC) or
N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC), phosgene
derivatives such as N,N'-carbonyldiimidazole (CDI) or isobutyl
chloroformate, .alpha.-chloroenamines such as
1-chloro-2-methyl-1-dimethylamino-1-propene, phosphorus compounds
such as propanephosphonic anhydride, diethyl cyanophosphonate,
bis(2-oxo-3-oxazolidinyl)phosphoryl chloride,
benzotriazol-1-yloxy-tris(dimethylamino)phosphonium
hexafluorophosphate (BOP) or
benzotriazol-1-yloxy-tris(pyrrolidino)phosphonium
hexafluorophosphate (PyBOP), and uronium compounds such as
O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TBTU),
O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate (HBTU),
2-(2-oxo-1-(2H)-pyridyl)-1,1,3,3-tetramethyluronium
tetrafluoroborate (TPTU),
O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate (HATU) or
O-(1H-6-chlorobenzotriazol-1-yl)-1,1,3,3-tetramethyluronium
tetrafluoroborate (TCTU), if appropriate in combination with
further auxiliaries, such as 1-hydroxybenzotriazole (HOBt) or
N-hydroxysuccinimide (HOSu), and/or bases such as alkali
carbonates, for example sodium or potassium carbonate, or organic
amine bases, such as triethylamine, N-methylpiperidine,
N-methylmorpholine (NMM), N,N-diisopropylethylamine (DIPEA),
pyridine or 4-N,N-dimethylaminopyridine (DMAP). Preference is given
to using O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate (HATU) or
O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TBTU) in combination with
N,N-diisopropylethylamine (DIPEA) and optionally
1-hydroxybenzotriazole (HOBt).
[0248] Inert solvents for process step [B-2] (X)+(XI).fwdarw.(I-B)
are, for example, ethers such as diethyl ether, tert-butyl methyl
ether, tetrahydrofuran, 1,4-dioxane or 1,2-dimethoxyethane,
hydrocarbons such as benzene, toluene, xylene, hexane or
cyclohexane, halogenated hydrocarbons such as dichloromethane,
trichloromethane, carbon tetrachloride, 1,2-dichloroethane,
trichloroethylene or chlorobenzene, or other solvents such as
acetone, acetonitrile, ethyl acetate, pyridine, dimethylsulfoxide
(DMSO), N,N-dimethylformamide (DMF), N,N'-dimethylpropylene urea
(DMPU) or N-methylpyrrolidinone (NMP). It is also possible to use
mixtures of these solvents. Preference is given to using
dichloromethane, tetrahydrofuran, N,N-dimethylformamide or mixtures
thereof. The reactions are generally carried out at a temperature
ranging from 0.degree. C. to +60.degree. C., preferably at
+10.degree. C. to +40.degree. C.
[0249] Oxidizing agents that are capable of converting the primary
alcohol (I-C) into the aldehyde (XIV) (process [C-1]) under mild
conditions include
1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one
("Dess-Martin periodinane"), 2,2,6,6-tetramethylpiperidin-1-oxyl
(TEMPO) in combination with secondary oxidants such as
iodosobenzene-I,I-diacetate or sodium hypochlorite, and
dimethylsulfoxide (DMSO)-based oxidation systems such as
DMSO/trifluoroacetic anhydride or
DMSO/N,N'-dicyclohexylcarbodiimide (DCC). Preference is given to
1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one. The
reaction is generally carried out in an inert solvent, preferably
using dichloromethane.
[0250] For the hydroxy-to-halogen transformation in process step
[C-2] (I-C).fwdarw.(XVI), various standard methods and reagents
that are well known in the art may be employed. Reagents of choice
are thionyl chloride [for X=Cl],
tetrabromomethane/triphenylphosphine [for X=Br], and
iodine/triphenylphosphine [for X=I]. The preparation of
6-(chloromethyl) derivatives (XVI) [X=Cl] is preferred for reasons
of convenience of work-up and compound stability.
[0251] Bases suitable for the process step [C-2]
(XVI)+(XVII).fwdarw.(I-E) [ether formation] are in particular
alkali carbonates such as lithium, sodium, potassium or caesium
carbonate, alkali acetates such as sodium or potassium acetate, or
customary tertiary amine bases such as triethylamine,
N-methylmorpholine, N-methylpiperidine, N,N-diisopropylethylamine
or pyridine. Preference is given to N,N-diisopropylethylamine
(DIPEA). The reaction (XVI)+(XVII).fwdarw.(I-E) is performed in an
inert solvent, such as tetrahydrofuran, or without solvent, using
an excess of alcohol (XVII), at a temperature ranging from
+20.degree. C. to +200.degree. C., preferably at +50.degree. C. to
+150.degree. C. Advantageously, the conversion is carried out by
means of a microwave reactor device.
[0252] The reaction sequence (I-C).fwdarw.(XVI).fwdarw.(I-E) may be
carried out in two separate steps, i.e. with isolation and
purification of the intermediate compound (XVI), or it may be
performed using a one-pot procedure, i.e. employing the crude
intermediate (XVI) as obtained in the preparation reaction.
[0253] In cases where a primary or secondary amine moiety forms
part of the G.sup.1 or G.sup.2 group in the target compounds of
formula (I), it may sometimes be appropriate in the preparation
reactions described above to use a protected derivative of this
amine as reaction component instead of the free amine. For this
purpose, conventional temporary amino-protecting groups, such as
acyl groups (e.g., acetyl or trifluoroacetyl) or carbamate-type
protecting groups (e.g., a Boc-, Cbz- or Fmoc-group), may be
employed. A Boc (tert-butoxycarbonyl) group is preferably used.
Similarly, a hydroxy function being part of the G.sup.1 or G.sup.2
group may temporarily be blocked in precursor compounds and process
intermediates, for example as a tetrahydropyranyl (THP) ether or as
a silyl ether derivative, such as a trimethylsilyl or
tert-butyldimethylsilyl ether.
[0254] These protecting groups may then be cleaved off
concomitantly during aqueous work-up and purification procedures,
or they are removed in a subsequent, separate reaction step using
standard methods well known in the art. The preparation of such
protected intermediates from the corresponding free amines or
alcohols is likewise readily accomplished following general
procedures described in the literature [see, for example, T. W.
Greene and P. Wuts, Protective Groups in Organic Synthesis, Wiley,
New York, 1999].
[0255] Certain types of protected (i.e. acylated) amine derivatives
exert significant FGFR-inhibiting activity by their own.
Accordingly, such compounds are also encompassed by the general
formula (I) as defined above.
[0256] The preparation of the compounds of the invention may be
illustrated by means of the following reaction schemes:
##STR00028##
##STR00029##
##STR00030##
##STR00031##
[0257] The 6-substituted 4-aminopyrrolo[2,1-f][1,2,4]triazines of
formula (II) can, for example, be prepared by two different routes
which are depicted in Scheme 5 below. In the first route,
4-amino-6-cyanopyrrolo[2,1-f][1,2,4]triazine (XVIII) is converted
into the ester (XIX) by acid-mediated alcoholysis and then reduced
to the 6-(hydroxymethyl) compound (IIa) [R.sup.3 in (II)=H] using
lithium triethylborohydride. Standard transformation into the
corresponding 6-(halomethyl)pyrrolotriazine, such as the chloro
compound (XX), followed by treatment with an alcohol of formula
(XVII) in the presence of a base readily provides the ether
derivatives of formula (IIb) [R.sup.3 in (II).noteq.H]. The
preparation of the starting compound (XVIII) has been described
previously [see Int. Pat. Appl. WO 2007/064883 (Intermediate
AX/Step 3)].
[0258] The second route starts from the protected
1-amino-4-bromo-2-cyanopyrrole (XXI) [preparation given in Int.
Pat. Appl. WO 2007/064883 (Intermediate AAE, Step 3)].
Deprotonation of the urethane nitrogen, subsequent metalation in
4-position and reaction with formaldehyde affords the
4-(hydroxymethyl) derivative (XXII). Treatment with hydrogen
chloride followed by addition of alcohol (XVII) and condensation
with formamidine using a one-pot procedure then provides the target
compound of formula (IIb). This route is especially suited for the
preparation of alkyl ether derivatives [R.sup.3A in
(IIb)=(C.sub.1-C.sub.4)-alkyl] in that the alcohol reactant (XVII)
may also serve as the reaction solvent in these cases.
##STR00032##
[0259] The 4-amino-5-bromopyrrolo[2,1-f][1,2,4]triazine derivative
of formula (XII) is readily available from
4-amino-6-(hydroxymethyl)pyrrolo[2,1-f][1,2,4]triazine (IIa) (cf.
Scheme 5) by initial 5,7-dibromination using
1,3-dibromo-5,5-dimethylhydantoin and subsequent selective
7-debromination via a halogen-metal exchange with n-butyllithium
followed by methanol quenching (see Scheme 6 below).
##STR00033##
[0260] The benzothiophen-2-yl boronates of formula (VI) can
conveniently be prepared starting from the substituted thiophenol
derivatives of formula (XXIV) (see Scheme 7 below). Alkylation with
bromoacetal (XXV) and subsequent polyphosphoric acid-mediated
cyclization provides the benzothiophene intermediates of formula
(XXVII) which are then metalated in 2-position and reacted with a
trialkyl borate. Alkaline work-up affords the free
(benzothiophen-2-yl)boronic acids of formula (VIa) which may be
transformed, if desired, into cyclic boronates, e.g. so-called MIDA
boronates of formula (VIb), by standard procedures known in the art
[see, for example, D. M. Knapp et al., J. Am. Chem. Soc. 131 (20),
6961-6963 (2009)].
##STR00034##
[cf P. A. Ple and L. J. Marnett, J. Heterocyclic Chem. 25 (4),
1271-1272 (1988); A. Venturelli et al., J. Med. Chem. 50 (23),
5644-5654 (2007)].
[0261] The compounds of the formulae (III), (XI), (XV), (XVII),
(XXIV) and (XXV) are either commercially available, known from the
literature, or can be prepared from readily available starting
materials by adaptation of standard methods described in the
literature. Detailed procedures and literature references for
preparing the starting materials can also be found in the
Experimental Part in the section on the preparation of the starting
materials and intermediates.
[0262] The preparation of further subgroups of the compounds of
general formula (I) is illustrated in the following reaction
schemes 8-14. The required pyrrolotriazine precursors can be
readily synthesized by customary methods well known in the art, and
further synthetic transformations, in most instances, follow the
preparation routes that have been outlined in the process section
above, using similar types of reactions, such as, for example,
bromination, boronate coupling, aminomethylation, reductive
amination, oxidation and/or ether or amide forming reactions.
Further details are provided in the Experimental Part on the
preparation of the exemplary embodiments and their respective
precursor compounds.
##STR00035## ##STR00036##
[R.sup.8A, R.sup.8B=(C.sub.1-C.sub.4)-alkyl, cyclopropyl or
cyclobutyl].
##STR00037##
[R.sup.8A=(C.sub.1-C.sub.4)-alkyl, cyclopropyl or cyclobutyl; X=Cl,
Br or I].
##STR00038##
[R=(C.sub.1-C.sub.4)-alkyl].
##STR00039##
[R=chloro or (C.sub.1-C.sub.4)-alkyl; for the preparation of the
starting material (XXVIII), see Int. Pat. Appl. WO 2007/064883 and
WO 2007/056170, respectively].
##STR00040##
[for the preparation of the starting compound (XXIX), see Scheme 14
below].
##STR00041## ##STR00042##
##STR00043## ##STR00044##
[0263] The compounds of the present invention have valuable
pharmacological properties and can be used for the prevention and
treatment of disorders in humans and other mammals.
[0264] The compounds of the present invention are potent inhibitors
of the activity or expression of receptor tyrosine kinases,
particularly of the FGFR kinases, and most notably of the FGFR-1
and FGFR-3 kinases. Accordingly, in another embodiment, the present
invention provides a method of treating disorders relating to or
mediated by the activity of FGFR kinases in a patient in need of
such treatment, comprising administering to the patient an
effective amount of a compound of formula (I) as defined above. In
certain embodiments, the disorders relating to the activity of FGFR
kinases are proliferative disorders, in particular cancer and tumor
diseases.
[0265] In the context of the present invention, the term
"treatment" or "treating" includes inhibiting, delaying, relieving,
mitigating, arresting, reducing, or causing the regression of a
disease, disorder, condition, or state, the development and/or
progression thereof, and/or the symptoms thereof. The term
"prevention" or "preventing" includes reducing the risk of having,
contracting, or experiencing a disease, disorder, condition, or
state, the development and/or progression thereof, and/or the
symptoms thereof. The term prevention includes prophylaxis.
Treatment or prevention of a disorder, disease, condition, or state
may be partial or complete.
[0266] The term "proliferative disorder" includes disorders
involving the undesired or uncontrolled proliferation of a cell.
The compounds of the present invention can be utilized to prevent,
inhibit, block, reduce, decrease, control, etc., cell proliferation
and/or cell division, and/or produce apoptosis. This method
comprises administering to a subject in need thereof, including a
mammal, including a human, an amount of a compound of this
invention, or a pharmaceutically acceptable salt, isomer,
polymorph, metabolite, hydrate or solvate thereof which is
effective to treat or prevent the disorder.
[0267] Throughout this document, for the sake of simplicity, the
use of singular language is given preference over plural language,
but is generally meant to include the plural language if not
otherwise stated. For example, the expression "A method of treating
a disease in a patient, comprising administering to a patient an
effective amount of a compound of formula (I)" is meant to include
the simultaneous treatment of more than one disease as well as the
administration of more than one compound of formula (I).
[0268] Proliferative disorders that can be treated and/or prevented
with the compounds of the present invention particularly include,
but are not limited to, the group of cancer and tumor diseases.
These are understood as meaning, in particular, the following
diseases, but without being limited to them: mammary carcinomas and
mammary tumors (ductal and lobular forms, also in situ), tumors of
the respiratory tract (small cell and non-small cell lung
carcinoma, parvicellular and non-parvicellular carcinoma, bronchial
carcinoma, bronchial adenoma, pleuropulmonary blastoma), cerebral
tumors (e.g. of the brain stem and of the hypothalamus,
astrocytoma, glioblastoma, medulloblastoma, ependymoma, and
neuro-ectodermal and pineal tumors), tumors of the digestive organs
(oesophagus, stomach, gall bladder, small intestine, large
intestine, rectum, anus), liver tumors (inter alia hepatocellular
carcinoma, cholangiocellular carcinoma and mixed hepatocellular and
cholangiocellular carcinoma), tumors of the head and neck region
(larynx, hypopharynx, nasopharynx, oropharynx, lips and oral
cavity), skin tumors (squamous epithelial carcinoma, Kaposi
sarcoma, malignant melanoma, Merkel cell skin cancer and
non-melanomatous skin cancer), tumors of soft tissue (inter alia
soft tissue sarcomas, osteosarcomas, malignant fibrous
histiocytomas, lymphosarcomas and rhabdomyosarcomas), tumors of the
eyes (inter alia intraocular melanoma, uveal melanoma and
retinoblastoma), tumors of the endocrine and exocrine glands (e.g.
thyroid and parathyroid glands, pancreas and salivary gland),
tumors of the urinary tract (tumors of the bladder, penis, kidney,
renal pelvis and ureter), tumors of the reproductive organs
(carcinomas of the endometrium, cervix, ovary, vagina, vulva and
uterus in women, and carcinomas of the prostate and testicles in
men), as well as distant metastases thereof. These disorders also
include proliferative blood diseases in solid form and as
circulating blood cells, such as lymphomas, leukaemias and
myeloproliferative diseases, e.g. acute myeloid, acute
lymphoblastic, chronic lymphocytic, chronic myelogenic and hairy
cell leukaemia, and AIDS-related lymphomas, Hodgkin's lymphomas,
nonHodgkin's lymphomas, cutaneous T-cell lymphomas, Burkitt's
lymphomas, and lymphomas in the central nervous system.
[0269] Due to their activity and selectivity profile, the compounds
of the present invention are believed to be particularly suitable
for the treatment of breast (mammary), lung, stomach (gastric),
bladder and ovary cancer and tumor diseases. Furthermore, the
compounds of the present invention may be especially suited for the
prevention or suppression of tumor metastasis in general.
[0270] Other proliferative disorders that can be treated and/or
prevented with the compounds and methods of the present invention
include psoriasis, keloids and other hyperplasias affecting the
skin, bullous disorders associated with subepidermal blister
formation including bullous pemphigoid, erythema multiforme and
dermatitis herpetiformis, fibrotic disorders such as lung fibrosis,
atherosclerosis, restenosis and hepatic cirrhosis, renal diseases
including mesangial cell proliferative disorders, glomerulopathies,
glomerulonephritis, diabetic nephropathy, malignant nephrosclerosis
and polycystic kidney disease, benign prostate hyperplasia (BPH),
angiogenic or blood vessel proliferative disorders, and thrombotic
microangiopathy syndromes.
[0271] The compounds of the present invention are also useful for
the treatment and/or prevention of ophthalmological diseases such
as, for example, age-related macular degeneration (AMD), dry
macular degeneration, ischemic retinal vein occlusion, diabetic
macula edema, diabetic retinopathy, retinopathy of prematurity, and
other retinopathies.
[0272] Other conditions that may be treated and/or prevented by
administering a compound of the present invention include
gynaecological diseases such as endometriosis, myoma and ovarian
cysts, metabolic disorders related to adipogenesis, bile
metabolism, phosphate metabolism, calcium metabolism and/or bone
mineralization, skeletal disorders such as, for example, dwarfism,
achondrodysplasia and Pfeiffer syndrome, cartilage diseases such as
osteoarthritis and polyarthritis, rheumatoid arthritis, calvities,
and transplant rejection.
[0273] The diseases mentioned above have been well characterized in
humans, but also exist with a comparable aetiology in other
mammals, and can be treated in those with the compounds and methods
of the present invention.
[0274] Thus, the present invention further relates to the use of
the compounds according to the invention for the treatment and/or
prevention of disorders, especially of the aforementioned
disorders.
[0275] The present invention further relates to the use of the
compounds according to the invention for preparing a pharmaceutical
composition for the treatment and/or prevention of disorders,
especially of the aforementioned disorders.
[0276] The present invention further relates to the use of the
compounds according to the invention in a method for the treatment
and/or prevention of disorders, especially of the aforementioned
disorders.
[0277] The present invention further relates to a method for the
treatment and/or prevention of disorders, especially of the
aforementioned disorders, by using an effective amount of at least
one of the compounds according to the invention.
[0278] Compounds of the present invention may be administered as
the sole pharmaceutical agent or in combination with one or more
additional therapeutic agents as long as this combination does not
lead to undesirable and/or unacceptable side effects. Such
combination therapy includes administration of a single
pharmaceutical dosage formulation which contains a compound of
formula (I), as defined above, and one or more additional
therapeutic agents, as well as administration of a compound of
formula (I) and each additional therapeutic agent in its own
separate pharmaceutical dosage formulation. For example, a compound
of formula (I) and a therapeutic agent may be administered to the
patient together in a single (fixed) oral dosage composition such
as a tablet or capsule, or each agent may be administered in
separate dosage formulations.
[0279] Where separate dosage formulations are used, the compound of
formula (I) and one or more additional therapeutic agents may be
administered at essentially the same time (i.e., concurrently) or
at separately staggered times (i.e., sequentially).
[0280] In particular, the compounds of the present invention may be
used in fixed or separate combination with other anti-cancer agents
such as alkylating agents, anti-metabolites, plant-derived
anti-tumor agents, hormonal therapy agents, topoisomerase
inhibitors, tubulin inhibitors, kinase inhibitors, targeted drugs,
antibodies, antibody-drug conjugates (ADCs), immunologicals,
biological response modifiers, anti-angiogenic compounds, and other
anti-proliferative, cytostatic and/or cytotoxic substances. In this
regard, the following is a non-limiting list of examples of
secondary agents that may be used in combination with the compounds
of the present invention:
[0281] Abarelix, abiraterone, aclarubicin, afatinib, aflibercept,
aldesleukin, alemtuzumab, alitretinoin, alpharadin, altretamine,
aminoglutethimide, amonafide, amrubicin, amsacrine, anastrozole,
andromustine, arglabin, asparaginase, axitinib, 5-azacitidine,
basiliximab, belotecan, bendamustine, bevacizumab, bexarotene,
bicalutamide, bisantrene, bleomycin, bortezomib, bosutinib,
brivanib alaninate, buserelin, busulfan, cabazitaxel, CAL-101,
calcium folinate, calcium levofolinate, camptothecin, capecitabine,
carboplatin, carmofur, carmustine, catumaxomab, cediranib,
celmoleukin, cetuximab, chlorambucil, chlormadinone, chlormethine,
cidofovir, cisplatin, cladribine, clodronic acid, clofarabine,
combretastatin, crisantaspase, crizotinib, cyclophosphamide,
cyproterone, cytarabine, dacarbazine, dactinomycin, darbepoetin
alfa, darinaparsin, dasatinib, daunorubicin, decitabine, degarelix,
denileukin diftitox, denosumab, deslorelin, dibrospidium chloride,
docetaxel, dovitinib, doxifluridine, doxorubicin, dutasteride,
eculizumab, edrecolomab, eflornithine, elliptinium acetate,
eltrombopag, endostatin, enocitabine, epimbicin, epirubicin,
epitiostanol, epoetin alfa, epoetin beta, epothilone, eptaplatin,
eribulin, erlotinib, estradiol, estramustine, etoposide,
everolimus, exatecan, exemestane, exisulind, fadrozole,
fenretinide, filgrastim, finasteride, flavopiridol, fludarabine,
5-fluorouracil, fluoxymesterone, flutamide, foretinib, formestane,
fotemustine, fulvestrant, ganirelix, gefitinib, gemcitabine,
gemtuzumab, gimatecan, gimeracil, glufosfamide, glutoxim,
goserelin, histrelin, hydroxyurea, ibandronic acid, ibritumomab
tiuxetan, idarubicin, ifosfamide, imatinib, imiquimod, improsulfan,
intedanib, interferon alpha, interferon alpha-2a, interferon
alpha-2b, interferon beta, interferon gamma, interleukin-2,
ipilimumab, irinotecan, ixabepilone, lanreotide, lapatinib,
lasofoxifene, lenalidomide, lenograstim, lentinan, lenvatinib,
lestaurtinib, letrozole, leuprorelin, levamisole, linifanib,
linsitinib, lisuride, lobaplatin, lomustine, lonidamine,
lurtotecan, mafosfamide, mapatumumab, masitinib, masoprocol,
medroxyprogesterone, megestrol, melarsoprol, melphalan,
mepitiostane, mercaptopurine, methotrexate, methyl aminolevulinate,
methyltestosterone, mifamurtide, mifepristone, miltefosine,
miriplatin, mitobronitol, mitoguazone, mitolactol, mitomycin,
mitotane, mitoxantrone, molgramostim, motesanib, nandrolone,
nedaplatin, nelarabine, neratinib, nilotinib, nilutamide,
nimotuzumab, nimustine, nitracrine, nolatrexed, ofatumumab,
oprelvekin, oxaliplatin, paclitaxel, palifermin, pamidronic acid,
panitumumab, pazopanib, pegaspargase, peg-epoetin beta,
pegfilgastrim, peg-interferon alpha-2b, pelitrexol, pemetrexed,
pemtumomab, pentostatin, peplomycin, perfosfamide, perifosine,
pertuzumab, picibanil, pirambicin, pirarubicin, plerixafor,
plicamycin, poliglusam, polyestradiol phosphate, ponatinib,
porfimer sodium, pralatrexate, prednimustine, procarbazine,
procodazole, PX-866, quinagolide, raloxifene, raltitrexed,
ranibizumab, ranimustine, razoxane, regorafenib, risedronic acid,
rituximab, romidepsin, romiplostim, rubitecan, saracatinib,
sargramostim, satraplatin, selumetinib, sipuleucelT, sirolimus,
sizofiran, sobuzoxane, sorafenib, streptozocin, sunitinib,
talaporfin, tamibarotene, tamoxifen, tandutinib, tasonermin,
teceleukin, tegafur, telatinib, temoporfin, temozolomide,
temsirolimus, teniposide, testolactone, testosterone, tetrofosmin,
thalidomide, thiotepa, thymalfasin, tioguanine, tipifarnib,
tivozanib, toceranib, tocilizumab, topotecan, toremifene,
tositumomab, trabectedin, trastuzumab, treosulfan, tretinoin,
triapine, trilostane, trimetrexate, triptorelin, trofosfamide,
ubenimex, valrubicin, vandetanib, vapreotide, varlitinib,
vatalanib, vemurafenib, vidarabine, vinblastine, vincristine,
vindesine, vinflunine, vinorelbine, volociximab, vorinostat,
zinostatin, zoledronic acid, and zorubicin.
[0282] Generally, the following aims may be pursued with the
combination of compounds of the present invention with other
anti-cancer agents: [0283] improved activity in slowing down the
growth of a tumor, in reducing its size or even in its complete
elimination compared with treatment with a single active compound;
[0284] possibility of employing the chemotherapeutics used in a
lower dosage than in monotherapy; [0285] possibility of a more
tolerable therapy with few side effects compared with individual
administration; [0286] possibility of treatment of a broader
spectrum of cancer and tumor diseases; [0287] achievement of a
higher rate of response to therapy; [0288] longer survival time of
the patient compared with standard therapy.
[0289] Thus, in a further embodiment, the present invention relates
to pharmaceutical compositions comprising at least one of the
compounds according to the invention and one or more additional
therapeutic agents for the treatment and/or prevention of
disorders, especially of the aforementioned disorders.
[0290] In cancer treatment, the compounds of the present invention
may also be employed in conjunction with radiation therapy and/or
surgical intervention.
[0291] Furthermore, the compounds of formula (I) may be utilized,
as such or in compositions, in research and diagnostics, or as
analytical reference standards, and the like, which are well known
in the art. When the compounds of the present invention are
administered as pharmaceuticals, to humans and other mammals, they
can be given per se or as a pharmaceutical composition containing,
for example, 0.1% to 99.5% (more preferably, 0.5% to 90%) of active
ingredient in combination with one or more pharmaceutically
acceptable excipients.
[0292] Thus, in another aspect, the present invention relates to
pharmaceutical compositions comprising at least one of the
compounds according to the invention, conventionally together with
one or more inert, non-toxic, pharmaceutically suitable excipients,
and to the use thereof for the treatment and/or prevention of
disorders, especially of the aforementioned disorders.
[0293] The compounds according to the invention can act
systemically and/or locally. For this purpose, they can be
administered in a suitable way such as, for example, by the oral,
parenteral, pulmonary, nasal, lingual, sublingual, buccal, rectal,
dermal, transdermal, conjunctival, otic or topical route, or as an
implant or stent.
[0294] For these application routes, the compounds of the invention
can be administered in suitable application forms.
[0295] Suitable for oral administration are application forms which
function according to the prior art and deliver the compounds
according to the invention rapidly and/or in modified fashion, and
which contain the compounds according to the invention in
crystalline, amorphous and/or dissolved form, such as, for example,
tablets (uncoated or coated tablets, for example having enteric
coatings or coatings which are insoluble or dissolve with a delay
and control the release of the compound according to the
invention), tablets which disintegrate rapidly in the mouth, or
films/wafers, films/lyophilisates, capsules (e.g. hard or soft
gelatin capsules), sugar-coated tablets, granules, pellets,
powders, emulsions, suspensions, aerosols or solutions.
[0296] Parenteral application can be carried out with avoidance of
an absorption step (intravenously, intraarterially, intracardially,
intraspinally or intralumbarly) or with inclusion of an absorption
(intramuscularly, subcutaneously, intracutaneously, percutaneously
or intraperitoneally). Useful parenteral application forms include
injection and infusion preparations in the form of solutions,
suspensions, emulsions, lyophilisates and sterile powders.
[0297] Forms suitable for other application routes include, for
example, inhalatory pharmaceutical forms (e.g. powder inhalers,
nebulizers), nasal drops, solutions or sprays, tablets or capsules
to be administered lingually, sublingually or buccally (e.g.
troches, lozenges), suppositories, ear and eye preparations (e.g.
drops, ointments), vaginal capsules, aqueous suspensions (lotions,
shaking mixtures), lipophilic suspensions, ointments, creams,
milks, pastes, foams, dusting powders, transdermal therapeutic
systems (e.g. patches), implants and stents.
[0298] In a preferred embodiment, the pharmaceutical composition
comprising a compound of formula (I) as defined above is provided
in a form suitable for oral administration. In another preferred
embodiment, the pharmaceutical composition comprising a compound of
formula (I) as defined above is provided in a form suitable for
intravenous administration.
[0299] The compounds according to the invention can be converted
into the recited application forms in a manner known per se by
mixing with inert, non-toxic, pharmaceutically suitable excipients.
These excipients include, inter alia, carriers (e.g.
microcrystalline cellulose, lactose, mannitol), solvents (e.g.
liquid polyethylene glycols), emulsifiers (e.g. sodium dodecyl
sulfate), surfactants (e.g. polyoxysorbitan oleate), dispersants
(e.g. polyvinylpyrrolidone), synthetic and natural polymers (e.g.
albumin), stabilizers (e.g. antioxidants such as, for example,
ascorbic acid), colorants (e.g. inorganic pigments such as, for
example, iron oxides), and taste and/or odour masking agents.
[0300] A preferred dose of the compound of the present invention is
the maximum that a patient can tolerate and not develop serious
side effects. Illustratively, the compound of the present invention
may be administered parenterally at a dose of about 0.001 mg/kg to
about 1 mg/kg, preferably of about 0.01 mg/kg to about 0.5 mg/kg of
body weight. On oral administration, an exemplary dose range is
about 0.01 to 100 mg/kg, preferably about 0.01 to 20 mg/kg, and
more preferably about 0.1 to 10 mg/kg of body weight. Ranges
intermediate to the above-recited values are also intended to be
part of the invention.
[0301] Nevertheless, actual dosage levels and time course of
administration of the active ingredients in the pharmaceutical
compositions of the invention may be varied so as to obtain an
amount of the active ingredient which is effective to achieve the
desired therapeutic response for a particular patient, composition
and mode of administration, without being toxic to the patient. It
may therefore be necessary where appropriate to deviate from the
stated amounts, in particular as a function of age, gender, body
weight, diet and general health status of the patient, the
bioavailability and pharmacodynamic characteristics of the
particular compound and its mode and route of administration, the
time or interval over which administration takes place, the dose
regimen selected, the response of the individual patient to the
active ingredient, the specific disease involved, the degree of or
the involvement or severity of the disease, the kind of concurrent
treatment (i.e., the interaction of the compound of the invention
with other co-administered therapeutics), and other relevant
circumstances.
[0302] Thus, it may be satisfactory in some cases to manage with
less than the aforementioned minimum amount, whereas in other cases
the stated upper limit must be exceeded. Treatment can be initiated
with smaller dosages, which are less than the optimum dose of the
compound. Thereafter, the dosage may be increased by small
increments until the optimum effect under the circumstances is
reached. For convenience, the total daily dosage may be divided and
administered in individual portions spread over the day.
[0303] The following exemplary embodiments illustrate the
invention. The invention is not restricted to the examples.
[0304] The percentages in the following tests and examples are,
unless stated otherwise, by weight; parts are by weight. Solvent
ratios, dilution ratios and concentrations reported for
liquid/liquid solutions are each based on volume.
A. EXAMPLES
Abbreviations and Acronyms
[0305] Ac acetyl [0306] Ac.sub.2O acetic anhydride [0307] AcOH
acetic acid [0308] aq. aqueous (solution) [0309] Boc
tert-butoxycarbonyl [0310] br. broad (.sup.1H-NMR signal) [0311] Bu
butyl [0312] cat. catalytic [0313] conc. concentrated [0314] d
doublet (.sup.1H-NMR signal) [0315] DBDMH
1,3-dibromo-5,5-dimethylhydantoin [0316] DCI direct chemical
ionization (MS) [0317] DCM dichloromethane [0318] Dess-Martin
periodinane 1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one
[0319] DIPEA N,N-diisopropylethylamine [0320] DMF
N,N-dimethylformamide [0321] DMSO dimethylsulfoxide [0322] EI
electron impact ionization (MS) [0323] eq. equivalent(s) [0324] ESI
electro-spray ionization (MS) [0325] Et ethyl [0326] EtOAc ethyl
acetate [0327] GC-MS gas chromatography-coupled mass spectroscopy
[0328] h hour(s) [0329] Hal halogen [0330] .sup.1H-NMR proton
nuclear magnetic resonance spectroscopy [0331] HPLC high
performance liquid chromatography [0332] iPr isopropyl [0333] LC-MS
liquid chromatography-coupled mass spectroscopy [0334] Me methyl
[0335] MeOH methanol [0336] min minute(s) [0337] MS mass
spectroscopy [0338] m/z mass-to-charge ratio (MS) [0339] NBS
N-bromosuccinimide [0340] n-Bu n-butyl [0341] NCS
N-chlorosuccinimide [0342] of th. of theory (chemical yield) [0343]
Pd/C palladium on charcoal [0344] PdCl.sub.2(dppf)
[1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) [0345]
Pd(dba).sub.2 bis(dibenzylideneacetone)palladium [0346] Ph phenyl
[0347] PPA polyphosphoric acid [0348] q quartet (.sup.1H-NMR
signal) [0349] quant. quantitative (yield) [0350] rac racemic
[0351] R.sub.f TLC retention factor [0352] RP reverse phase (HPLC)
[0353] rt room temperature [0354] R.sub.t retention time (HPLC)
[0355] s singlet (.sup.1H-NMR signal) [0356] sat. saturated
(solution) [0357] t triplet (.sup.1H-NMR signal) [0358] TBAF
tetra-n-butylammonium fluoride [0359] TBDMS tert-butyldimethylsilyl
[0360] TBTU
N-[(1H-benzotriazol-1-yloxy)(dimethylamino)methylene]-N-methylmethanamini-
um tetrafluoroborate [0361] tBu tert-butyl [0362] tert tertiary
[0363] TFA trifluoroacetic acid [0364] THF tetrahydrofuran [0365]
TLC thin layer chromatography
LC-MS and GC-MS Methods:
Method 1 (LC-MS):
[0366] Instrument: Micromass Quattro Premier with Waters UPLC
Acquity; column: Thermo Hypersil GOLD 1.9.mu., 50 mm.times.1 mm;
eluent A: 1 L water+0.5 mL 50% aq. formic acid, eluent B: 1 L
acetonitrile+0.5 ml 50% aq. formic acid; gradient: 0.0 min 90%
A.fwdarw.0.1 min 90% A.fwdarw.1.5 min 10% A.fwdarw.2.2 min 10% A;
temperature: 50.degree. C.; flow rate: 0.33 mL/min; UV detection:
210 nm.
Method 2 (LC-MS):
[0367] Instrument: Waters Acquity SQD UPLC System; column: Waters
Acquity UPLC HSS T3 1.8.mu., 50 mm.times.1 mm; eluent A: 1 L
water+0.25 mL 99% formic acid, eluent B: 1 L acetonitrile+0.25 mL
99% formic acid; gradient: 0.0 min 90% A.fwdarw.1.2 min 5%
A.fwdarw.2.0 min 5% A; oven: 50.degree. C.; flow rate: 0.40 mL/min;
UV detection: 210-400 nm.
Method 3 (LC-MS):
[0368] Instrument: Micromass Quattro Micro with HPLC Agilent 1100
Series; column: YMC-Triart C18 3.mu., 50 mm.times.3 mm; eluent A: 1
L water+0.01 mol ammonium carbonate, eluent B: 1 L acetonitrile;
gradient: 0.0 min 100% A.fwdarw.2.75 min 5% A.fwdarw.4.5 min 5% A;
oven: 40.degree. C.; flow rate: 1.25 mL/min; UV detection: 210
nm.
Method 4 (LC-MS):
[0369] Instrument: Waters Acquity SQD UPLC System; column: Waters
Acquity UPLC HSS T3 1.8.mu., 30 mm.times.2 mm; eluent A: 1 L
water+0.25 mL 99% formic acid, eluent B: 1 L acetonitrile+0.25 ml.
99% formic acid; gradient: 0.0 min 90% A.fwdarw.1.2 min 5%
A.fwdarw.2.0 min 5% A; oven: 50.degree. C.; flow rate: 0.60 mL/min;
UV detection: 208-400 nm.
Method 5 (LC-MS):
[0370] Instrument: Micromass Quattro Premier with Waters UPLC
Acquity; column: Thermo Hypersil GOLD 1.9.mu., 50 mm.times.1 mm;
eluent A: 1 L water+0.5 ml, 50% aq. formic acid, eluent B: 1 L
acetonitrile+0.5 ml 50% aq. formic acid; gradient: 0.0 min 97%
A.fwdarw.0.5 min 97% A.fwdarw.3.2 min 5% A.fwdarw.4.0 min 5% A;
temperature: 50.degree. C.; flow rate: 0.3 m/min; UV detection: 210
nm.
Method 6 (GC-MS):
[0371] Instrument: Micromass GCT, GC6890; column: Restek RTX-35, 15
m.times.200 m.times.0.33 .mu.m; constant flow with helium: 0.88
mL/min; oven: 70.degree. C.; inlet: 250.degree. C.; gradient:
70.degree. C., 30.degree. C./min.fwdarw.310.degree. C. (maintain
for 3 min).
Method 7 (LC-MS):
[0372] Instrument MS: Waters Micromass QM; Instrument HPLC: Agilent
1100 series; column: Agilent ZORBAX Extend-C18 3.0 mm.times.50 mm,
3.5.mu.; eluent A: 1 L water+0.01 mol ammonium carbonate, eluent B:
1 L acetonitrile; gradient: 0.0 min 98% A.fwdarw.0.2 min 98%
A.fwdarw.3.0 min 5% A.fwdarw.4.5 min 5% A; temperature: 40.degree.
C.; flow rate: 1.75 mL/min; UV detection: 210 nm.
Method 8 (LC-MS):
[0373] Instrument MS: Waters Micromass ZQ; Instrument HPLC: Agilent
1100 series; column: Agilent ZORBAX Extend-C18 3.0 mm.times.50 mm,
3.5 g; eluent A: 1 L water+0.01 mol ammonium carbonate, eluent B: 1
L acetonitrile; gradient: 0.0 min 98% A.fwdarw.0.2 min 98%
A.fwdarw.3.0 min 5% A.fwdarw.4.5 min 5% A; temperature: 40.degree.
C.; flow rate: 1.75 mL/min; UV detection: 210 nm.
General Purification Methods (See Table I and II Below):
Method P1:
[0374] Preparative RP-HPLC (Reprosil C18, gradient
acetonitrile/0.2% aq. trifluoroacetic acid).
Method P2:
[0375] Preparative RP-HPLC (XBridge C18, gradient
acetonitrile/water+0.1% aq. ammonia).
Method P3:
[0376] Preparative RP-HPLC (Sunfire C18, gradient
acetonitrile/water).
Method P4:
[0377] Preparative RP-HPLC (XBridge C18, gradient
acetonitrile/water+0.05% aq. ammonia).
Method P5:
[0378] The product obtained from the preceding RP-HPLC purification
is dissolved in methanol and filtered through an anion exchange
cartridge (Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge
is eluted with methanol, and the filtrate is evaporated.
Method P6:
[0379] A solution of the product in ethyl acetate is washed with
sat. aq. sodium hydrogencarbonate solution followed by sat. aq.
sodium chloride solution, dried over magnesium sulfate, filtered
and evaporated.
Starting Materials and Intermediates:
Intermediate 1A
2-Methoxy-4-methylaniline
##STR00045##
[0381] A mixture of 5-methyl-2-nitroanisol (265 g, 1.58 mol) and
10% Pd/C (39.75 g) in THF (1.32 L) was stirred overnight at rt
under 1 atm of hydrogen. Filtration over kieselguhr and evaporation
afforded 216.1 g of the crude product which was used in the next
step without further purification.
[0382] LC-MS (method 3): R.sub.t=2.39 min; MS (ESIpos): m/z=138
(M+H).sup.+
[0383] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=6.45-6.63 (m,
3H), 4.46 (s, 2H), 3.72 (s, 3H), 2.16 (s, 3H) ppm.
Intermediate 2A
2-Methoxy-4-methylbenzenethiol
##STR00046##
[0384] Method 1:
[0385] A solution of sodium nitrite (7 g, 101.4 mmol) in water (25
ml) was added dropwise to a cooled (0.degree.-5.degree. C.)
solution of Intermediate 1A (13.7 g, 100 mmol) in concentrated
hydrochloric acid (30 ml) and water (85 ml). After stirring at
0.degree. C. for 10 min, sodium acetate (15 g, 182.8 mmol) was
added. The resulting mixture was added dropwise to a hot solution
(70.degree.-80.degree. C.) of potassium O-ethyl dithiocarbonate (30
g, 187.1 mmol) in water (140 ml), stirred between 70.degree. C. and
80.degree. C. for 1 h and then cooled to rt. The mixture was
extracted twice with ethyl acetate, and the combined organic
extracts were dried over sodium sulfate and evaporated. The residue
was taken up in a 1.3 M solution of potassium hydroxide in ethanol
(300 ml). Glucose (8 g) was added, and the resulting mixture was
refluxed for 3 h. Then, the ethanol solvent was evaporated, and the
residue was diluted with water and acidified with 6 N aqueous
sulfuric acid. Zinc powder (15 g) was added carefully, and the
resulting mixture was heated to 50.degree. C. for 30 min. The
mixture was then cooled to rt, diluted with dichloromethane and
filtered. The filtrate was extracted twice with dichloromethane,
and the combined organic extracts were dried over sodium sulfate
and evaporated affording 14.3 g of the crude product which was used
in the next step without further purification.
Method 2:
[0386] To 2.9 L of THF was added a warm solution of 355 ml (6.67
mol) concentrated sulfuric acid in 1.1 L of water. At 50.degree.
C., 293 g (1.33 mol) 2-methoxy-4-methylbenzenesulfonyl chloride
were added under stirring. Then, 521 g (7.97 mol) of zinc powder
were added carefully in portions (foaming), and the slightly
exothermic reaction was cooled in a water bath to maintain a
temperature of 50.degree.-55.degree. C. The mixture was
subsequently stirred at 55.degree. C. for 3 h. The progress of the
reaction was monitored by TLC (silica gel, petrolether/ethyl
acetate 95:5). The reaction mixture was poured into 13.6 L of
water, 6.8 L dichloromethane were added, and the mixture was
stirred for 5 min. After decanting from remaining zinc and phase
separation, the aqueous phase was extracted once more with 6.8 L
dichloromethane. The combined organic phases were washed with 10%
brine, dried and evaporated at 40.degree. C. under reduced pressure
yielding 237 g of crude product. This material was used in the next
step without further purification. An analytical sample was
obtained by silica gel chromatography with petrolether/ethyl
acetate (97:3) as eluent.
[0387] LC-MS (method 1): R.sub.t=1.21 min; MS (ESIneg): m/z=153
(M-H).sup.-
[0388] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.17 (d, 1H),
6.81 (s, 1H), 6.66 (d, 1H), 4.63 (br. s, 1H), 3.80 (s, 3H), 2.26
(s, 3H) ppm.
Intermediate 3A
1-[(2,2-Diethoxyethyl)sulfanyl]-2-methoxy-4-methylbenzene
##STR00047##
[0390] 237 g crude material from Intermediate 2A, 287 g (1.46 mol)
bromoacetaldehyde-diethylacetal and 862 g (2.65 mol) caesium
carbonate were suspended in 2 L DMF. The reaction temperature
increased initially to 40.degree. C., then stirring was continued
overnight at ambient temperature. The reaction mixture was
partitioned between 10 L of water and 2.7 L of ethyl acetate. The
aqueous phase was extracted with another portion of 2.7 L ethyl
acetate. The combined organic phases were washed with 10% brine,
dried and evaporated. The resulting oily residue was purified by
silica gel chromatography with petrolether/ethyl acetate (95:5) as
eluent.
[0391] Yield: 236 g of an oil (66% of th.)
[0392] GC-MS (method 6): R.sub.t=6.03 min; MS (EIpos): m/z=270
(M).sup.+
[0393] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.16 (d, 1H),
6.82 (s, 1H), 6.73 (d, 1H), 4.55 (t, 1H), 3.80 (s, 3H), 3.52-3.64
(m, 2H), 3.39-3.51 (m, 2H), 2.96 (d, 2H), 2.33 (s, 3H), 1.09 (t,
6H) ppm.
Intermediate 4A
7-Methoxy-5-methyl-1-benzothiophene
##STR00048##
[0395] To a refluxing mixture of 13 g polyphosphoric acid and 150
ml chlorobenzene was added dropwise a solution of 5.2 g (19.2 mmol)
of Intermediate 3A, and refluxing was continued overnight. After
cooling, the organic layer was decanted, and the residue and flask
were rinsed twice with DCM. The combined organic phases were
evaporated at reduced pressure. The residue (3.76 g) was
chromatographed on silica gel with isohexane/0-10% ethyl acetate as
eluent.
[0396] Yield: 1.69 g of an oil (49% of th.)
[0397] GC-MS (method 6): R.sub.t=5.20 min; MS (EIpos): m/z=178
(M).sup.+
[0398] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.68 (d, 1H),
7.34 (d, 1H), 7.28 (s, 1H), 6.78 (s, 1H), 3.93 (s, 3H), 2.43 (s,
3H) ppm.
[0399] Intermediate 5A
(7-Methoxy-5-methyl-1-benzothiophen-2-yl)boronic acid
##STR00049##
[0401] Under argon atmosphere, 26.7 g (150 mmol) of Intermediate 4A
were dissolved in 270 ml of THF and cooled to -70.degree. C.
Between -70.degree. C. and -65.degree. C., 66 ml (165 mmol) of a
2.5 N solution of n-butyllithium in hexane were added dropwise
within 20 min, resulting in formation of a white precipitate. After
stirring for 1 h at -70.degree. C., 41.5 ml (180 mmol) triisopropyl
borate were added at this temperature within 10 min (resulting in a
thick suspension). Stirring was continued for 1 h at -70.degree.
C., before the reaction mixture was allowed to warm up to rt
overnight. Then, 400 ml of saturated aq. ammonium chloride solution
were added, the layers were separated, and the aqueous layer was
extracted once more with THF. The combined organic phases were
evaporated under reduced pressure. To the residue thus obtained,
200 ml of water and 86 ml of 2 N aq. sodium hydroxide solution were
added. The solution was washed twice with DCM, then acidified with
35 ml of 3 M sulfuric acid, and the resulting suspension was
stirred vigorously for 1 h. The precipitate was filtered off by
suction and dried overnight at 45.degree. C. in vacuo.
[0402] Yield: 28.25 g of a colorless solid (94% pure by LC-MS, 80%
of th.)
[0403] LC-MS (method 2): R.sub.t=0.87 min; MS (ESIpos): m/z=223
(M+H).sup.+
[0404] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.17 (d, 1H),
6.81 (s, 1H), 6.66 (d, 1H), 4.63 (br. s, 1H), 3.80 (s, 3H), 2.26
(s, 3H) ppm.
Intermediate 6A
2-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-6-methyl-1,3,6,2-dioxazaboroca-
ne-4,8-dione
##STR00050##
[0406] 6.3 g (28.4 mmol) of Intermediate 5A and 4.2 g (28.4 mmol)
2,2'-(methylimino)diacetic acid were dissolved in a mixture of 45
ml DMSO and 400 ml toluene and refluxed for 16 h using a Dean-Stark
trap. After evaporation, the residue was taken up in ethyl acetate
and washed three times with water and once with brine. The organic
phase was dried over magnesium sulfate and evaporated to a volume
of about 200 ml. A white solid precipitated which was filtered,
washed with ethyl acetate and dried in vacuo to give a first crop
(5.52 g) of the title compound. A second crop (3.32 g) was obtained
after evaporation of the mother liquor and flash-chromatography
over a layer of silica gel using cyclohexane/0-100% ethyl acetate
as the eluent.
[0407] Yield: 8.84 g (overall purity 92.5% by LC-MS, 87% of
th.)
[0408] LC-MS (method 2): R.sub.t=0.93 min; MS (ESIpos): m/z=334
(M+H).sup.+
[0409] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.42 (s, 1H),
7.26 (s, 1H), 6.76 (s, 1H), 4.40 (d, 2H), 4.17 (d, 2H), 3.92 (s,
3H), 2.63 (s, 3H), 2.42 (s, 3H) ppm.
Intermediate 7A
Ethyl 4-aminopyrrolo[2,1-f][1,2,4]triazine-6-carboxylate
##STR00051##
[0411] A solution of
4-aminopyrrolo[2,1-f][1,2,4]triazine-6-carbonitrile (3.9 g, 24.5
mmol; preparation described in PCT Int. Pat. Appl. WO 2007/064883)
in ethanol (124.8 ml) was stirred with concentrated sulfuric acid
(62.4 ml) at 80.degree. C. overnight. After cooling to rt, the
reaction mixture was poured onto 800 g of ice and brought to pH 6-7
with concentrated aq. sodium hydroxide solution. Ethyl acetate (500
ml) and dichloromethane (500 ml) were added to the suspension, and
the resulting mixture was filtered over kieselguhr. The organic
layer was separated from the aqueous layer. The solid was dissolved
in hot water (1 L), and the aqueous layer was extracted twice with
ethyl acetate. The combined organic layers were dried over sodium
sulfate and evaporated. The residue was triturated with an
isopropanol/diethylether mixture, and the solid was filtered off
yielding 2.5 g (49% of th.) of the title compound.
[0412] LC-MS (method 2): R.sub.t=0.59 min; MS (ESIpos): m/z=206
(M+H).sup.+
[0413] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.11-7.97 (m,
3H), 7.88 (s, 1H), 7.34 (br. s, 1H), 4.27 (q, 2H), 1.30 (t, 3H)
ppm.
Intermediate 8A
(4-Aminopyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00052##
[0415] An ice-cooled solution of Intermediate 7A (3.0 g, 14.5 mmol)
in THF (30 ml) was treated with a 1 M solution of lithium
triethylborohydride in THF (58 ml) and stirred at rt for 45 min.
The reaction mixture was then cooled to 0.degree. C., quenched with
methanol, warmed slowly to rt and adsorbed on kieselguhr.
Purification by column chromatography over silica gel
(dichloromethane/methanol 20:1.fwdarw.4:1 gradient) afforded 2.21 g
(92.5% of th.) of the title compound.
[0416] LC-MS (method 3): R.sub.t=1.46 min; MS (ESIpos): m/z=164
(M+H).sup.+
[0417] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.75 (s, 1H),
7.64 (br. s, 2H), 7.50 (br. d, 1H), 6.79 (br. d, 1H), 5.01 (t, 1H),
4.50 (d, 2H) ppm.
Intermediate 9A
tert-Butyl [2-cyano-4-(hydroxymethyl)-1H-pyrrol-1-yl]carbamate
##STR00053##
[0419] Under argon, a 1 M solution of methylmagnesium bromide in
THF (13.3 ml) was added over 15 min to a solution of tert-butyl
(4-bromo-2-cyano-1H-pyrrol-1-yl)carbamate (3.7 g, 12.09 mmol;
preparation described in PCT Int. Pat. Appl. WO 2007/064883,
Intermediate AAE, Step 3) in THF (37 ml) cooled to -60.degree. C.
After 30 min, a 1.6 M solution of n-butyllithium in hexane (15.1
ml, 24.2 mmol) was added over 10 min to the reaction, and the
resulting mixture was stirred between -60.degree. C. and
-40.degree. C. for 1 h. Then, paraformaldehyde (1.09 g, 36.3 mmol)
was added to the reaction, and the reaction mixture was slowly
warmed to rt and stirred overnight. After quenching with sat. aq.
ammonium chloride solution, the aqueous layer was extracted twice
with ethyl acetate. The combined organic phases were washed with
sat. aq. sodium chloride solution, dried over sodium sulfate and
evaporated. Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 2:1.fwdarw.1:1) afforded 2.04 g (69% of
th.) of the title compound.
[0420] LC-MS (method 4): R.sub.t=0.70 min; MS (ESIpos): m/z=238
(M+H).sup.+
[0421] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.79 (br. s,
1H), 7.09 (d, 1H), 6.86 (d, 1H), 4.97 (t, 1H), 4.28 (d, 2H), 1.45
(s, 9H) ppm.
Intermediate 10A
6-(Methoxymethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00054##
[0422] Method 1:
[0423] A solution of Intermediate 8A (1.3 g, 7.9 mmol) in THF (25
ml) was treated with thionyl chloride (1.15 ml, 15.8 mmol) and
stirred at rt for 2 h. After evaporation, the residue was dissolved
in methanol (25 ml) and treated with sodium acetate (1.3 g, 15.8
mmol). The mixture was stirred for 3 h at 65.degree. C. and then
evaporated again. Purification by column chromatography on silica
gel (dichloromethane/methanol 100:2) afforded 787 mg (55% of th.)
of the title compound.
Method 2:
[0424] A solution of Intermediate 9A (6.14 g, 25.88 mmol) in a 4 M
solution of hydrogen chloride in 1,4-dioxane (15 ml) was stirred at
rt for 5 h. After dilution with methanol (73 ml), stirring at rt
was continued overnight. Then, potassium phosphate (54.9 g, 258.65
mmol) and formamidinium acetate (13.46 g, 129.32 mmol) were added,
and the resulting mixture was stirred at 65.degree. C. for 17 h.
The reaction mixture was evaporated, sat. aq. sodium chloride
solution was added, and the mixture was extracted with
dichloromethane followed by ethyl acetate. The combined organic
phases were dried over sodium sulfate and evaporated. Purification
by column chromatography on silica gel (dichloromethane/methanol
40:1.fwdarw.20:1) afforded 2.36 g (49% of th.) of the title
compound.
[0425] LC-MS (method 3): R.sub.t=1.72 min; MS (ESIpos): m/z=179
(M+H).sup.+
[0426] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.77 (s, 1H),
7.69 (br. s, 2H), 7.57 (s, 1H), 6.81 (s, 1H), 4.42 (s, 2H), 3.25
(s, 3H) ppm.
Intermediate 11A
4-Amino-6-(methoxymethyl)pyrrolo[2,1-f][1,2,4]triazine-7-carbaldehyde
##STR00055##
[0428] Phosphoryl chloride (13.7 ml, 147.18 mmol) was added to a
solution of Intermediate 10A (5.24 g, 29.43 mmol) in DMF (80 ml) at
0.degree. C. The resulting mixture was stirred at 60.degree. C. for
8 h, then carefully quenched with water and neutralized with 4 M
aq. sodium hydroxide solution. The aqueous layer was extracted with
ethyl acetate. The combined organic phases were washed with sat.
aq. sodium chloride solution, dried over magnesium sulfate and
evaporated. A solution of the residue in methanol (50 ml) was
treated with sodium acetate (2.41 g, 29.43 mmol) and refluxed
overnight. The reaction mixture was diluted with water and
extracted with ethyl acetate. The combined organic phases were
washed with sat. aq. sodium chloride solution, dried over magnesium
sulfate and evaporated, affording 2.66 g of the crude product which
was used in the next step without further purification.
[0429] LC-MS (method 4): R.sub.t=0.50 min; MS (ESIpos): m/z=207
(M+H).sup.+
[0430] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.35 (s, 1H),
8.20 (br. s, 2H), 8.07 (s, 1H), 7.06 (s, 1H), 4.72 (s, 2H), 3.39
(s, 3H) ppm.
Intermediate 12A
4-Amino-5-bromo-6-(methoxymethyl)pyrrolo[2,1-f][1,2,4]triazine-7-carbaldeh-
yde
##STR00056##
[0432] A solution of Intermediate 11A (crude, 2.66 g) in DMF (30
ml), cooled to -30.degree. C., was treated with a solution of
N-bromosuccinimide (NBS; 2.52 g, 14.19 mmol) in DMF (14 ml). The
resulting mixture was slowly warmed to 0.degree. C. After 1 hour,
the mixture was warmed to rt, stirred for further 15 min and then
quenched with 1 M aq. sodium thiosulfate solution. The precipitate
was filtered off and washed with ethyl acetate, affording 1.1 g
(100% purity, 30% of th.) as a first crop of the title compound.
The remaining filtrate was extracted with ethyl acetate. The
combined organic phases were washed with sat. aq. sodium chloride
solution, dried over sodium sulfate and evaporated. Purification of
the residue by column chromatography on silica gel
(cyclohexane/ethyl acetate 1:1.fwdarw.1:3) afforded further 1.39 g
(70% purity, 26% of th.) of the title compound.
[0433] LC-MS (method 2): R.sub.t=0.67 min; MS (ESIpos): m/z=283/285
(M+H).sup.+
[0434] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.36 (s, 1H),
8.63 (br. s, 1H), 8.13 (s, 1H), 7.23 (br. s, 1H), 4.64 (s, 2H),
3.26 (s, 3H) ppm.
Intermediate 13A
4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazine-7-carbaldehyde
##STR00057##
[0436] Under argon, a degassed 0.5 M aq. potassium phosphate
solution (9.9 ml) was added to a solution of Intermediate 12A (710
mg, 2.49 mmol), Intermediate 5A (921 mg, 3.73 mmol) and
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 196 mg, 249 .mu.mol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010)) in
degassed THF (28.4 ml). The resulting mixture was stirred at
60.degree. C. for 2 h and then evaporated. Purification of the
residue by column chromatography on silica gel (cyclohexane/ethyl
acetate 5:1.fwdarw.1:1) afforded 550 mg (51% of th.) of the title
compound.
[0437] LC-MS (method 2): R.sub.t=1.06 min; MS (ESIpos): m/z=383
(M+H).sup.+
[0438] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.50 (s, 1H),
8.43 (br. s, 1H), 8.21 (s, 1H), 7.42 (s, 1H), 7.33 (s, 1H), 6.87
(s, 1H), 6.09 (br. s, 1H), 4.58 (s, 2H), 3.96 (s, 3H), 3.19 (s,
3H), 2.46 (s, 3H) ppm.
Intermediate 14A
6-(Ethoxymethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00058##
[0439] Method 1:
[0440] To a suspension of 2 g (12.2 mmol) of Intermediate 8A in 40
ml THF were added 1.78 ml (24.4 mmol) thionyl chloride at rt within
20 see. The mixture was stirred for 1.5 h, then evaporated to
dryness, and the residue was dissolved in 40 ml ethanol. 2 g (24.4
mmol) sodium acetate were added, and the mixture was stirred at
70.degree. C. for 1 h 45 min. The reaction mixture was evaporated
again, and sat. aq. sodium hydrogencarbonate solution was added.
The mixture was extracted five times with ethyl acetate. The
combined organic phases were washed with sat. aq. sodium chloride
solution, dried with magnesium sulfate and evaporated to dryness,
giving 2.02 g of crude product which was purified by column
chromatography on silica gel with dichloromethane/methanol (0-2%)
as the eluent.
[0441] Yield: 1.37 g (58% of th.).
Method 2:
[0442] Step 1: A solution of Intermediate 9A (2.3 g, 9.69 mmol) in
1,4-dioxane (5 ml) was treated with a 4 M solution of hydrogen
chloride in 1,4-dioxane (24 ml, 96.9 mmol) and stirred at rt for
130 min. Then, the suspension was filtered, and the precipitate was
washed with 1,4-dioxane (5 ml) and dried in vacuo yielding 1.01 g
(54% of th.) of the intermediate compound
1-amino-4-(chloromethyl)-1H-pyrrole-2-carbonitrile
hydrochloride.
[0443] Step 2: Freshly prepared
1-amino-4-(chloromethyl)-1H-pyrrole-2-carbonitrile hydrochloride
from Step 1 (0.3 g, 1.82 mmol) was dissolved in ethanol (10 ml) and
stirred at rt for 5 min. The clear solution was treated with
formamidine acetate (813 mg, 7.81 mmol) and potassium phosphate
(1.66 g, 7.81 mmol) and stirred first at rt for 3 days, then at
80.degree. C. for 10.5 h. More formamidine acetate (488 mg, 4.69
mmol) was added, and the mixture was stirred for further 18 h at
80.degree. C. The mixture was then cooled to rt, and water and
ethyl acetate were added. The organic phase was separated, and the
aqueous phase was extracted twice with ethyl acetate. The combined
organic phases were washed with sat. aq. sodium chloride solution,
dried with magnesium sulfate and evaporated. The residue was
dissolved in a mixture of methanol and dichloromethane, adsorbed on
diatomaceous earth, dried in vacuo and finally purified by
chromatography on silica gel (gradient 0-10%
methanol/dichloromethane) yielding 260 mg (78% of th.) of the title
compound.
[0444] LC-MS (method 3): R.sub.t=2.02 min; MS (ESIpos): m/z=193
(M+H).sup.+
[0445] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.77 (s, 1H),
7.59-7.74 (br. s, 2H), 7.56 (s, 1H), 6.82 (s, 1H), 5.76 (s, 1H),
4.46 (s, 2H), 3.46 (q, 2H), 1.13 (t, 3H) ppm.
Intermediate 15A
4-Amino-6-(ethoxymethyl)pyrrolo[2,1-f][1,2,4]triazine-7-carbaldehyde
##STR00059##
[0447] To a solution of 2.1 g (10.9 mmol) of Intermediate 14A in 40
ml dry DMF, 5.1 ml (54.6 mmol) phosphoryl chloride were added
dropwise at 0.degree. C. under an argon atmosphere. The mixture was
stirred at 60.degree. C. for 10 h. Then, water was added carefully,
and the mixture was stirred at ambient temperature until all
reactive intermediates were destroyed (HPLC control). The acidic
solution was neutralized with 1 M aq. sodium hydroxide solution and
extracted three times with ethyl acetate. The combined organic
phases were washed with sat. aq. sodium chloride solution, dried
over magnesium sulfate and evaporated at reduced pressure.
[0448] Yield: 1.94 g (purity 90%, 81% of th.)
[0449] LC-MS (method 5): R.sub.t=1.49 min; MS (ESIpos): m/z=221
(M+H).sup.+
[0450] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.35 (s, 1H),
8.14-8.26 (m, 2H), 8.07 (s, 1H), 7.07 (s, 1H), 4.76 (s, 2H), 3.58
(q, 2H), 1.20 (t, 3H) ppm.
Intermediate 16A
4-Amino-5-bromo-6-(ethoxymethyl)pyrrolo[2,1-f][1,2,4]triazine-7-carbaldehy-
de
##STR00060##
[0452] To a solution of 73 g (0.33 mol) of Intermediate 15A in 1.9
L DMF was added dropwise a solution of 65 g (0.37 mol) NBS in 200
ml DMF at -15.degree. C. The mixture was allowed to warn to
0.degree. C. and stirred for 3 h at this temperature. The reaction
mixture was poured into 2% aq. sodium thiosulfate solution under
stirring, and the precipitate was filtered off, washed with water
and dried over phosphorous pentoxide in vacuo.
[0453] Yield: 85.6 g (86% of th.)
[0454] LC-MS (method 2): R.sub.t=0.76 min; MS (ESIpos): m/z=299/301
(M+H).sup.+
[0455] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.36 (s, 1H),
8.62 (br. s, 1H), 8.13 (s, 1H), 7.22 (br. s, 1H), 4.68 (s, 2H),
3.49 (q, 2H), 1.10 (t, 3H) ppm.
Intermediate 17A
4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrol-
o[2,1-f][1,2,4]-triazine-7-carbaldehyde
##STR00061##
[0457] Under an argon atmosphere, 714 mg (purity 85%, 2.03 mmol) of
Intermediate 16A, 946 mg (2.84 mmol) of Intermediate 6A and 160 mg
(0.2 mmol)
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-trii-
sopropylbiphenyl-2-yl)phosphine (1:1; see S. L. Buchwald et al., J.
Am. Chem. Soc. 132 (40), 14073-14075 (2010)) were suspended in 15.5
ml THF. Then, 15.5 ml of a degassed 0.5 M aq. potassium phosphate
solution were added, and the mixture was stirred at 50.degree. C.
for 16 h. After addition of water, the mixture was extracted with
ethyl acetate, and the combined organic phases were dried over
magnesium sulfate and evaporated under reduced pressure. The
residue was purified by column chromatography on silica gel (100 g)
with 10-50% ethyl acetate/cyclohexane as the eluent.
[0458] Yield: 452 mg (75% pure by HPLC, 42% of th.)
[0459] LC-MS (method 5): R.sub.t=2.38 min; MS (ESIpos): m/z=397
(M+H).sup.+
[0460] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.50 (s, 1H),
8.42 (br. s, 1H), 8.21 (s, 1H), 7.43 (s, 1H), 7.33 (s, 1H), 6.87
(s, 1H), 6.07 (br. s, 1H), 4.63 (s, 2H), 3.96 (s, 3H), 3.40 (q,
2H), 2.46 (s, 3H), 1.02 (t, 3H) ppm.
Intermediate 18A
(4-Amino-5,7-dibromopyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00062##
[0462] A solution of Intermediate 8A (5 g, 30.4 mmol) in THF (100
ml) was treated with 1,3-dibromo-5,5-dimethylhydantoin (9.58 g,
33.5 mmol) and stirred at rt for 2 h. The precipitate was filtered
off and dried in vacuo to afford 6.60 g (64% of th.) of the title
compound.
[0463] LC-MS (method 2): R.sub.t=0.56 min; MS (ESIpos):
m/z=321/323/325 (M+H).sup.+
[0464] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.23 (br. s,
1H), 7.96 (s, 1H), 6.94 (br. s, 1H), 5.09 (br. s, 1H), 4.43 (s, 2H)
ppm.
Intermediate 19A
(4-Amino-5-bromopyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00063##
[0466] A suspension of Intermediate 18A (3.7 g, 11.5 mmol) in THF
(800 ml) was heated under stirring until complete dissolution. The
mixture was then cooled to -78.degree. C., and a 1.6 M solution of
n-butyllithium in hexanes (20 ml, 32.1 mmol) was added dropwise.
After 5 min, a further portion of 1.6 M n-butyllithium solution
(1.5 ml, 2.29 mmol) was added. The resulting mixture was stirred at
-78.degree. C. for 5 min, then quenched with methanol (5 ml) and
warmed to rt. The reaction mixture was diluted with sat. aq.
ammonium chloride solution, sat. aq. sodium hydrogencarbonate
solution, sat. aq. sodium chloride solution and ethyl acetate.
After phase separation, the organic layer was washed with sat. aq.
sodium chloride solution. The combined aqueous phases were
re-extracted with ethyl acetate. The combined organic phases were
washed again with sat. aq. sodium chloride solution, dried over
magnesium sulfate and evaporated to afford 2.87 g of the crude
product which was used in subsequent steps without further
purification.
[0467] LC-MS (method 3): R.sub.t=1.73 min; MS (ESIpos): m/z=243/245
(M+H).sup.+
[0468] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.41-7.89 (br.
s, 1H), 7.82 (s, 1H), 7.66 (s, 1H), 7.13-6.48 (br. s, 1H), 5.11 (t,
1H), 4.45 (d, 2H) ppm.
Intermediate 20A
5
[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4-
]triazin-6-yl]methanol
##STR00064##
[0470] A suspension of Intermediate 19A (70% purity, 2.52 g, 7.26
mmol), Intermediate 6A (3.63 g, 10.9 mmol) and caesium fluoride
(5.51 g, 36.3 mmol) in a THF/water mixture (10:1; 80 ml) was
degassed under argon.
4-(Di-tert-butylphosphino)-N,N-dimethylaniline-dichloropalladium
(2:1; 176 mg, 0.248 mmol) was added, and the resulting mixture was
degassed again and stirred at 50.degree. C. for 16 h. The reaction
mixture was then washed with sat. aq. sodium chloride solution, and
the organic layer was separated, dried over magnesium sulfate,
filtered and evaporated. The residue was suspended in methanol, and
the resulting solid was filtered off and dried in vacuo to afford
1.97 g (90% purity, 72% of th.) of the title compound.
[0471] LC-MS (method 2): R.sub.t=0.85 min; MS (ESIpos): m/z=340
(M+H).sup.+
[0472] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.91 (s, 1H),
7.5-8.1 (br. s, 1H), 7.72 (s, 1H), 7.35 (s, 1H), 7.30 (s, 1H), 6.84
(s, 1H), 5.5-6.0 (br. s, 1H), 5.06 (t, 1H), 4.49 (d, 2H), 3.95 (s,
3H), 2.45 (s, 3H) ppm.
Intermediate 21A
6-(Methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f-
][1,2,4]triazin-4-amine
##STR00065##
[0474] A solution of Intermediate 20A (400 mg, 1.17 mmol) in
dichloromethane (12 ml) was treated with thionyl chloride (128
.mu.l, 1.76 mmol) and stirred at rt for 15 min. After evaporation,
the residue was taken up in methanol (12 ml) and treated with DIPEA
(409 .mu.l, 2.35 mmol). The mixture was refluxed overnight and then
evaporated again. Purification by column chromatography on silica
gel (dichloromethane/methanol 98:2.fwdarw.95:5) afforded 388 mg
(93% of th.) of the title compound.
[0475] LC-MS (method 2): R.sub.t=1.00 min; MS (ESIpos): m/z=355
(M+H).sup.+
[0476] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.93 (s, 1H),
7.82 (s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H), 4.38 (s,
2H), 3.95 (s, 3H), 3.22 (s, 3H), 2.45 (s, 3H) ppm.
Intermediate 22A
6-(Ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f]-
[1,2,4]triazin-4-amine
##STR00066##
[0478] Intermediate 20A (200 mg, 587 .mu.mol) in dichloromethane (5
ml) was treated with thionyl chloride (64 .mu.l, 881 .mu.mol) and
stirred at rt for 15 min. After evaporation, the residue was
refluxed in ethanol (5 ml) for 1 h, then treated with DIPEA (204
.mu.l, 1.17 mmol) and refluxed again overnight. The reaction
mixture was evaporated, and the crude product was purified by
column chromatography on silica gel (dichloromethane/methanol
98:2.fwdarw.95:5) affording 202 mg (90% of th.) of the title
compound.
[0479] LC-MS (method 5): R.sub.t=2.32 min; MS (ESIpos): m/z=369
(M+H).sup.+
[0480] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.31-7.59 (br.
s, 1H), 7.93 (s, 1H), 7.81 (s, 1H), 7.35 (s, 1H), 7.30 (s, 1H),
6.84 (s, 1H), 6.20-5.50 (br. s, 1H), 4.41 (s, 2H), 3.95 (s, 3H),
3.41 (q, 2H), 2.45 (s, 3H), 1.08 (t, 3H) ppm.
Intermediate 23A
tert-Butyl
4-{[4-amino-6-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothio-
phen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piperazine-1-carboxylat-
e
##STR00067##
[0482] A solution of Intermediate 20A (9.5 g, 27.9 mmol) in acetic
acid (136.8 ml) was treated with tert-butyl
piperazine-1-carboxylate (6.24 g, 33.49 mmol) and 37% aq.
formaldehyde solution (2.5 ml, 33.49 mmol). The mixture was stirred
at 60.degree. C. for 2.5 h. After evaporation, the residue was
taken up in ethyl acetate, and the mixture was washed twice with
sat. aq. sodium hydrogencarbonate solution, dried over sodium
sulfate and evaporated. The residue was triturated in boiling
ethanol (100 ml). The solid was filtered off and washed with
ethanol and diethylether affording 9.70 g (58% of th.) of the title
compound.
[0483] LC-MS (method 2): R.sub.t=0.88 min; MS (ESIpos): m/z=539
(M+H).sup.+
[0484] .sup.1H-NMR (400 MHz, CDCl.sub.3): .delta.=7.94 (s, 1H),
7.13-7.35 (m, 2H, overlap with CHCl.sub.3 peak), 6.67 (s, 1H), 5.86
(br. s, 1H), 5.54 (br. s, 2H), 4.68 (s, 2H), 4.08 (s, 2H), 4.00 (s,
3H), 3.45 (br. s, 4H), 2.59-2.48 (m, 7H), 1.45 (s, 9H) ppm.
Intermediate 24A
tert-Butyl
4-({4-amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-6-[(2-m-
ethoxy-2-oxoethoxy)methyl]pyrrolo[2,1-f][1,2,4]triazin-7-yl}methyl)piperaz-
ine-1-carboxylate
##STR00068##
[0486] A solution of Intermediate 23A (300 mg, 556 .mu.mol) in
dichloromethane (12 ml) was treated with thionyl chloride (81
.mu.l, 1.11 mmol) and stirred at rt for 15 min. After evaporation,
the residue was dissolved in freshly distilled methylglycolate (2.5
ml) and treated with DIPEA (485 .mu.l, 2.78 mmol). The mixture was
stirred at 70.degree. C. for 2 h, then evaporated again, and excess
methylglycolate was removed by distillation. Purification of the
residue by column chromatography on silica gel (cyclohexane/ethyl
acetate 1:5) afforded 136 mg (33% of th.) of the title
compound.
[0487] LC-MS (method 2): R.sub.t=0.88 min; MS (ESIpos): m/z=611
(M+H).sup.+
[0488] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.39 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.58 (s, 2H), 4.11 (s,
2H), 3.96 (s, 3H), 3.93 (s, 2H), 3.57 (s, 3H), 3.30-3.24 (m, 4H),
2.45-2.38 (m, 7H), 1.39 (s, 9H) ppm.
Intermediate 25A
tert-Butyl
4-{[4-amino-6-formyl-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl-
)pyrrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazine-1-carboxylate
##STR00069##
[0490] A solution of Intermediate 23A (300 mg, 556 .mu.mol) in
dichloromethane (4.2 ml) was treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 307 mg,
724 mol) and stirred at rt for 2 h. The reaction mixture was
quenched with sat. aq. sodium hydrogencarbonate solution and sat.
aq. sodium thiosulfate solution (1:1) and stirred at rt for 30 min.
The aqueous layer was extracted three times with dichloromethane,
and the combined organic layers were dried over magnesium sulfate
and evaporated. Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 1:1.fwdarw.100% ethyl acetate) afforded
273 mg (87% of th.) of the title compound.
[0491] LC-MS (method 2): R.sub.t=0.87 min; MS (ESIpos): m/z=537
(M+H).sup.+
[0492] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.02 (s, 1H),
8.44-8.18 (br. s, 1H), 8.08 (s, 1H), 7.51 (s, 1H), 7.32 (s, 1H),
6.88 (s, 1H), 6.01-5.74 (br. s, 1H), 4.16 (s, 2H), 3.96 (s, 3H),
3.30-3.22 (m, 4H), 2.48-2.40 (m, 7H), 1.38 (s, 9H) ppm.
Intermediate 26A
4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmethy-
l)pyrrolo[2,1-f]-[1,2,4]triazine-6-carbaldehyde bis(formiate)
##STR00070##
[0494] A solution of Intermediate 23A (80 mg, 148 .mu.mol) in THF
(3.9 ml) was treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 94 mg, 222
.mu.mol) and stirred at rt for 30 min. The reaction mixture was
quenched with sat. aq. sodium hydrogencarbonate solution and sat.
aq. sodium thiosulfate solution (1:1). The aqueous layer was
extracted three times with ethyl acetate, and the combined organic
layers were dried over magnesium sulfate and evaporated. The
residue was dissolved in a 4 M solution of hydrogen chloride in
1,4-dioxane (4 ml) and stirred at rt for 1 h. After evaporation,
the residue was purified by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid) affording 34 mg
(52% of th.) of the title compound.
[0495] LC-MS (method 2): R.sub.t=0.78 min; MS (ESIpos): m/z=437
(M+H).sup.+
[0496] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=10.02 (s, 1H),
8.29 (br. s, 2H), 8.09 (s, 1H), 7.50 (s, 1H), 7.33 (s, 1H), 6.88
(s, 1H), 4.17 (s, 2H), 3.96 (s, 3H), 2.90-2.81 (m, 4H), 2.63-2.56
(m, 4H), 2.46 (s, 3H) ppm.
Intermediate 27A
tert-Butyl
4-({4-amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-6-[(3-o-
xopiperazin-1-yl)methyl]pyrrolo[2.1-f][1,2,4]triazin-7-yl}methyl)piperazin-
e-1-carboxylate
##STR00071##
[0498] A solution of Intermediate 25A (185 mg, 344 .mu.mol) in THF
(4.6 ml) was treated with 2-oxopiperazine (344 mg, 3.4 mmol),
sodium triacetoxyborohydride (365 mg, 1.7 mmol) and acetic acid (39
.mu.l, 689 .mu.mol). The resulting mixture was stirred at rt for 2
h, then adsorbed on kieselguhr and purified by column
chromatography on silica gel
(dichloromethane.fwdarw.dichloromethane/methanol 100:8) affording
221 mg (quant.) of the title compound.
[0499] LC-MS (method 4): R.sub.t=0.77 min; MS (ESIpos): m/z=621
(M+H).sup.+.
Intermediate 28A
N-Ethylethanaminium
4-amino-7-{[4-(tert-butoxycarbonyl)piperazin-1-yl]carbonyl}-5-(7-methoxy--
5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazine-6-carboxylate
##STR00072##
[0501] A solution of Intermediate 25A (70 mg, 130 .mu.mol) in
THF/water (10:1, 4.85 ml) was treated with a 2 M solution of
2-methyl-2-buten in THF (521 .mu.l, 1.04 mmol) and sodium
dihydrogenphosphate (107 mg, 783 .mu.mol) and stirred at rt for 5
min. Sodium chlorite (70 mg, 783 .mu.mol) was added, and the
resulting mixture was stirred at rt overnight. After dilution with
water, the aqueous phase was extracted three times with ethyl
acetate. The combined organic layers were dried over sodium sulfate
and evaporated, and the residue was purified by preparative RP-HPLC
(XBridge C18, gradient 5-50% acetonitrile/water+0.05% diethylamine)
affording 18 mg (21% of th.) of the title compound.
[0502] LC-MS (method 1): R.sub.t=1.14 min; MS (ESIpos): m/z=567
(M+H).sup.+
[0503] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=8.26-7.89 (br. s, 1H), 7.94 (s, 1H), 7.33 (s, 1H), 7.28 (s,
1H), 6.83 (s, 1H), 5.64-5.33 (br. s, 1H), 3.95 (s, 3H), 2.80 (q,
4H), 2.45 (s, 3H), 1.40 (s, 9H), 1.09 (t, 6H) ppm.
Intermediate 29A
tert-Butyl
4-{[4-amino-6-(azidomethyl)-5-(7-methoxy-5-methyl-1-benzothioph-
en-2-yl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piperazine-1-carboxylate
##STR00073##
[0505] A solution of Intermediate 23A (150 mg, 0.278 mmol) in
dichloromethane (7.5 ml) was treated with thionyl chloride (40
.mu.l, 0.56 mmol) and stirred at rt for 15 min. After evaporation,
the residue was dissolved in DMF (6 ml) and treated with sodium
azide (362 mg, 5.57 mmol) and sodium iodide (208 mg, 1.39 mmol).
The mixture was heated to 80.degree. C. for 1 h, then diluted with
water and extracted three times with ethyl acetate. The combined
organic layers were washed with water and with sat. aq. sodium
chloride solution, dried over magnesium sulfate and evaporated.
Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 98:2.fwdarw.100% ethyl acetate) afforded
95.8 mg (57% of th.) of the title compound.
[0506] LC-MS (method 4): R.sub.t=0.99 min; MS (ESIpos): m/z=564
(M+H).sup.+
[0507] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.20-7.80 (br.
s, 1H), 8.01 (s, 1H), 7.39 (s, 1H), 7.33 (s, 1H), 6.86 (s, 1H),
6.05-5.55 (br. s, 1H), 4.50 (s, 2H), 3.96 (s, 3H), 3.94 (s, 2H),
3.32-3.25 (m, 4H), 2.45 (s, 3H), 2.43-2.36 (m, 4H), 1.39 (s, 9H)
ppm.
Intermediate 30A
tert-Butyl
4-{[6-(acetamidomethyl)-4-amino-5-(7-methoxy-5-methyl-1-benzoth-
iophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piperazine-1-carboxyl-
ate
##STR00074##
[0509] A mixture of Intermediate 29A (320 mg, 567 .mu.mol), 10%
Pd/C (320 mg) and acetic anhydride (106 .mu.l, 1.13 mmol) in
methanol (32 ml) was stirred for 90 min under 1 atm of hydrogen at
rt. The mixture was then filtered through kieselguhr, and the
filtrate was evaporated. Purification by column chromatography on
silica gel (cyclohexane/ethyl acetate 1:1.fwdarw.100% ethyl
acetate) afforded 440 mg (quant.) of the title compound.
[0510] LC-MS (method 2): R.sub.t=0.91 min; MS (ESIpos): m/z=580
(M+H).sup.+
[0511] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.02-7.97 (m,
2H), 7.38 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.27 (br. d, 2H),
3.95 (s, 3H), 3.89 (s, 2H), 3.31-3.24 (m, 4H), 2.45 (s, 3H),
2.43-2.36 (m, 4H), 1.74 (s, 3H), 1.39 (s, 9H) ppm.
Intermediate 31A
7-[(4-Acetylpiperazin-1-yl)methyl]-4-amino-5-(7-methoxy-5-methyl-1-benzoth-
iophen-2-yl)pyrrolo[2,1-f][1,2,4]triazine-6-carbaldehyde
##STR00075##
[0513] A solution of Example 55 (678 mg, purity 89%, 1.26 mmol) in
dichloromethane (4 ml) containing molecular sieves (4 .ANG.) was
treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 623 mg,
1.47 mmol) and stirred at rt for 5 min. The reaction mixture was
then adsorbed on diatomaceous earth and purified by column
chromatography on silica gel (gradient of 30-100% ethyl
acetate/cyclohexane, then 0-10% methanol/dichloromethane) yielding
449 mg (49% of th.) of the title compound.
[0514] LC-MS (method 4): R.sub.t=0.70 min; MS (ESIpos): m/z=479
(M+H).sup.+.
Intermediate 32A
4-{[4-Amino-6-(azidomethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyr-
rolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazin-2-one
##STR00076##
[0516] A solution of Example 13 (59 mg, 130 .mu.mol) in
dichloromethane (3.5 ml) was treated with thionyl chloride (19
.mu.l, 261 .mu.mol) and stirred at rt for 15 min. After
evaporation, the residue was dissolved in DMF (2.8 ml) and treated
with sodium iodide (97 mg, 652 .mu.mol) and sodium azide (169 mg,
2.6 mmol). The mixture was stirred at 80.degree. C. for 1 h. After
dilution with sat. aq. sodium chloride solution, the aqueous phase
was extracted four times with ethyl acetate, and the combined
organic layers were dried over magnesium sulfate and evaporated.
Purification by column chromatography on silica gel
(dichloromethane/methanol 98:2.fwdarw.90:10) afforded 35 mg (56% of
th.) of the title compound.
[0517] LC-MS (method 5): R.sub.t=2.04 min; MS (ESIpos): m/z=478
(M+H).sup.+
[0518] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.02 (s, 1H),
7.75 (br. s, 1H), 7.41 (s, 1H), 7.33 (s, 1H), 6.86 (s, 1H), 4.51
(s, 2H), 4.01 (s, 2H), 3.96 (s, 3H), 3.16-3.08 (m, 2H), 3.04-2.98
(m, 2H), 2.65-2.58 (m, 2H), 2.45 (s, 3H) ppm.
Intermediate 33A
4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin-4-ylmethy-
l)pyrrolo[2,1-f]-[1,2,4]triazine-6-carbaldehyde
##STR00077##
[0520] A solution of Example 50 (710 mg, purity 70%, 1.13 mmol) in
dichloromethane (5 ml) containing molecular sieves (4 .ANG.) was
treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 623 mg,
1.47 mmol) and stirred at rt for 5 min. The reaction mixture was
then adsorbed on diatomaceous earth and purified by column
chromatography on silica gel (gradient of 30-100% ethyl
acetate/cyclohexane) yielding 386 mg (72% of th.) of the title
compound.
[0521] LC-MS (method 2): R.sub.t=0.76 min; MS (ESIpos): m/z=438
(M+H).sup.+.
Intermediate 34A
(4-Amino-7-bromopyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00078##
[0523] A solution of 1,3-dibromo-5,5-dimethylhydantoin (87 mg,
0.305 mmol) in THF (1 ml) was added dropwise to a solution of
Intermediate 8A (100 mg, 0.609 mmol) in THF (4 ml) and methanol (2
ml) at -78.degree. C. The mixture was stirred at -78.degree. C. for
16 h, then diluted with water and extracted with ethyl acetate. The
combined organic layers were washed with sat. aq. sodium chloride
solution, dried over magnesium sulfate and evaporated. Purification
by column chromatography on silica gel (dichloromethane/methanol
20:1.fwdarw.10:1) afforded 55 mg (32% of th.) of the title
compound.
[0524] LC-MS (method 3): R.sub.t=1.71 min; MS (ESIpos): m/z=243/245
(M+H).sup.+
[0525] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.74-7.94 (m,
3H), 7.04 (s, 1H), 5.12 (t, 1H), 4.48 (d, 2H) ppm.
Intermediate 35A
7-Bromo-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f][1,2,4]tri-
azin-4-amine
##STR00079##
[0527] A solution of Intermediate 34A (885 mg, 3.64 mmol) in DMF
(11 ml) was treated with tert-butyldimethylsilyl chloride (823 mg,
5.46 mmol) and imidazole (743 mg, 10.92 mmol) and stirred at rt for
2 h. The reaction mixture was combined with the reaction mixture of
a 100 mg test run, diluted with water and extracted twice with
ethyl acetate. The combined organic phases were washed with water
and sat. aq. sodium chloride solution, dried over magnesium sulfate
and evaporated. Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 2:1.fwdarw.100% ethyl acetate) afforded
1.36 g (93% of th.) of the title compound.
[0528] LC-MS (method 2): R.sub.t=1.13 min; MS (ESIpos): m/z=357/359
(M+H).sup.+
[0529] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.98-7.68 (m,
3H), 7.04 (s, 1H), 4.68 (s, 2H), 0.89 (s, 9H), 0.09 (s, 6H)
ppm.
Intermediate 36A
4-Amino-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f][1,2,4]tri-
azine-7-carbonitrile
##STR00080##
[0531] Under argon, a mixture of Intermediate 35A (880 mg, 2.46
mmol),
[1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II)-dichlorometha-
ne complex [PdCl.sub.2(dppf).times.DCM] (120 mg, 0.148 mmol),
bis(dibenzylideneacetone)palladium [Pd(dba).sub.2] (135 mg, 0.148
mmol), zinc cyanide (578 mg, 4.92 mmol), zinc powder (64 mg, 0.985
mmol) and zinc acetate (180 mg, 0.985 mmol) in degassed DMF/water
(100:1, 9.2 ml) was stirred at 160.degree. C. overnight. The
reaction mixture was then combined with the reaction mixture of a
100 mg test run, and the combined mixtures were adsorbed on
kieselguhr, filtered over another layer of kieselguhr and eluted
with tert-butyl methyl ether. The filtrate was washed with sat. aq.
sodium hydrogencarbonate solution, and the aqueous layer was
re-extracted three times with tert-butyl methyl ether. The combined
organic phases were dried over sodium sulfate and evaporated.
Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 1:1) afforded 453 mg (44% of th.) of the
title compound.
[0532] LC-MS (method 2): R.sub.t=1.09 min; MS (ESIpos): m/z=304
(M+H).sup.+
[0533] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.22-8.35 (m,
2H), 8.06 (s, 1H), 6.95 (s, 1H), 4.83 (s, 2H), 0.91 (s, 9H), 0.11
(s, 6H) ppm.
Intermediate 37A
4-Amino-5-bromo-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f][1-
,2,4]triazine-7-carbonitrile
##STR00081##
[0535] A solution of 1,3-dibromo-5,5-dimethylhydantoin (621 mg,
2.17 mmol) in THF (20 ml) was added dropwise to a solution of
Intermediate 36A (1.1 g, 3.62 mmol) in THF (80 ml) at -50.degree.
C. The resulting mixture was slowly warmed to rt, stirred for 2 h
and then quenched with 10% aq. sodium thiosulfate solution and sat.
aq. sodium hydrogencarbonate solution. The aqueous phase was
extracted three times with ethyl acetate. A solid precipitating
from the ethyl acetate solution was filtered off and washed with
ethyl acetate, affording 508 mg (100% purity, 36% of th.) as a
first crop of the title compound. The remaining filtrate was dried
over sodium sulfate and evaporated. The residue was precipitated
from DMSO and washed with DMSO and ethyl acetate affording further
498 mg (85% purity, 26% of th.) of the title compound.
[0536] LC-MS (method 5): R.sub.t=2.70 min; MS (ESIpos): m/z=382/384
(M+H).sup.+
[0537] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.10 (s, 1H),
4.76 (s, 2H), 0.90 (s, 9H), 0.12 (s, 6H) ppm.
Intermediate 38A
4-Amino-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)-5-(7-methoxy-5-methyl-1-
-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazine-7-carbonitrile
##STR00082##
[0539] Under argon, a solution of Intermediate 37A (459 mg, 1.29
mmol) in degassed THF (14.8 ml) was added to
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 152 mg, 0.19 mmol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010))
and Intermediate 6A (647 mg, 1.94 mmol). Degassed 0.5 M aq.
potassium phosphate solution (5.1 ml) was added dropwise, and the
resulting mixture was stirred at 60.degree. C. for 2 h. The
reaction mixture was then combined with the reaction mixtures of
previous 70 mg, 90 mg and 500 mg test runs and evaporated.
Purification of the residue by column chromatography on silica gel
(cyclohexane/ethyl acetate 3:1.fwdarw.100% ethyl acetate) afforded
1.0 g (58% of th.) of the title compound.
[0540] LC-MS (method 2): R.sub.t=1.49 min; MS (ESIpos): m/z=480
(M+H).sup.+
[0541] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.57-8.35 (br.
s, 1H), 8.18 (s, 1H), 7.41 (s, 1H), 7.30 (s, 1H), 6.86 (s, 1H),
6.28-6.03 (br. s, 1H), 4.74 (s, 2H), 3.95 (s, 3H), 2.44 (s, 3H),
0.83 (s, 9H), 0.00 (s, 6H) ppm.
Intermediate 39A
4-Amino-6-formyl-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f]-
[1,2,4]triazine-7-carbonitrile
##STR00083##
[0543] A solution of Example 68 (250 mg, 0.684 mmol) in
dichloromethane (5 ml) was treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 377 mg,
0.889 mmol) and stirred at rt for 1 h. The reaction mixture was
combined with the reaction mixture from a 50 mg test run, quenched
with sat. aq. sodium hydrogencarbonate solution and sat. aq. sodium
thiosulfate solution (1:1) and stirred at rt for 30 min. The
aqueous phase was extracted three times with ethyl acetate, and the
combined organic phases were dried over sodium sulfate and
evaporated. Purification by column chromatography on silica gel
(cyclohexane/25% ethyl acetate.fwdarw.100% ethyl acetate) afforded
102 mg (24% of th.) of the title compound.
[0544] LC-MS (method 2): R.sub.t=1.02 min; MS (ESIpos): m/z=364
(M+H).sup.+
[0545] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=9.86 (s, 1H),
8.78 (br. s, 1H), 8.30 (s, 1H), 7.62 (s, 1H), 7.36 (s, 1H), 6.91
(s, 1H), 6.51 (br. s, 1H), 3.97 (s, 3H), 2.46 (s, 3H) ppm.
Intermediate 40A
4-Amino-6-(azidomethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo-
[2,1-f][1,2,4]-triazine-7-carbonitrile
##STR00084##
[0547] A solution of Example 68 (100 mg, 273 .mu.mol) in
dichloromethane (5 ml) was treated with thionyl chloride (39 .mu.l,
547 .mu.mol) and stirred at rt for 15 min. After evaporation, the
residue was dissolved in DMF (6 ml) and treated with sodium iodide
(205 mg, 1.36 mmol) and sodium azide (355 mg, 5.47 mmol). The
mixture was stirred at 80.degree. C. overnight, then diluted with
water and extracted three times with ethyl acetate. The combined
organic layers were washed with water, followed by sat. aq. sodium
chloride solution, dried over sodium sulfate and evaporated
affording 91 mg of the crude product which was used in the next
step without further purification.
[0548] LC-MS (method 2): R.sub.t=1.13 min; MS (ESIpos): m/z=391
(M+H).sup.+
[0549] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): d=8.65-8.45 (br. s,
1H), 8.23 (s, 1H), 7.46 (s, 1H), 7.34 (s, 1H), 6.88 (s, 1H),
6.41-6.10 (br. s, 1H), 4.57 (s, 2H), 3.96 (s, 3H), 2.46 (s, 3H)
ppm.
Intermediate 41A
5,7-Dibromo-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f][1,2,4-
]triazin-4-amine
##STR00085##
[0551] A solution of Intermediate 18A (2 g, 6.21 mmol) in DMF (20
ml) was treated with imidazole (846 mg, 12.4 mmol) and
tert-butyldimethylsilyl chloride (1.12 g, 7.45 mmol) and stirred at
rt for 20 h. The reaction mixture was then diluted with water (200
ml) and stirred at rt for further 2 h. Filtration of the solid
afforded 2.46 g (88% of th.) of the title compound.
[0552] LC-MS (method 2): R.sub.t=1.37 min; MS (ESIpos): m/z=437
(M+H).sup.+
[0553] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.49-8.05 (br.
s, 1H), 7.96 (s, 1H), 7.15-6.76 (br. s, 1H), 4.64 (s, 2H), 0.87 (s,
9H), 0.09 (s, 6H) ppm.
Intermediate 42A
2-[4-Amino-5-bromo-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f-
][1,2,4]triazin-7-yl]-propan-2-ol
##STR00086##
[0555] Under argon, a solution of Intermediate 41A (1 g, 2.29 mmol)
in THF (40 ml) was cooled to -78.degree. C. and treated with a 1.6
M solution of methyllithium in diethylether (1.5 ml, 2.40 mmol).
After stirring for 10 min at -78.degree. C., a 1.6 M solution of
n-butyllithium in hexanes (1.58 ml, 2.52 mmol) was added, and
stirring was continued for 10 min. Acetone (1.68 ml, 22.92 mmol)
was added, and the resulting mixture was slowly warmed to rt and
stirred at rt for 18 h. The reaction was then quenched with water,
and the aqueous phase was extracted twice with ethyl acetate. The
combined organic phases were washed with sat. aq. sodium chloride
solution, dried over magnesium sulfate and evaporated. Purification
by flash chromatography on silica gel (cyclohexane/ethyl acetate
2:1) afforded 306 mg (30% of th.) of the title compound.
[0556] LC-MS (method 4): R.sub.t=1.39 min; MS (ESIpos): m/z=415/417
(M+H).sup.+
[0557] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.23-7.85 (br.
s, 1H), 7.89 (s, 1H), 7.05-6.82 (br. s, 1H), 5.49 (s, 1H), 4.88 (s,
2H), 1.66 (s, 6H), 0.87 (s, 9H), 0.08 (s, 6H) ppm.
Intermediate 43A
2-[4-Amino-5-bromo-6-(hydroxymethyl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]prop-
an-2-ol
##STR00087##
[0559] A solution of Intermediate 42A (304 mg, 0.732 mmol) in THF
(15 ml) was treated with a 1 M solution of tetra-n-butylammonium
fluoride in THF (768 .mu.l, 768 .mu.mol) and stirred at rt for 2
min. The reaction mixture was diluted with acetonitrile (20 ml),
then evaporated, and the residue was purified by preparative
RP-HPLC (Reprosil C18, gradient 10-30% acetonitrile/0.2% aq. TFA).
The product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 180 mg (67% of th.) of the
title compound.
[0560] LC-MS (method 2): R.sub.t=0.55 min; MS (ESIpos): m/z=301/303
(M+H).sup.+
[0561] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.21-7.90 (m,
1H), 7.87 (s, 1H), 7.09-6.60 (br. s, 1H), 5.90 (br. s, 1H), 5.03
(br. s, 1H), 4.63 (s, 2H), 1.66 (s, 6H) ppm.
Intermediate 44A
4-Amino-7-(2-hydroxypropan-2-yl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-y-
l)pyrrolo[2,1-f]-[1,2,4]triazine-6-carbaldehyde
##STR00088##
[0563] A solution of Example 73 (135 mg, purity 89%, 302 .mu.mol)
in dichloromethane (7 ml) was treated with Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one; 166 mg,
392 .mu.mol) and stirred at rt for 70 min. The reaction mixture was
combined with the reaction mixture of a 18 mg (45 .mu.mol) test run
and quenched with sat. aq. sodium hydrogencarbonate solution and
sat. aq. sodium thiosulfate solution (1:1). The aqueous phase was
extracted three times with dichloromethane. The combined organic
phases were washed with sat. aq. sodium chloride solution, dried
over magnesium sulfate and evaporated yielding 143 mg (purity 77%,
92% of th.) of the title compound.
[0564] LC-MS (method 2): R.sub.t=1.08 min; MS (ESIpos): m/z=397
(M+H).sup.+.
Intermediate 45A
6-({[tert-Butyl(dimethyl)silyl]oxy}methyl)pyrrolo[2,1-f][1,2,4]triazin-4-a-
mine
##STR00089##
[0566] A solution of 1.5 g (9.14 mmol) of Intermediate 8A in 15 ml
dry DMF was treated with 1.65 g (10.96 mmol)
tert-butyldimethylsilyl chloride and 1.24 g (18.27 mmol) imidazole
and stirred at rt overnight. The reaction mixture was poured into
250 ml water and stirred for 5 min. The resulting precipitate was
filtered off and dried in vacuo at 45.degree. C. Yield: 2.28 g (90%
of th.).
[0567] LC-MS (method 5): R.sub.t=2.12 min; MS (ESIpos): m/z=279
(M+H).sup.+.
Intermediate 46A
6-({[tert-Butyl(dimethyl)silyl]oxy}methyl)-7-chloropyrrolo[2,1-f][1,2,4]tr-
iazin-4-amine
##STR00090##
[0569] A solution of 2 g (7.18 mmol) of Intermediate 45A in 20 ml
THF was treated with 893 mg (6.47 mmol) N-chlorosuccinimide in 6
portions over 60 min at -10.degree. C. Stirring was continued for
15 min at -10.degree. C., then the mixture was allowed to warm to
rt. Another 192 mg (1.44 mmol) N-chlorosuccinimide were added at
rt, and stirring was continued overnight. About one-tenth of the
reaction mixture was evaporated to dryness, and the residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) furnishing 94 mg (4% of th.) of
the title compound. The major part of the reaction mixture was
adsorbed on silica gel and subjected to chromatography on silica
gel with isohexane/ethyl acetate 5-66% as eluent, yielding 899 mg
(40% of th.) of the title compound. Total yield: 993 mg (44% of
th.).
[0570] LC-MS (method 5): R.sub.t=2.45 min; MS (ESIpos): m/z=313
(M+H).sup.+
[0571] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.69-8.04 (m,
3H), 7.0 (s, 1H), 4.70 (s, 2H), 0.88 (s, 9H), 0.08 (s, 6H) ppm.
Intermediate 47A
5-Bromo-6-({[tert-butyl(dimethyl)silyl]oxy}methyl)-7-chloropyrrolo[2,1-f][-
1,2,4]triazin-4-amine
##STR00091##
[0573] A solution of 890 mg (2.85 mmol) of Intermediate 46A in 20
ml DMF was treated with 506 mg (2.85 mmol) N-bromosuccinimide in
portions over 1 h at -10.degree. C. Stirring was continued for 3 h
at -10.degree. C. and then overnight at rt. Water (200 ml) was
added, and the mixture was stirred for 2 h. The precipitated solid
was filtered off, washed with water and dried in vacuo at
45.degree. C. Yield: 997 mg (89% of th.).
[0574] LC-MS (method 5): R.sub.t=2.82 min; MS (ESIpos):
m/z=391/393/395 (M+H).sup.+
[0575] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.32 (br. s,
1H), 7.97 (s, 1H), 6.97 (br. s, 1H), 4.66 (s, 2H), 0.87 (s, 9H),
0.09 (s, 6H) ppm.
Intermediate 48A
6-({[tert-Butyl(dimethyl)silyl]oxy}methyl)-7-chloro-5-(7-methoxy-5-methyl--
1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00092##
[0577] Under an argon atmosphere, a flask was charged with 800 mg
(2.04 mmol) of Intermediate 47A, 680 mg (2.04 mmol) of Intermediate
6A, 80 mg (0.1 mmol)
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; see S. L. Buchwald et al., J. Am.
Chem. Soc. 132 (40), 14073-14075 (2010)) and 1.3 g (6.13 mmol)
potassium phosphate. Then, 30 ml of a degassed 1,4-dioxane/water
mixture (5:1) were added, and the solution was stirred at
70.degree. C. for 1 h. Further 680 mg (2.04 mmol) of Intermediate
6A and 32 mg (0.04 mmol)
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1) were added, and stirring at
70.degree. C. was continued for another 1 h. This procedure was
repeated three times until the starting material was consumed
(control by LC-MS). With the last portion of reagents, also 1.6 ml
of 5 M aq. sodium hydroxide solution were added to achieve a pH
value of 8-9. At the end of the reaction period, 30 ml water and 5
M aq. formic acid were added (pH 3-4) whereupon an oil separated. A
part of this oil was purified by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid). Two fractions
of the title compound were obtained: 103 mg of a solid (89% purity
by LC-MS, 9% of th.), and 23 mg of a solid (100% purity by LC-MS,
2% of th.). The rest of the oil and the supernatant were diluted
with water, adjusted to pH 8-9 with sat. aq. sodium
hydrogencarbonate solution and extracted three times with
dichloromethane. The combined organic phases were washed with
water, dried and evaporated under reduced pressure, yielding 2.1 g
of an oil. This material was purified by column chromatography on
silica gel with dichloromethane/0-5% methanol as eluent to afford
further 709 mg (43% purity by LC-MS, 31% of th.) of the title
compound. Total yield: 42% of th.
[0578] LC-MS (method 2): R.sub.t=1.57 min; MS (ESIpos): m/z=489
(M+H).sup.+
[0579] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.08 (s, 1H),
7.43 (s, 1H), 7.32 (s, 1H), 6.88 (s, 1H), 4.66 (s, 2H), 3.98 (s,
3H), 2.47 (s, 3H), 0.84 (s, 9H), -0.03 (s, 6H) ppm.
Intermediate 49A
4-Amino-7-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f]-
[1,2,4]triazine-6-carbaldehyde
##STR00093##
[0581] To a suspension of 166 mg (0.44 mmol) of Example 79 and
molecular sieves (3 .ANG.) in 3 ml dichloromethane were added 207
mg (0.49 mmol) Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one) at
0-5.degree. C. The mixture was stirred for 10 min at this
temperature, then further 56 mg (0.13 mmol) Dess-Martin periodinane
were added, and stirring was continued for 15 min at 5.degree. C.
and 10 min at ambient temperature. After this, the mixture was
adsorbed on diatomaceous earth and purified by column
chromatography on silica gel with dichloromethane/0-10% methanol as
eluent. Yield: 100 mg of a solid (92.7% pure by LC-MS, 56% of
th.).
[0582] LC-MS (method 2): R.sub.t=1.08 min; MS (ESIpos): m/z=373
(M+H).sup.+
[0583] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=9.90 (s, 1H),
8.46 (br. s, 1H), 8.15 (s, 1H), 7.54 (s, 1H), 7.34 (s, 1H), 6.89
(s, 1H), 6.07 (br. s, 1H), 3.96 (s, 3H), 2.46 (s, 3H) ppm.
Intermediate 50A
7-Chloro-6-(chloromethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazin-4-amine
##STR00094##
[0585] The title compound was isolated as a by-product in the
preparation of Example 79 (see below).
[0586] Yield: 9.2 mg (10% of th.).
[0587] LC-MS (method 3): R.sub.t=2.98 min; MS (ESIpos): m/z=393/395
(M+H).sup.+.
Intermediate 51A
6-({[tert-Butyl(dimethyl)silyl]oxy}methyl)-7-methylpyrrolo[2,1-f][1,2,4]tr-
iazin-4-amine
##STR00095##
[0589] To a solution of 3 g (8.4 mmol) of Intermediate 35A in 60 ml
1,4-dioxane under an argon atmosphere were added 171 mg (0.21 mmol)
of PdCl.sub.2(dppf).times.DCM and then dropwise over 10 min 16.8 ml
of a 2 M solution of dimethylzine in toluene (causing a rise in
temperature from 22.degree. C. to 31.degree. C.). Stirring was
continued first for 10 min at ambient temperature, then for 13 h at
90.degree. C. After this, water (10 ml) was added to the reaction
mixture at rt, and the suspension was stirred for 1 h. The mixture
was evaporated under reduced pressure, and the residue was taken up
in water and ethyl acetate and stirred for another 1 h. The
precipitate was filtered off and discarded, the phases were
separated, and the aqueous layer was extracted twice with ethyl
acetate. The combined organic phases were dried and evaporated
affording 2.45 g (92% purity by LC-MS, 92% of th.) of the title
compound.
[0590] LC-MS (method 5): R.sub.t=2.18 min; MS (ESIpos): m/z=293
(M+H).sup.+
[0591] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.98-7.78 (m,
3H), 7.04 (s, 1H), 4.68 (s, 2H), 0.89 (s, 9H), 0.09 (s, 6H)
ppm.
Intermediate 52A
(4-Amino-7-methylpyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00096##
[0592] Method 1:
[0593] To a solution of 100 mg (0.28 mmol) of Intermediate 35A in 2
ml 1,4-dioxane under an argon atmosphere were added 6 mg (0.01
mmol) PdCl.sub.2(dppf).times.DCM and then dropwise over 10 min 0.56
ml of a 2 M solution of dimethylzinc in toluene. The mixture was
stirred at 90.degree. C. overnight, then evaporated, and the
residue was treated with acetonitrile and 5 M aq. formic acid. The
precipitate was filtered off, the filtrate was evaporated, and the
residue was taken up in DMSO/acetonitrile and purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid). Yield: 33 mg (66% of th.).
[0594] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.79 (s, 1H),
7.47 (br. s, 2H), 6.80 (s, 1H), 4.84 (t, 1H), 4.49 (d, 2H), 2.36
(s, 3H) ppm.
Method 2:
[0595] Under an argon atmosphere, a microwave reaction vessel was
charged with 750 mg (3.1 mmol) of Intermediate 34A, 515 .mu.l (3.7
mmol) trimethylboroxine, 786 mg (3.7 mmol) potassium phosphate and
73 mg (0.09 mmol)
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-trii-
sopropylbiphenyl-2-yl)phosphine (1:1; see S. L. Buchwald et al., J.
Am. Chem. Soc. 132 (40), 14073-14075 (2010)). Then, 13 ml of a
degassed 1,4-dioxane/water mixture (5:1) were added, the vessel was
sealed, and the mixture was heated to 140.degree. C. for 20 min in
the microwave (4 bar, 50 watt). Further 50 mg
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1) were added, and the mixture was
heated again to 140.degree. C. for 20 min in the microwave. Another
portion of 515 .mu.l trimethylboroxine was added, and the vessel
was heated again to 140.degree. C. for 20 min. The latter procedure
was repeated two more times with heating periods of 30 min each
until LC-MS showed only minor amounts of starting material left.
The mixture was filtered over kieselguhr, washed with 1,4-dioxane,
and the combined filtrates were evaporated to dryness. This residue
was combined with a previous 100 mg test run and purified by
preparative RP-HPLC (XBridge C18, gradient 5-42% acetonitrile/0.05%
aq. ammonium hydroxide solution). Yield: 238 mg (38% of th.).
[0596] LC-MS (method 5): R.sub.t=0.51 min; MS (ESIneg): m/z=177
(M-H).sup.1H-NMR
[0597] (400 MHz, DMSO-d.sub.6): .delta.=7.79 (s, 1H), 7.48 (br. s,
2H), 6.80 (s, 1H), 4.84 (t, 1H), 4.49 (d, 2H), 2.36 (s, 3H)
ppm.
Intermediate 53A
(4-Amino-5-bromo-7-methylpyrrolo[2,1-f][1,2,4]triazin-6-yl)methanol
##STR00097##
[0599] A solution of 245 mg (1.38 mmol) of Intermediate 52A in 8.8
ml DMF was treated with 288 mg (1.62 mmol) N-bromosuccinimide in
portions over 2 h at -10.degree. C. Stirring was continued at
-10.degree. C. for 30 min and then for 2 h at ambient temperature.
The mixture was poured into 50 ml water and extracted with ethyl
acetate. The combined organic phases were dried and evaporated. The
residue was flash-chromatographed on silica gel with
dichloromethane/0-15% methanol as eluent. Yield: 148 mg (42% of
th.).
[0600] LC-MS (method 4): R.sub.t=0.42 min; MS (ESIpos): m/z=257/259
(M+H).sup.+
[0601] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.85 (s, 1H),
6.14-8.22 (broad, 2H), 4.88 (t, 1H), 4.45 (d, 2H), 2.43 (s, 3H)
ppm.
Intermediate 54A
4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-methylpyrrolo[2,1-f]-
[1,2,4]triazine-6-carbaldehyde
##STR00098##
[0603] To a suspension of 175 mg (0.49 mmol) of Example 75 and
molecular sieves (3 .ANG.) in 3.5 ml dichloromethane were added 230
mg (0.54 mmol) Dess-Martin periodinane
(1,1,1-triacetoxy-1,1-dihydro-1,2-benziodoxol-3(1H)-one) at
0-5.degree. C. The mixture was stirred for 5 min at this
temperature, followed by 30 min at rt. After this, the mixture was
adsorbed on diatomaceous earth and purified by flash-chromatography
on silica gel with isohexane/10-100% ethyl acetate as eluent.
Yield: 139 mg of a solid (79% of th.).
[0604] LC-MS (method 5): R.sub.t=2.34 min; MS (ESIpos): m/z=353
(M+H).sup.+
[0605] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=9.93 (s, 1H),
8.06 (s, 1H), 7.50 (s, 1H), 7.33 (s, 1H), 6.88 (s, 1H), 3.96 (s,
3H), 2.71 (s, 3H), 2.46 (s, 3H) ppm.
Intermediate 55A
tert-Butyl
4-[(4-amino-6-methylpyrrolo[2,1-f][1,2,4]triazin-7-yl)methyl]pi-
perazine-1-carboxylate
##STR00099##
[0607] A solution of 6-methylpyrrolo[2,1-f][1,2,4]triazin-4-amine
(500 mg, 3.3 mmol; preparation described in PCT Int. Pat. Appl. WO
2007/056170) in acetic acid (8 ml) was treated with 37% aq.
formaldehyde solution (328 .mu.l, 4.04 mmol) and tert-butyl
piperazine-1-carboxylate (754 mg, 4.04 mmol). The mixture was
stirred at 60.degree. C. overnight. After evaporation, the residue
was taken up in ethyl acetate and washed with sat. aq. sodium
hydrogencarbonate solution. The aqueous layer was extracted twice
with ethyl acetate. The combined organic phases were dried over
magnesium sulfate and evaporated affording 1.2 g of the crude
product which was used in the next step without further
purification.
[0608] LC-MS (method 2): R.sub.t=0.56 min; MS (ESIpos): m/z=347
(M+H).sup.+
[0609] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.79 (s, 1H),
7.51 (br. s, 2H), 6.67 (s, 1H), 3.76 (br. s, 2H), 3.29-3.17 (m,
4H), 2.36-2.29 (m, 4H), 2.22 (s, 3H), 1.37 (s, 9H) ppm.
Intermediate 56A
tert-Butyl
4-[(4-amino-5-bromo-6-methylpyrrolo[2,1-f][1,2,4]triazin-7-yl)m-
ethyl]piperazine-1-carboxylate
##STR00100##
[0611] A solution of Intermediate 55A (1.17 g, 3.37 mmol) in THF
(20 ml) was cooled to -60.degree. C. and treated with
1,3-dibromo-5,5-dimethylhydantoin (5.78 mg, 2.02 mmol). The mixture
was stirred for 4 h at -60.degree. C. to -20.degree. C. After this,
the reaction mixture was quenched with 10% aq. sodium thiosulfate
solution. Most of the THF solvent was evaporated whereupon a solid
precipitated. Filtration and recrystallization from acetone
afforded 862 mg (59% of th.) of the title compound.
[0612] LC-MS (method 2): R.sub.t=0.66 min; MS (ESIpos): m/z=425/427
(M+H).sup.+
[0613] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.29-7.62 (br.
s, 1H), 7.85 (s, 1H), 7.17-6.44 (br. s, 1H), 3.80 (s, 2H),
3.29-3.22 (m, 4H), 2.37-2.27 (m, 4H), 2.16 (s, 3H), 1.37 (s, 9H)
ppm.
Intermediate 57A
tert-Butyl
4-{[4-amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-6-methy-
lpyrrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazine-1-carboxylate
##STR00101##
[0615] A solution of Intermediate 56A (100 mg, 235 .mu.mol) in
degassed 1,4-dioxane (3 ml) was treated with Intermediate 6A (93
mg, 282 .mu.mol),
dicyclohexyl(2',6'-dimethoxybiphenyl-2-yl)phosphine (S-Phos; 9.6
mg, 23 .mu.mol) and palladium diacetate (2.6 mg, 11 .mu.mol).
Degassed 3 M aq. potassium phosphate solution (588 .mu.l) was
added, and the resulting mixture was stirred at 60.degree. C. for 1
h. A further portion of Intermediate 6A (78 mg, 235 .mu.mol) was
added, and stirring at 60.degree. C. was continued overnight. The
reaction mixture was quenched with 2 M aq. sodium hydroxide
solution, and the aqueous phase was extracted with ethyl acetate.
The combined organic phases were washed with sat. aq. sodium
chloride solution, dried over magnesium sulfate and evaporated.
Purification by column chromatography on silica gel
(cyclohexane/ethyl acetate 3:2) afforded 82 mg (62% of th.) of the
title compound.
[0616] LC-MS (method 2): R.sub.t=0.97 min; MS (ESIpos): m/z=523
(M+H).sup.+
[0617] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): 5=7.94 (s, 1H), 7.34
(s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 3.95 (s, 3H), 3.84 (s, 2H),
3.30-3.23 (m, 4H, overlap with water peak), 2.45 (s, 3H), 2.42-2.33
(m, 4H), 2.19 (s, 3H), 1.39 (s, 9H) ppm.
Intermediate 58A
tert-Butyl
4-[(4-amino-6-chloropyrrolo[2,1-f][1,2,4]triazin-7-yl)methyl]pi-
perazine-1-carboxylate
##STR00102##
[0619] Following the procedure of Intermediate 55A, 4 g (23.7 mmol)
6-chloropyrrolo[2,1-f][1,2,4]triazin-4-amine (preparation described
in PCT Int. Pat. Appl. WO 2007/064883) were reacted to give 11.2 g
of the title compound as crude material which was used in the next
step without further purification.
[0620] LC-MS (method 5): R.sub.t=0.74 min; MS (ESIpos): m/z=367
(M+H).sup.+
[0621] A sample of 67 mg of the corresponding formiate salt,
tert-butyl
4-[(4-amino-6-chloropyrrolo[2,1-f]-[1,2,4]triazin-7-yl)methyl]piperazine--
1-carboxylate formiate, was isolated after preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic acid)
of 100 mg of the crude material obtained above.
[0622] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.14
(s, 1H), 7.91 (s, 1H), 7.83 (br. s, 2H), 6.95 (s, 1H), 3.80 (s,
2H), 3.17 (s, 2H), 2.38 (br. s, 4H), 1.37 (s, 9H) ppm.
Intermediate 59A
tert-Butyl
4-[(4-amino-5-bromo-6-chloropyrrolo[2,1-f][1,2,4]triazin-7-yl)m-
ethyl]piperazine-1-carboxylate
##STR00103##
[0624] Following the procedure of Intermediate 56A, 11 g (30 mmol)
of Intermediate 58A were reacted to give 1.17 g (9% of th.) of the
title compound after flash-chromatography on silica gel
(dichloromethane/methanol 10:1) and subsequent preparative RP-HPLC
(Daiso C18, gradient 40-65% acetonitrile/water).
[0625] LC-MS (method 2): R.sub.t=0.74 min; MS (ESIpos):
m/z=445/447/449 (M+H).sup.+
[0626] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=7.96
(s, 1H), 3.84 (s, 2H), 2.37 (br. s, 4H), 1.37 (s, 9H) ppm.
Intermediate 60A
tert-Butyl
4-{[4-amino-6-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl-
)pyrrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazine-1-carboxylate
##STR00104##
[0628] Under an argon atmosphere, a flask was charged with 140 mg
(0.31 mmol) of Intermediate 59A, 102 mg (0.46 mmol) of Intermediate
5A, 16 mg (0.02 mmol) PdCl.sub.2(dppf).times.DCM and 122 mg (1.15
mmol) sodium carbonate. After addition of 3 ml degassed
1,2-dimethoxyethane/water (3:1), the suspension was stirred at
60.degree. C. for 2.5 h. Further portions of Intermediate 5A (50
mg, 0.23 mmol) and PdCl.sub.2(dppf).times.DCM (8 mg, 0.01 mmol)
were added, and stirring was continued at 40.degree. C. for 2.5 h.
The latter procedure was repeated once more until the starting
material was consumed. Then, the reaction mixture was partially
evaporated under reduced pressure, water was added, and the mixture
was extracted with dichloromethane. The combined organic phases
were washed with water and sat. aq. sodium chloride solution, dried
and evaporated under reduced pressure. The residue (322 mg) was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) to give 49 mg of a mixture of
the title compound and the Boc-deprotected derivative,
6-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmet-
hyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine. This mixture was used as
such in the next reaction step (see Example 85).
[0629] LC-MS (method 2): R.sub.t=1.00 min; MS (ESIpos): m/z=543
(M+H).sup.+, and R.sub.t=0.81 min; MS (ESIpos): m/z=443
(M+H).sup.+.
Intermediate 61A
4-{[4-Amino-6-(methoxymethyl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}pipe-
razin-2-one
##STR00105##
[0631] A solution of Intermediate 10A (5 g, 28.1 mmol) and
piperazin-2-one (3.09 g, 30.9 mmol) in acetic acid (85 ml) was
treated with 37% aq. formaldehyde solution (3.15 ml, 42.1 mmol) and
stirred at 60.degree. C. for 16 h. The volatiles were evaporated
under reduced pressure, and the residue was dissolved in methanol
and adsorbed on diatomaceous earth. Purification by column
chromatography on silica gel (5-10% methanol/dichloromethane)
afforded 3.91 g (46% of th.) of the title compound.
[0632] LC-MS (method 3): R.sub.t=1.68 min; MS (ESIpos): m/z=291
(M+H).sup.+.
Intermediate 62A
4-{[4-Amino-5-bromo-6-(methoxymethyl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]met-
hyl}piperazin-2-one
##STR00106##
[0634] A solution of Intermediate 61A (3.9 g, 13.4 mmol) in DMF (50
ml) was cooled to 0.degree. C. and treated with a solution of
N-bromosuccinimide (2.63 g, 14.8 mmol) in DMF (6 ml). The mixture
was stirred at 0.degree. C. for 1 h. Then, the solvent was
evaporated, and the residue was dissolved in methanol and adsorbed
on diatomaceous earth. Purification by column chromatography on
silica gel (5-10% methanol/dichloromethane) afforded 1.99 g (39% of
th.) of the title compound.
[0635] LC-MS (method 3): R.sub.t=1.86 min; MS (ESIpos): m/z=369/371
(M+).sup.+
[0636] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96-8.23 (br.
s, 1H), 7.90 (s, 1H), 7.71 (s, 1H), 6.72-6.98 (br. s, 1H), 4.45 (s,
2H), 3.91 (s, 2H), 3.27 (s, 3H), 3.07 (br. s, 2H), 2.96 (s, 2H),
2.56 (br. s, 2H) ppm.
Intermediate 63A
7-(Chloromethyl)-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2--
yl)pyrrolo[2,1-f]-[1,2,4]triazin-4-amine hydrochloride
##STR00107##
[0638] A suspension of Example 86 (1 g, 2.51 mmol) in toluene (60
ml) was treated dropwise with thionyl chloride (1.83 ml, 25.1
mmol), and the mixture was stirred at rt overnight. The volatiles
were evaporated under reduced pressure. The residue was
co-evaporated with toluene under reduced pressure for three times
affording 0.85 g (74% of th.) of the title compound which was
immediately used for the next step without further
purification.
Intermediate 64A
4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrol-
o[2,1-f][1,2,4]-triazine-7-carboxylic acid
##STR00108##
[0640] A suspension of Intermediate 17A (5 g, 12.6 mmol) in
THF/water (10:1, 220 ml) was treated with a 2 M solution of
2-methyl-2-butene in THF (31.5 ml, 63.1 mmol) and with sodium
dihydrogenphosphate (6.96 g, 50.4 mmol). The mixture was stirred at
rt for 5 min. Then, sodium chlorite (4.56 g, 50.44 mmol) was added,
and the resulting mixture was stirred at rt for 20 h. The
suspension was filtered, and the resulting solid was washed with
water affording 4.24 g (74% of th.) of the title compound which was
used in the next step without further purification.
[0641] LC-MS (method 4): R.sub.t=1.06 min; MS (ESIpos): m/z=413
(M+H).sup.+
[0642] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=13.22 (br. s,
1H), 8.53-8.00 (br. s, 1H), 8.14 (s, 1H), 7.41 (s, 1H), 7.33 (s,
1H), 6.86 (s, 1H), 6.28-5.65 (br. s, 1H), 4.61 (s, 2H), 3.96 (s,
3H), 3.36 (q, 2H), 2.46 (s, 3H), 1.01 (t, 3H) ppm.
Intermediate 65A
5,7-Dimethoxy-1-benzothiophene
##STR00109##
[0644] To a solution of 1-benzothiophene-5,7-diol (1.16 g, 6.98
mmol) in acetone (20 ml) under argon were added potassium carbonate
(2.89 g, 20.9 mmol) and iodomethane (912 .mu.l, 14.6 mmol). The
resulting mixture was stirred under reflux for 18 h. After cooling
to rt, the mixture was treated with a 7 M solution of ammonia in
methanol (10 ml) for 30 min and then adsorbed on silica gel.
Purification by column chromatography over silica gel
(cyclohexane/ethyl acetate 40:1) afforded 0.52 g (32% of th.) of
the title compound.
[0645] LC-MS (method 4): R.sub.t=1.02 min; MS (ESIpos): m/z=195
(M+H).sup.+
[0646] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.69 (d, 1H),
7.35 (d, 1H), 7.02 (d, 1H), 6.57 (d, 1H), 3.92 (s, 3H), 3.81 (s,
3H) ppm.
Intermediate 66A
(5,7-Dimethoxy-1-benzothiophen-2-yl)boronic acid
##STR00110##
[0648] Under an argon atmosphere, a 1.6 M solution of
n-butyllithium in hexane (1.84 ml, 2.95 mmol) was added dropwise to
a solution of Intermediate 65A (520 mg, 2.68 mmol) in dry THF (5
ml) at -70.degree. C. After 1 h at -70.degree. C., triisopropyl
borate (742 .mu.l, 3.21 mmol) was added, and the mixture was
stirred for 16 h while slowly warming up to rt. Dichloromethane and
sat. aq. ammonium chloride solution were added, and the pH value
was adjusted to 6 by addition of 1 M hydrochloric acid. The organic
phase was separated, and the aqueous phase was extracted with
dichloromethane. The combined organic phases were dried with
magnesium sulfate, filtered and evaporated. The resulting residue
was purified by column chromatography over silica gel (at first
eluting with dichloromethane/methanol 40:1, then methanol, finally
methanol/4 M hydrogen chloride in 1,4-dioxane 10:1) yielding 631 mg
(710% purity, 71% of th.) of the title compound.
[0649] LC-MS (method 4): R.sub.t=0.83 min; MS (ESIpos): m/z=239
(M+H).sup.+.
Intermediate 67A
4-Amino-6-(chloromethyl)pyrrolo[2,1-f][1,2,4]triazine-7-carbaldehyde
##STR00111##
[0651] Intermediate 67A was isolated as a side-product of the
synthesis of Intermediate 11A after column chromatography on silica
gel (dichloromethane/acetone 8:2.fwdarw.7:3).
[0652] LC-MS (method 2): R.sub.t=0.6 min; MS (ESIpos): m/z=211/213
(M+H).sup.+.
Intermediate 68A
4-Amino-6-[(3-oxopiperazin-1-yl)methyl]pyrrolo[2,1-f][1,2,4]triazine-7-car-
baldehyde
##STR00112##
[0654] A solution of Intermediate 67A (11.03 g, 52.4 mmol) and
2-oxopiperazine (6.82 g, 68.1 mmol) in 331 ml DMF was treated at rt
with DIPEA (13.7 ml, 78.6 mmol) and stirred overnight. The
precipitate was filtered off, washed with DMF and diethylether and
then dried in vacuo to yield 11.64 g of the title compound (89%
purity, 72% of th.).
[0655] LC-MS (method 7): R.sub.t=1.30 min; MS (ESIpos): m/z=275
(M+H).sup.+.
Intermediate 69A
4-{[4-Amino-7-(hydroxymethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}pipe-
razin-2-one
##STR00113##
[0657] To a solution of Intermediate 68A (10.17 g, 89% purity,
33.01 mmol) in 1 M hydrochloric acid (370 ml) and methanol (370 ml)
at rt was added zinc dust (12.1 g, 185 mmol), and the mixture was
stirred at rt for 18 h. Silica gel (100 g) was added, and the
volatiles were evaporated under reduced pressure. The residue was
suspended in methanol, the volatiles were evaporated under reduced
pressure again, and the residue was dried in vacuo. The solid was
subjected to column chromatography on silica gel (gradient
dichloromethane/7 M ammonia in methanol 10:1.fwdarw.3:1) to yield
6.39 g of the title compound (60% of th.).
[0658] LC-MS (method 8): R.sub.t=1.12 min; MS (ESIpos): m/z=277
(M+H).sup.+
[0659] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.83 (s, 1H),
7.76 (br. s, 1H), 7.64 (br. s, 2H), 6.83 (s, 1H), 5.01 (br. s, 1H),
4.74 (s, 2H), 3.64 (br. s, 2H), 3.14 (br. s, 2H), 2.95 (br. s, 2H),
2.58 (br. s, 2H) ppm.
Intermediate 70A
4-{[4-Amino-5-bromo-7-(hydroxymethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]met-
hyl}piperazin-2-one trifluoroacetate
##STR00114##
[0661] A suspension of Intermediate 69A (1 g) in methanol/water
(10:1, 33 ml) was treated with trifluoroacetic acid (0.56 ml, 7.24
mmol) to result in a clear solution. A solution of
N-bromosuccinimide (708 mg, 3.98 mmol) in methanol (30 ml) was
added dropwise at 0.degree. C., and the mixture was stirred at
0.degree. C. for 1 h. The formed precipitate was filtered off and
dried in vacuo to yield 700 mg of the title compound (41% of
th.).
[0662] LC-MS (method 7): R.sub.t=1.38 min; MS (ESIpos): m/z=355/357
(M+H).sup.+
[0663] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=7.90 (s, 1H), 5.00
(s, 2H), 4.44 (s, 2H), 3.85 (s, 2H), 3.58 (br. t, 2H), 3.50 (br. t,
2H) ppm.
PREPARATION EXAMPLES
Example 1
4-{[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)p-
yrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}piperazin-2-one
##STR00115##
[0664] Method 1:
[0665] A solution of Intermediate 13A (3 g, 7.84 mmol) in methanol
(87 ml) was treated with acetic acid (0.898 ml, 15.68 mmol),
2-oxopiperazine (1.17 g, 11.76 mmol) and sodium
triacetoxyborohydride (4.98 g, 23.53 mmol). The mixture was stirred
at rt for 4.5 h. Further portions of 2-oxopiperazine (392 mg, 3.9
mmol) and sodium triacetoxyborohydride (3.3 g, 15.68 mmol) were
added, and the resulting mixture was stirred at 60.degree. C.
overnight. After evaporation, the residue was taken up in sat. aq.
sodium hydrogencarbonate solution and extracted with ethyl acetate.
The combined organic phases were dried over sodium sulfate and
evaporated, and the residue was purified by column chromatography
on silica gel (dichloromethane/methanol 40:1.fwdarw.10:1). The
product thus obtained was triturated in methanol and filtered off
affording 540 mg (14% of th.) of the title compound. The methanolic
mother liquor was evaporated and the residue purified by two-fold
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid). The fractions containing pure product were combined and
neutralized with sat. aq. sodium hydrogencarbonate solution. The
acetonitrile solvent was evaporated, and the aqueous phase was
extracted with ethyl acetate. The combined organic phases were
dried over sodium sulfate and evaporated affording 395 mg (13% of
th.) as a second batch of the title compound.
Method 2:
[0666] A solution of Intermediate 21A (291 mg, 0.82 mmol) in acetic
acid (2.9 ml) was treated with 37% aq. formaldehyde solution (104
.mu.l, 1.39 mmol) and 2-oxopiperazine (139 mg, 1.39 mmol). The
mixture was stirred at 60.degree. C. for 3 h and then evaporated.
Purification by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 184 mg (44% of th.) of
the title compound.
[0667] LC-MS (method 4): R.sub.t=0.77 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0668] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.73 (br. s, 1H), 7.37 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H), 4.41
(s, 2H), 3.97 (s, 2H), 3.96 (s, 3H), 3.20 (s, 3H), 3.15-3.08 (br.
s, 2H), 3.01 (s, 2H), 2.64 (br. t, 2H), 2.45 (s, 3H) ppm.
Example 2
4-{[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)p-
yrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piperazin-2-one
dihydrochloride
##STR00116##
[0670] A solution of Example 1 (100 mg, 214 .mu.mol) in 1,4-dioxane
(2 ml) was treated with a 4 M solution of hydrogen chloride in
1,4-dioxane (2 ml, 8 mmol). The solvent was evaporated leaving 130
mg (quant.) of the title compound.
[0671] LC-MS (method 2): R.sub.t=0.79 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0672] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.39 (br. s,
1H), 8.31-8.53 (br. s, 1H), 8.17 (s, 1H), 7.38 (s, 1H), 7.34 (s,
1H), 6.87 (s, 1H), 6.21-6.42 (br. s, 1H), 4.81 (br. s, 2H), 4.52
(br. s, 2H), 3.96 (s, 3H), 3.57 (s, 2H), 3.32-3.72 (m, 4H), 3.26
(s, 3H), 2.46 (s, 3H) ppm.
Example 3
(3R)-3-({[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen--
2-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
dihydrochloride
##STR00117##
[0674] A solution of Intermediate 13A (520 mg, 1.36 mmol) in
methanol (15 ml) and acetic acid (156 .mu.l, 2.7 mmol) was treated
with (R)-3-aminopyrrolidin-2-one (503 mg, 5.0 mmol) and
triacetoxyborohydride (1.06 g, 5.0 mmol). The mixture was stirred
at rt overnight and then evaporated. Purification by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) and lyophilization of the product thus obtained from a 4 M
solution of hydrogen chloride in 1,4-dioxane afforded 272 mg (36%
of th.) of the title compound.
[0675] LC-MS (method 2): R.sub.t=0.70 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0676] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): d=9.62 (br. s, 1H),
9.46 (br. s, 1H), 8.56-8.22 (br. s, 1H), 8.44 (s, 1H), 8.19 (s,
1H), 7.38 (s, 1H), 7.34 (s, 1H), 6.87 (s, 1H), 6.54-6.12 (br. s,
1H), 4.89-4.65 (m, 2H), 4.58-4.46 (m, 2H), 4.17-4.07 (br. s, 1H,
overlap with water peak), 3.96 (s, 3H), 3.36-3.16 (m, 2H), 3.25 (s,
3H), 2.48-2.39 (m, 1H), 2.46 (s, 3H), 2.23-2.06 (m, 1H) ppm.
Example 4
(3R)-3-({[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen--
2-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
##STR00118##
[0678] A solution of Intermediate 13A (2.0 g, 5.2 mmol) in methanol
(58 ml) and acetic acid (0.6 ml) was treated with
(R)-3-aminopyrrolidin-2-one (785 mg, 7.8 mmol) and
triacetoxyborohydride (3.32 g, 15.6 mmol). The mixture was stirred
at rt overnight. After this, the reaction mixture was diluted with
sat. aq. sodium hydrogencarbonate solution and extracted three
times with ethyl acetate. The combined organic phases were dried
over sodium sulfate and evaporated. The residue was purified by
two-fold column chromatography on silica gel
(dichloromethane/methanol 40:1 to 10:1) to afford 957 mg (37% of
th.) of the title compound.
[0679] LC-MS (method 4): R.sub.t=0.72 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0680] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.01 (s, 1H),
7.75 (s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.46-4.34
(m, 2H), 4.24-4.02 (m, 2H), 3.96 (s, 3H), 3.24-3.04 (m, 3H), 3.21
(s, 3H), 2.45 (s, 3H), 2.40-2.27 (m, 1H), 1.80-1.65 (m, 1H)
ppm.
Example 5
4-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazin-2-one
##STR00119##
[0681] Method 1:
[0682] A solution of Intermediate 17A (2 g, 5.05 mmol) in THF (100
ml) was treated at 0.degree. C. with 2-oxopiperazine (1.01 g, 10.1
mmol), sodium triacetoxyborohydride (1.07 g, 5.04 mmol) and acetic
acid (0.29 ml, 5.04 mmol). The resulting mixture was stirred at
0.degree. C. for 30 min. Four additional portions of sodium
triacetoxyborohydride (1.07 g, 5.04 mmol) and of acetic acid (0.29
ml, 5.04 mmol) were added every 30 minutes, and the resulting
mixture was stirred for further 30 min at 0.degree. C., then for 25
min at 35.degree. C. and finally at rt overnight. The reaction was
quenched with 10% aq. sodium chloride solution and extracted twice
with ethyl acetate. The combined organic layers were evaporated.
The residue was purified by column chromatography over silica gel
(dichloromethane/methanol 95:5.fwdarw.90:10) affording 360 mg (17%
of th.) of the compound described in Example 86 (see below) and
1.82 g of the title compound in separate fractions. The title
product thus obtained was suspended in ethanol (20 ml), refluxed
for 2 h and then cooled to 15.degree. C. The solid was filtered off
and washed with ethanol affording 1.63 g (67% of th.) of the pure
title compound.
Method 2:
[0683] A solution of Example 13 (930 mg, 1.9 mmol) in
dichloromethane (18 ml) was treated with thionyl chloride (210
.mu.l, 2.8 mmol) and stirred at rt for 15 min. After evaporation,
the residue was dissolved in ethanol (18 ml) and treated with DIPEA
(670%, 3.8 mmol). The mixture was stirred at 70.degree. C. for 2 h
and then evaporated. The residue was purified by column
chromatography on silica gel (dichloromethane/methanol
98:2.fwdarw.90:10). The product thus obtained was triturated in an
acetonitrile/diethylether mixture and filtered. The filtrate was
evaporated, and the residue was re-purified by column
chromatography on silica gel (dichloromethane/methanol
98:2.fwdarw.90:10). Again, the product thus obtained was triturated
in acetonitrile/diethylether and filtered. This procedure was
repeated one more time. The three batches of solids obtained in
this way were combined, triturated in acetonitrile/diethylether
once again and finally filtered off affording 600 mg (62% of th.)
of the title compound.
Method 3:
[0684] A solution of Intermediate 22A (720 mg, purity 89%, 1.74
mmol) in acetic acid (10 ml) was stirred with piperazin-2-one (261
mg, 2.61 mmol) at 60.degree. C. To this, 37% aq. formaldehyde
solution (260 .mu.l, 3.48 mmol) was added in three portions after
0, 3 and 12 h, respectively, and the mixture was stirred at
60.degree. C. for a total of 24 h. Then, the volatiles were removed
under reduced pressure, and the residue was partitioned between
ethyl acetate and sat. aq. sodium hydrogencarbonate solution. The
organic phase was washed with sat. aq. sodium chloride solution,
dried with magnesium sulfate and evaporated. The residue was
dissolved in a mixture of methanol and dichloromethane, adsorbed on
diatomaceous earth, dried in vacuo and purified by
flash-chromatography on silica gel (gradient 0-6%
methanol/dichloromethane). The product fractions were combined,
evaporated and re-purified by preparative RP-HPLC (Reprosil C18,
gradient 30-50% acetonitrile/0.2% aq. TFA). The product fractions
were combined again, diluted with sat. aq. sodium hydrogencarbonate
solution and extracted with ethyl acetate. The organic phase was
washed with sat. aq. sodium chloride solution, dried with magnesium
sulfate and evaporated yielding 281 mg (31% of th.) of the title
compound.
[0685] LC-MS (method 2): R.sub.t=0.86 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0686] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.01 (s, 1H),
7.8-8.05 (br. s, 1H), 7.74 (br. s, 1H), 7.38 (s, 1H), 7.31 (s, 1H),
6.85 (s, 1H), 5.6-5.9 (br. s, 1H), 4.45 (s, 2H), 3.97 (s, 2H), 3.94
(s, 3H), 3.41 (q, 2H), 3.09-3.13 (m, 2H), 3.02 (s, 2H), 2.63-2.69
(br. s, 2H), 2.45 (s, 3H), 1.07 (t, 3H) ppm.
Example 6
4-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazin-2-one
dihydrochloride
##STR00120##
[0688] A solution of Intermediate 17A (60 mg, purity 69%, 104
.mu.mol) in methanol (6 ml) was treated with 2-oxopiperazine (22
mg, 209 .mu.mol), sodium cyanoborohydride (33 mg, 522 mmol) and
acetic acid (12 .mu.l, 209 .mu.mol). The mixture was stirred at
60.degree. C. for 16 h and then filtered. The filtrate and the
residue were purified separately by preparative RP-HPLC (Reprosil
C18, gradient 40-60% acetonitrile/0.2% aq. TFA). The product
fractions were combined, diluted with 1 M hydrochloric acid (3 ml)
and evaporated to dryness yielding 46 mg (79% of th.) of the title
compound.
[0689] LC-MS (method 2): R.sub.t=0.87 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0690] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.37 (br. s,
1H), 8.31 (br. s, 1H), 8.14 (s, 1H), 7.38 (s, 1H), 7.33 (s, 1H),
6.87 (s, 1H), 6.13 (br. s, 1H), 4.80 (br. s, 2H), 4.52 (s, 2H),
3.96 (s, 3H), 3.86 (br. s, 2H), 3.36-3.51 (m, 5H), 2.46 (s, 3H),
1.11 (t, 3H) ppm.
Example 7
(3R)-3-({[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-
-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
dihydrochloride
##STR00121##
[0692] A solution of Intermediate 17A (60 mg, purity 69%, 104
.mu.mol) in methanol (3 ml) was treated with
(R)-3-aminopyrrolidin-2-one (22 mg, 209 .mu.mol), sodium
cyanoborohydride (33 mg, 522 .mu.mol) and acetic acid (12 .mu.l,
209 .mu.mol). The mixture was stirred at 60.degree. C. for 4 h and
then filtered. The filtrate was purified by preparative RP-HPLC
(Reprosil C18, gradient 40-60% acetonitrile/0.2% aq. TFA). The
product fractions were combined, diluted with 1 M hydrochloric acid
and evaporated to dryness yielding 46 mg (79% of th.) of the title
compound.
[0693] LC-MS (method 4): R.sub.t=0.74 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0694] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=9.58-9.75 (m,
1H), 9.33-9.54 (m, 1H), 8.44 (s, 1H), 8.27-8.71 (br. s, 1H), 8.19
(s, 1H), 7.38 (s, 1H), 7.34 (s, 1H), 6.87 (s, 1H), 6.18-6.48 (br.
s, 1H), 4.68-4.87 (m, 2H), 4.49-4.62 (q, 2H), 3.96 (s, 3H), 3.45
(q, 2H), 3.18-3.35 (m, 2H), 2.46 (s, 4H), 2.08-2.22 (m, 1H), 1.10
(t, 3H) ppm.
Example 8
(3R)-3-({[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-
-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
##STR00122##
[0696] A solution of Intermediate 17A (226 mg, purity 75%, 428
.mu.mol) in methanol (4 ml) was treated with
(R)-3-aminopyrrolidin-2-one (85 mg, 855 .mu.mol), sodium
cyanoborohydride (134 mg, 2.14 mmol) and acetic acid (49 .mu.l, 855
.mu.mol). The mixture was stirred at rt for 1.5 h. After this, the
mixture was directly separated by preparative RP-HPLC (Reprosil
C18, gradient 40-60% acetonitrile/0.2% aq. TFA). The product
fractions were combined, diluted with sat. aq. sodium
hydrogencarbonate solution and extracted with ethyl acetate. The
combined organic phases were washed with sat. aq. sodium chloride
solution, dried over magnesium sulfate and evaporated to dryness
yielding 180 mg (88% of th.) of the title compound.
[0697] LC-MS (method 4): R.sub.t=0.74 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0698] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.76 (s, 1H), 7.8-8.1 (br. s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.84
(s, 1H), 5.6-5.9 (br. s, 1H), 4.39-4.49 (m, 2H), 4.04-4.23 (m, 2H),
3.96 (s, 3H), 3.41 (q, 2H), 3.05-3.23 (m, 3H), 2.45 (s, 3H),
2.31-2.40 (m, 1H), 1.68-1.79 (m, 1H), 1.08 (t, 3H) ppm.
Example 9
N.sup.2-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-
-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}glycinamide
dihydrochloride
##STR00123##
[0700] A solution of Intermediate 17A (60 mg, purity 69%, 104
.mu.mol) in methanol (3 ml) was treated with glycinamide
hydrochloride (23 mg, 209 .mu.mol), sodium cyanoborohydride (32 mg,
522 mmol) and acetic acid (12 .mu.l, 209 .mu.mol). The mixture was
stirred at 60.degree. C. for 16 h. After filtration, the filtrate
was separated by preparative RP-HPLC (Reprosil C18, gradient 20-40%
acetonitrile/0.2% aq. TFA). The product fractions were combined,
diluted with 1 M hydrochloric acid, evaporated to dryness and
combined with the residue from the filtration step. This material
was re-purified by two-fold preparative RP-HPLC (Reprosil C18,
gradient 20-40% acetonitrile/0.2% aq. TFA). The product fractions
were combined again, diluted with 1 M hydrochloric acid and
evaporated to dryness yielding 7.4 mg (13% of th.) of the title
compound.
[0701] LC-MS (method 2): R.sub.t=0.80 min; MS (ESIpos): m/z=455
(M+H).sup.+
[0702] .sup.1H-NMR (500 MHz, DMSO-d.sub.6): .delta.=9.33 (br. s,
2H), 9.05 (br. s, 1H), 8.31 (s, 1H), 7.93 (br. s, 1H), 7.58 (br. s,
1H), 7.40 (s, 1H), 7.35 (s, 1H), 7.05 (br. s, 1H), 6.88 (s, 1H),
4.65 (br. m, 2H), 4.56 (s, 2H), 3.96 (s, 3H), 3.74 (br. m, 2H),
3.46 (q, 2H), 2.46 (s, 3H), 1.10 (t, 3H) ppm.
Example 10
6-(Ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin--
4-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00124##
[0704] A solution of Intermediate 17A (55 mg, purity 73%, 101
.mu.mol) in methanol (3 ml) was treated with morpholine (18 mg, 202
.mu.mol), sodium cyanoborohydride (19 mg, 303 mmol) and acetic acid
(18 .mu.l, 304 .mu.mol). The mixture was stirred at 60.degree. C.
for 18 h. Further amounts of morpholine (18 mg, 202 .mu.mol),
sodium cyanoborohydride (19 mg, 303 mmol) and acetic acid (18
.mu.l, 304 .mu.mol) were added, and stirring at 60.degree. C. was
continued for another 3 h. The resulting mixture was diluted with
THF to dissolve precipitates and separated by preparative RP-HPLC
(Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA). The
product fractions were combined and evaporated to dryness. The
residue was dissolved in methanol and filtered through an anion
exchange cartridge (Stratospheres SPE, PL-HCO.sub.3 MP-resin). The
cartridge was eluted with methanol, and the filtrate was evaporated
yielding 32 mg (68% of th.) of the title compound.
[0705] LC-MS (method 2): R.sub.t=0.78 min; MS (ESIpos): m/z=468
(M+H).sup.+
[0706] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.98 (s, 1H),
7.9 (br. s, 1H), 7.37 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H), 5.75
(br. s, 1H), 4.45 (s, 2H), 3.95 (s, 3H), 3.88 (s, 2H), 3.51-3.56
(m, 4H), 3.40 (q, 2H), 2.4-2.5 (m, 4H), 2.45 (s, 3H), 1.06 (t, 3H)
ppm.
Example 11
1-(4-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl-
)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piperazin-1-yl)ethanone
dihydrochloride
##STR00125##
[0708] A solution of Intermediate 17A (130 mg, 0.328 mmol) in
methanol (8 ml) was treated with N-acetylpiperazine (63 mg, 0.492
mmol), sodium cyanoborohydride (103 mg, 1.63 mmol) and acetic acid
(37 .mu.l, 0.655 mmol). The mixture was stirred at 60.degree. C.
for 3 h. It was then combined with the reaction mixture of a 30 mg
test run, evaporated and purified by preparative RP-HPLC (Reprosil
C18, gradient 10-95% acetonitrile/0.1% aq. formic acid). The
product thus obtained was lyophilized from 1,4-dioxane, then
dissolved in ethyl acetate and washed with sat. aq. sodium
hydrogencarbonate solution. The organic layer was dried over
magnesium sulfate, evaporated and lyophilized again from
1,4-dioxane. Re-purification by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.10% aq. formic acid), followed by
two-fold column chromatography on silica gel
(dichloromethane/methanol 98:2.fwdarw.4:1) and lyophilization from
a 4 M solution of hydrogen chloride in 1,4-dioxane afforded 49 mg
(18% of th.) of the title compound.
[0709] LC-MS (method 2): R.sub.t=0.82 min; MS (ESIpos): m/z=509
(M+H).sup.+
[0710] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=8.65-8.26 (br. s, 1H), 8.17 (s, 1H), 7.38 (s, 1H), 7.34 (s,
1H), 6.87 (s, 1H), 6.53-6.05 (br. s, 1H), 4.75 (br. s, 2H), 4.56
(br. s, 2H), 3.96 (s, 3H), 3.45 (q, 2H, overlap with water peak),
2.46 (s, 3H), 2.03 (s, 3H), 1.09 (t, 3H) ppm.
Example 12
[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmeth-
yl)pyrrolo[2,1-f]-[1,2,4]triazin-6-yl]methanol bis(formiate)
##STR00126##
[0712] A solution of Intermediate 23A (95 mg, 152 .mu.mol) in a 4 M
solution of hydrogen chloride in 1,4-dioxane (3.7 ml) was stirred
at rt for 2 h. After evaporation, the residue was purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) affording 44 mg (62% of th.) of
the title compound.
[0713] LC-MS (method 2): R.sub.t=0.63 min; MS (ESIpos): m/z=439
(M+H).sup.+
[0714] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.28 (br. s,
2H), 7.98 (s, 1H), 7.37 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.50
(s, 2H, overlap with water peak), 3.98 (s, 2H, overlap with water
peak), 3.95 (s, 3H, overlap with water peak), 2.95-2.84 (m, 4H),
2.61-2.55 (m, 4H, overlap with DMSO peak), 2.45 (s, 3H) ppm.
Example 13
4-{[4-Amino-6-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)p-
yrrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazin-2-one
##STR00127##
[0716] A solution of Intermediate 20A (1.34 g, 3.96 mmol) in acetic
acid (13.9 ml) was treated with 37% aq. formaldehyde solution (501
.mu.l, 6.6 mmol) and 2-oxopiperazine (670 mg, 6.6 mmol). The
mixture was stirred at 50.degree. C. for 2 h and then evaporated.
Purification by column chromatography on silica gel
(dichloromethane/methanol 98:2.fwdarw.90:10) afforded 942 mg (49%
of th.) of the title compound.
[0717] LC-MS (method 2): R.sub.t=0.67 min; MS (ESIpos): m/z=453
(M+H).sup.+
[0718] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.76 (br. s, 1H), 7.39 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H), 4.51
(s, 2H), 4.03 (br. s, 2H), 3.95 (s, 3H), 3.18-3.13 (m, 2H),
3.07-2.98 (m, 2H), 2.72-2.60 (m, 2H), 2.45 (s, 3H) ppm.
Example 14
7-{[(3S)-3-Amino-3-methylpyrrolidin-1-yl]methyl}-6-(methoxymethyl)-5-(7-me-
thoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00128##
[0720] A solution of Intermediate 21A (100 mg, 0.28 mmol) in acetic
acid (1 ml) was treated with 37% aq. formaldehyde solution (25
.mu.l, 0.33 mmol) and tert-butyl
[(3S)-3-methylpyrrolidin-3-yl]carbamate (Yoshida et al., Chem.
Pharm. Bull. 1996, 44 (7), 1376-1386; 67 mg, 0.33 mmol). The
mixture was stirred at rt for 3 h and then evaporated. The residue
was diluted with sat. aq. sodium hydrogencarbonate solution, and
solid potassium carbonate was added until no more gas generation
occurred. The aqueous phase was extracted three times with ethyl
acetate. The combined organic phases were dried over sodium sulfate
and evaporated. The residue was dissolved in a 4 M solution of
hydrogen chloride in 1,4-dioxane (2 ml) and stirred at rt for 2 h.
After evaporation, the residue was purified by two-fold preparative
RP-HPLC (first Reprosil C18, gradient 10-95% acetonitrile/0.1% aq.
formic acid; then Shield RP18, gradient 5-50% methanol+0.1% aq.
TFA/0.1% aq. TFA). The product thus obtained was lyophilized from a
4 M solution of hydrogen chloride in 1,4-dioxane affording 14 mg
(8% of th.) of the title compound.
[0721] LC-MS (method 2): R.sub.t=0.67 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0722] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=8.50-8.22 (br. s, 1H), 8.14 (s, 1H), 7.36 (s, 1H), 7.34 (s,
1H), 6.87 (s, 1H), 6.36-6.01 (br. s, 1H), 4.88 (br. s, 2H), 4.56
(br. s, 2H), 3.96 (s, 3H, overlap with water peak), 3.26 (s, 3H),
2.46 (s, 3H), 1.52 (s, 3H) ppm.
Example 15
7-{[(3S)-3-Amino-3-methylpyrrolidin-1-yl]methyl}-6-(ethoxymethyl)-5-(7-met-
hoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00129##
[0724] A solution of Intermediate 17A (100 mg, 271 .mu.mol) in
acetic acid (1 ml) was treated with 37% aq. formaldehyde solution
(24 .mu.l, 326 .mu.mol) and tert-butyl
[(3S)-3-methylpyrrolidin-3-yl]carbamate (Yoshida et al., Chem.
Pharm. Bull. 1996, 44 (7), 1376-1386; 65 mg, 326 .mu.mol). The
mixture was stirred at 60.degree. C. for 4 h. Then, further amounts
of 37% aq. formaldehyde solution (10 .mu.l, 136 .mu.mol) and
tert-butyl [(3S)-3-methylpyrrolidin-3-yl]carbamate (27 mg, 136
.mu.mol) were added, and stirring at 60.degree. C. was continued
overnight. After evaporation, the residue was partitioned between
ethyl acetate and sat. aq. sodium hydrogencarbonate solution. Solid
potassium carbonate was added until no more gas generation
occurred. The aqueous phase was extracted three times with ethyl
acetate. The combined organic phases were dried over sodium
sulfate, evaporated and purified by preparative RP-HPLC (Reprosil
CIS, gradient 10-95% acetonitrile/0.1% aq. formic acid). The
product thus obtained was dissolved in a 4 M solution of hydrogen
chloride in 1,4-dioxane (2 ml) and stirred at rt for 1 h. After
evaporation, the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic acid)
affording 24 mg (18% of th.) of the title compound.
[0725] LC-MS (method 4): R.sub.t=0.70 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0726] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.19
(s, 1H), 7.38 (s, 1H), 7.34 (s, 1H), 6.87 (s, 1H), 4.90 (br. s,
2H), 3.96 (s, 3H), 3.46 (q, 3H), 2.46 (s, 3H), 1.54 (s, 3H),
1.14-1.05 (m, 3H) ppm.
Example 16
1-(4-{[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-y-
l)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}piperazin-1-yl)ethanone
dihydrochloride
##STR00130##
[0728] A solution of Intermediate 21A (50 mg, 141 .mu.mol) in
acetic acid (1 ml) was treated with 37% aq. formaldehyde solution
(4.6 .mu.l, 169 .mu.mol) and N-acetylpiperazine (21.6 mg, 169
.mu.mol). The mixture was stirred at 75.degree. C. for 3 h. After
evaporation, the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic acid).
Lyophilization from a 4 M solution of hydrogen chloride in
1,4-dioxane afforded 34 mg (39% of th.) of the title compound.
[0729] LC-MS (method 5): R.sub.t=1.78 min; MS (ESIpos): m/z=495
(M+H).sup.+
[0730] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=8.70-8.35 (br. s, 1H), 8.18 (s, 1H), 7.37 (s, 1H), 7.34 (s,
1H), 6.86 (s, 1H), 6.61-6.19 (br. s, 1H), 4.74 (br. s, 2H), 4.54
(br. s, 2H), 3.96 (s, 3H), 3.25 (s, 3H), 2.46 (s, 3H), 2.03 (s, 3H)
ppm.
Example 17
6-(Methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-
-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine formiate
##STR00131##
[0732] A solution of Intermediate 23A (110 mg, 204 .mu.mol) in
dichloromethane (2.2 ml) was treated with thionyl chloride (29
.mu.l, 408 .mu.mol) and stirred at rt for 15 min. After
evaporation, the residue was dissolved in methanol (2.2 ml) and
treated with DIPEA (39 .mu.l, 224 .mu.mol). The mixture was stirred
at 70.degree. C. for 1 h and then evaporated. The residue was taken
up in a 4 M solution of hydrogen chloride in 1,4-dioxane (2.2 ml)
and stirred at rt for 2 h. Evaporation and purification by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 44.9 mg (46% of th.) of
the title compound.
[0733] LC-MS (method 2): R.sub.t=0.73 min; MS (ESIpos): m/z=453
(M+H).sup.+
[0734] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.28 (br. s,
1H), 8.00 (s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.41
(s, 2H), 4.00-3.90 (m, 5H), 3.00-2.90 (m, 4H), 2.65-2.56 (m, 4H),
2.45 (s, 3H) ppm.
Example 18
6-(Ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin--
1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00132##
[0736] A solution of Intermediate 23A (80 mg, 317 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (22 .mu.l,
297 .mu.mol) and stirred at rt for 15 min. After evaporation, the
residue was dissolved in ethanol (2 ml) and treated with DIPEA (28
.mu.l, 163 .mu.mol). The mixture was stirred at 70.degree. C. for 1
h and then evaporated. The residue was taken up in a 4 M solution
of hydrogen chloride in 1,4-dioxane (2 ml) and stirred at rt for 1
h. Evaporation and purification by preparative RP-HPLC (Reprosil
C18, gradient 10-95% acetonitrile/0.1% aq. formic acid) afforded 35
mg (50% of th.) of the title compound.
[0737] LC-MS (method 2): R.sub.t=0.82 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0738] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.09
(br. s, 1H), 7.36 (s, 1H), 7.32 (s, 1H), 6.86 (s, 1H), 4.50 (br. s,
2H), 3.96 (s, 3H), 3.42 (q, 2H, overlap with water peak), 2.45 (s,
3H), 1.05 (t, 3H) ppm.
Example 19
6-(Ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin--
1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine dihydrochloride
##STR00133##
[0740] A solution of Example 18 (50 mg, 107 .mu.mol) in a 4 M
solution of hydrogen chloride in 1,4-dioxane (1 ml) was stirred at
rt for 15 min. After evaporation 55 mg (93% of th.) of the title
compound were obtained.
[0741] LC-MS (method 4): R.sub.t=0.74 min; MS (ESIpos): m/z=467
(M+H).sup.+
[0742] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.18
(s, 1H), 7.37 (s, 1H), 7.34 (s, 1H), 6.87 (s, 1H), 4.76 (br. s,
2H), 4.60 (br. s, 2H), 3.96 (s, 3H), 3.45 (q, 2H), 2.46 (s, 3H),
1.09 (t, 3H) ppm.
Example 20
1-(4-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl-
)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}piperazin-1-yl)ethanone
##STR00134##
[0744] A solution of Example 18 (70 mg, 150 .mu.mol) in
dichloromethane (2 ml) and THF (0.8 ml) was treated with acetyl
chloride (21 .mu.l, 300 .mu.mol) and sodium carbonate (127 mg, 1.2
mmol). The mixture was stirred at rt overnight. After evaporation,
the residue was purified by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid) affording 27 mg
(31% of th.) of the title compound.
[0745] LC-MS (method 4): R.sub.t=0.77 min; MS (ESIpos): m/z=509
(M+H).sup.+
[0746] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): 5=7.98 (s, 1H), 7.37
(s, 1H), 7.31 (s, 1H), 6.84 (s, 1H), 4.45 (s, 2H), 3.95 (s, 3H),
3.92 (s, 2H), 3.45-3.36 (m, 6H), 2.48-2.38 (m, 7H, overlap with
DMSO peak), 1.97 (s, 3H), 1.06 (t, 3H) ppm.
Example 21
4-({4-Amino-6-[(2-hydroxyethoxy)methyl]-5-(7-methoxy-5-methyl-1-benzothiop-
hen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-7-yl}methyl)piperazin-2-one
formiate
##STR00135##
[0748] A solution of Example 13 (60 mg, 132 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (14 .mu.l,
198 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in ethylene glycol (500
.mu.l) and stirred at 100.degree. C. for 90 min. Purification by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 34 mg (47% of th.) of
the title compound.
[0749] LC-MS (method 2): R.sub.t=0.69 min; MS (ESIpos): m/z=497
(M+H).sup.+
[0750] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.14 (s, 1H),
8.00 (s, 1H), 7.74 (br. s, 1H), 7.42 (s, 1H), 7.31 (s, 1H), 6.84
(s, 1H), 4.49 (s, 2H), 3.99 (s, 2H), 3.96 (s, 3H), 3.50-3.43 (m,
2H), 3.42-3.36 (m, 2H, overlap with water peak), 3.15-3.07 (m, 2H),
3.03 (s, 2H), 2.69-2.61 (m, 2H), 2.45 (s, 3H) ppm.
Example 22
2-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f]-[1,2,4]triazin-6-yl]methoxy}ethanol
dihydrochloride
##STR00136##
[0752] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (4 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 20 min and then
evaporated. The residue was dissolved in ethylene glycol/THF (2:1,
1.5 ml) and stirred at 100.degree. C. for 2 h. After evaporation,
the residue was purified by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid), followed by
column chromatography on silica gel (dichloromethane/methanol+0.1%
aq. ammonia 98:2.fwdarw.90:10). Lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 67 mg (64% of th.) of
the title compound.
[0753] LC-MS (method 2): R.sub.t=0.66 min; MS (ESIpos): m/z=483
(M+H).sup.+
[0754] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.16 (s, 1H),
7.49 (s, 1H), 7.43 (s, 1H), 6.98 (s, 1H), 4.86 (s, 2H), 4.69 (s,
2H), 4.03 (s, 3H), 3.81-3.49 (m, 12H), 2.50 (s, 3H) ppm.
Example 23
6-(Butoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin--
1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine formiate
##STR00137##
[0756] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in 1-butanol (2 ml) and
heated to 70.degree. C. for 1 h. After evaporation, the residue was
taken up in a 4 M solution of hydrogen chloride in 1,4-dioxane (2
ml) and stirred at rt for 2 h. Evaporation and purification by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 22 mg (24% of th.) of
the title compound.
[0757] LC-MS (method 2): R.sub.t=0.82 min; MS (ESIpos): m/z=495
(M+H).sup.+
[0758] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.29 (s, 1H),
7.99 (s, 1H), 7.35 (s, 1H), 7.29 (s, 1H), 6.84 (s, 1H), 4.43 (s,
2H), 3.95 (s, 3H), 3.92 (s, 2H), 3.35 (t, 2H), 2.90-2.79 (m, 4H),
2.45 (s, 3H), 1.47-1.37 (m, 2H), 1.31-1.18 (m, 2H), 0.80 (t, 3H)
ppm.
Example 24
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmethyl)-6-(pr-
opoxymethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine bis(formiate)
##STR00138##
[0760] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in 1-propanol (2 ml), treated
with DIPEA (48 .mu.l, 278 .mu.mol) and stirred at rt for 1 h. After
evaporation, the residue was taken up in a 4 M solution of hydrogen
chloride in 1,4-dioxane (2 ml) and stirred at rt for 2 h.
Evaporation and purification by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid) afforded 15 mg
(16% of th.) of the title compound.
[0761] LC-MS (method 2): R.sub.t=0.78 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0762] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.26 (br. s,
2H), 7.98 (s, 1H), 7.36 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 4.44
(s, 2H), 3.95 (s, 3H), 3.92 (s, 2H), 3.32 (t, 2H), 2.87-2.79 (m,
4H), 1.52-1.39 (m, 2H), 0.81 (t, 3H) ppm.
Example 25
6-[(Cyclopropylmethoxy)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-
-7-(piperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
bis(formiate)
##STR00139##
[0764] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in cyclopropylmethanol (2
ml), treated with DIPEA (48 .mu.l, 278 .mu.mol) and stirred at
70.degree. C. for 2 h. After evaporation, the residue was taken up
in a 4 M solution of hydrogen chloride in 1,4-dioxane (2 ml) and
stirred at rt for 2 h. Evaporation and purification by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) afforded 29 mg (30% of th.) of the title compound.
[0765] LC-MS (method 2): R.sub.t=0.78 min; MS (ESIpos): m/z=493
(M+H).sup.+
[0766] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.28 (br. s,
2H), 7.99 (s, 1H), 7.37 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 4.45
(s, 2H), 3.95 (s, 3H), 3.93 (s, 2H), 3.21 (d, 2H), 2.95-2.83 (m,
4H), 2.61-2.56 (m, 4H), 2.45 (s, 3H), 1.01-0.91 (m, 1H), 0.46-0.37
(m, 2H), 0.15-0.08 (m, 2H) ppm.
Example 26
6-[(Cyclobutyloxy)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(p-
iperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
##STR00140##
[0768] A solution of Intermediate 23A (85 mg, 157 .mu.mol) in
dichloromethane (1.7 ml) was treated with thionyl chloride (23
.mu.l, 315 .mu.mol). The mixture was stirred at rt for 15 min and
then evaporated. The residue was dissolved in cyclobutanol (1.7
ml), treated with DIPEA (41 .mu.l, 236 .mu.mol) and stirred at
70.degree. C. for 2 h. After evaporation, the residue was taken up
in a 4 M solution of hydrogen chloride in 1,4-dioxane (1.7 ml) and
stirred at rt for 2 h. Evaporation and purification by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) afforded 23 mg (28% of th.) of the title compound.
[0769] LC-MS (method 2): R.sub.t=0.82 min; MS (ESIpos): m/z=493
(M+H).sup.+
[0770] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.01 (s, 1H),
7.37 (s, 1H), 7.32 (s, 1H), 6.85 (s, 1H), 4.37 (s, 2H), 4.06-3.86
(m, 6H), 3.14-2.97 (m, 4H), 2.80-2.62 (m, 4H), 2.45 (s, 3H),
2.10-1.96 (m, 2H), 1.83-1.69 (m, 2H), 1.65-1.52 (m, 1H), 1.49-1.33
(m, 1H) ppm.
Example 27
6-(Isopropoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(pipera-
zin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine formiate
##STR00141##
[0772] A solution of Intermediate 23A (65 mg, 120 .mu.mol) in
dichloromethane (1.3 ml) was treated with thionyl chloride (17
.mu.l, 241 .mu.mol). The mixture was stirred at rt for 15 min and
then evaporated. The residue was dissolved in 2-propanol (1.3 ml),
treated with DIPEA (23 .mu.l, 132 .mu.mol) and stirred at
70.degree. C. for 1 h. Further DIPEA (23 .mu.l, 132 .mu.mol) was
added, and the mixture was stirred again at 70.degree. C. for 1 h.
Then, another portion of DIPEA (63 .mu.l, 362 .mu.mol) was added,
and stirring was continued at 90.degree. C. for 3 h. After
evaporation, the residue was taken up in a 4 M solution of hydrogen
chloride in 1,4-dioxane (1.3 ml) and stirred at rt for 2 h.
Evaporation and purification by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid) afforded 23 mg
(34% of th.) of the title compound.
[0773] LC-MS (method 4): R.sub.t=0.74 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0774] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.30 (s, 1H),
7.98 (s, 1H), 7.37 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 4.43 (s,
2H), 3.95 (s, 3H), 3.90 (s, 2H), 3.57 (m, 1H, overlap with water
peak), 2.88-2.78 (m, 4H), 2.45 (s, 3H), 1.04 (d, 6H) ppm.
Example 28
6-[(2-Methoxyethoxy)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7--
(piperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
formiate
##STR00142##
[0776] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in 2-methoxyethanol (2 ml),
treated with DIPEA (35 .mu.l, 204 .mu.mol) and stirred at
70.degree. C. for 1 h. After evaporation, the residue was taken up
in a 4 M solution of hydrogen chloride in 1,4-dioxane (2 ml) and
stirred at rt for 2 h. Evaporation and purification by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) afforded 50 mg (50% of th.) of the title compound.
[0777] LC-MS (method 2): R.sub.t=0.75 min; MS (ESIpos): m/z=497
(M+H).sup.+
[0778] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.26 (br. s,
1H), 7.99 (s, 1H), 7.39 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 4.48
(s, 2H), 3.95 (s, 3H), 3.94 (s, 2H), 3.52-3.46 (m, 2H, overlap with
water peak), 3.44-3.37 (m, 2H, overlap with water peak), 3.20 (s,
3H), 2.94-2.84 (m, 4H), 2.61-2.54 (m, 4H, overlap with DMSO peak),
2.45 (s, 3H) ppm.
Example 29
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmethyl)-6-[(2-
,2,2-trifluoroethoxy)methyl]pyrrolo[2,1-f][1,2,4]triazin-4-amine
formiate
##STR00143##
[0780] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in 2,2,2-trifluoroethanol (2
ml), treated with DIPEA (35 .mu.l, 204 .mu.mol) and stirred at
70.degree. C. for 1 h. Further DIPEA (35 .mu.l, 204 .mu.mol) was
added, and stirring was continued at rt for 1 h. After evaporation,
the residue was taken up in a 4 M solution of hydrogen chloride in
1,4-dioxane (2 ml) and stirred at rt for 2 h. Evaporation and
purification by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 23 mg (23% of th.) of
the title compound.
[0781] LC-MS (method 2): R.sub.t=0.79 min; MS (ESIpos): m/z=521
(M+H).sup.+
[0782] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.28 (br. s,
1H), 8.00 (s, 1H), 7.37 (s, 1H), 7.30 (s, 1H), 6.85 (s, 1H), 4.66
(s, 2H), 4.08 (q, 2H), 3.95 (s, 3H), 3.92 (s, 2H), 2.85-2.75 (m,
4H), 2.45 (s, 3H) ppm.
Example 30
6-[(2-Aminoethoxy)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(p-
iperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00144##
[0784] A solution of Intermediate 23A (150 mg, 278 .mu.mol) in
dichloromethane (5 ml) was treated with thionyl chloride (40 .mu.l,
556 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in THF (0.5 ml) and treated
with tert-butyl (2-hydroxyethyl)carbamate (1 ml) and DIPEA (242
.mu.l, 1.39 mmol). The mixture was stirred at 100.degree. C.
overnight. After evaporation, the residue was taken up in
1,4-dioxane (10 ml), treated with a 4 M solution of hydrogen
chloride in 1,4-dioxane (10 ml) and stirred at rt for 1 h. The
volatiles were removed under reduced pressure, and the residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid). Further purification by
preparative RP-HPLC (Shield RP18, 25% acetonitrile/75% 0.01% aq.
TFA) followed by lyophilization from a 4 M solution of hydrogen
chloride in 1,4-dioxane afforded 10 mg (6% of th.) of the title
compound.
[0785] LC-MS (method 2): R.sub.t=0.58 min; MS (ESIpos): m/z=482
(M+H).sup.+
[0786] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.12 (s, 1H), 7.50
(s, 1H), 7.44 (s, 1H), 6.99 (s, 1H), 4.61 (s, 2H), 4.04 (s, 3H),
3.88-3.80 (m, 1H), 3.70-3.60 (m, 3H), 3.52-3.46 (m, 4H), 3.40-3.34
(m, 4H), 3.07 (t, 2H), 2.51 (s, 3H) ppm.
Example 31
Methyl
{[4-amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin--
1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methoxy}acetate
##STR00145##
[0788] A solution of Intermediate 23A (50 mg, 92 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (13 .mu.l,
186 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in methylglycolate (1 ml),
treated with DIPEA (80 .mu.l, 464 .mu.mol) and stirred at
70.degree. C. for 2 h. After evaporation, the residue was taken up
in dichloromethane (1.6 ml), treated with trifluoroacetic acid (400
.mu.l, 5.19 mmol) and stirred at rt for 1 h. Then, the mixture was
evaporated, and the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. TFA).
Fractions containing the title compound were neutralized with sat.
aq. sodium hydrogencarbonate solution and evaporated. The residue
was taken up in water, and the mixture was extracted three times
with dichloromethane. The combined organic phases were dried over
sodium sulfate and evaporated affording 21 mg (43% of th.) of the
title compound.
[0789] LC-MS (method 2): R.sub.t=0.72 min; MS (ESIpos): m/z=511
(M+H).sup.+
[0790] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.39 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.59 (s, 2H), 4.11 (s,
2H), 3.95 (s, 3H), 3.90 (s, 2H), 3.57 (s, 3H), 2.81-2.69 (m, 4H),
2.48-2.39 (m, 7H) ppm.
Example 32
{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmet-
hyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methoxy}acetic acid
##STR00146##
[0792] A solution of Intermediate 24A (200 mg, 327 .mu.mol) in THF
(14 ml) was treated with 2.5 M aq. lithium hydroxide solution (16
ml) and stirred at 80.degree. C. for 2 h. The mixture was then
combined with the reaction mixture of a 27 mg test run. The aqueous
phase was extracted twice with THF, and the combined organic phases
were evaporated. The residue was dissolved in a 4 M solution of
hydrogen chloride in 1,4-dioxane (2 ml) and stirred at rt for 3 h.
After evaporation, the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. TFA).
Lyophilization from a 4 M solution of hydrogen chloride in
1,4-dioxane and re-purification by preparative RP-HPLC (XBridge
C18, gradient 5-95% acetonitrile/0.1% aq. ammonium hydroxide
solution) afforded 13 mg (7% of th.) of the title compound.
[0793] LC-MS (method 2): R.sub.t=0.69 min; MS (ESIpos): m/z=497
(M+H).sup.+
[0794] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.38 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H), 4.54 (s, 2H), 4.08 (s,
2H), 3.95 (s, 3H), 3.79 (s, 2H), 2.99-2.89 (m, 4H), 2.73-2.63 (m,
4H), 2.44 (s, 3H) ppm.
Example 33
2-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methoxy}acetamide
##STR00147##
[0796] Intermediate 24A (200 mg, 327 .mu.mol) was treated with a 7
M solution of ammonia in methanol (10 ml) and stirred at rt
overnight. The mixture was then combined with the reaction mixture
of a 20 mg test run, and the solvents were evaporated. The residue
was treated with a 4 M solution of hydrogen chloride in 1,4-dioxane
(2 ml) and stirred at rt for 2 h. Evaporation and subsequent
purifications by preparative RP-HPLC (first Reprosil C18, gradient
10-95% acetonitrile/0.1% aq. TFA; then XBridge C18, gradient 5-95%
acetonitrile/0.1% aq. ammonium hydoxide solution) afforded 5.5 mg
(3% of th.) of the title compound.
[0797] LC-MS (method 2): R.sub.t=0.69 min; MS (ESIpos): m/z=496
(M+H).sup.+
[0798] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.14-7.59 (br.
s, 1H), 7.99 (s, 1H), 7.39 (s, 1H), 7.35-7.22 (m, 3H), 6.85 (s,
1H), 4.54 (s, 2H), 3.95 (s, 3H), 3.90 (s, 2H), 3.79 (s, 2H),
2.73-2.66 (m, 4H), 2.45 (s, 3H), 2.43-2.35 (m, 4H) ppm.
Example 34
2-({7-[(4-Acetylpiperazin-1-yl)methyl]-4-amino-5-(7-methoxy-5-methyl-1-ben-
zothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl}methoxy)acetamide
##STR00148##
[0800] A solution of Example 33 (105 mg, 68% purity, 144 .mu.mol)
in THF/dichloromethane (1:2.5, 3.9 ml) was treated with sodium
carbonate (179 mg, 1.6 mmol) and stirred at rt for 30 min. Acetyl
chloride (30 .mu.l, 424 .mu.mol) was added, and the resulting
mixture was stirred at rt for 30 min, then quenched with methanol
(2 ml) and evaporated. Purification by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic acid)
afforded 30 mg (85% purity, 34% of th.) of the title compound.
[0801] LC-MS (method 5): R.sub.t=1.67 min; MS (ESIpos): m/z=538
(M+H).sup.+
[0802] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.40 (s, 1H), 7.33-7.19 (m, 3H), 6.85 (s, 1H), 4.55 (s, 2H), 3.96
(s, 5H), 3.80 (s, 2H), 3.43-3.36 (m, 4H), 2.47-2.38 (m, 7H), 1.97
(s, 3H) ppm.
Example 35
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-6-(phenoxymethyl)-7-(piperazin-
-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine bis(formiate)
##STR00149##
[0804] A solution of Intermediate 23A (100 mg, 185 .mu.mol) in
dichloromethane (2 ml) was treated with thionyl chloride (27 .mu.l,
371 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in THF (2 ml), treated with
phenol (174 mg, 1.85 mmol) and DIPEA (48 .mu.l, 278 .mu.mol) and
stirred at 70.degree. C. for 2 h. Further amounts of phenol (174
mg, 1.85 mmol) and DIPEA (64 .mu.l, 371 .mu.mol) were added, and
stirring was continued at 70.degree. C. overnight. After
evaporation, the residue was taken up in a 4 M solution of hydrogen
chloride in 1,4-dioxane (2 ml) and stirred at rt for 2 h.
Evaporation and purification by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. formic acid) afforded 8 mg
(8% of th.) of the title compound.
[0805] LC-MS (method 2): R.sub.t=0.84 min; MS (ESIpos): m/z=515
(M+H).sup.+
[0806] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.26 (s, 2H),
8.02 (s, 1H), 7.36 (s, 1H), 7.30-7.22 (m, 3H), 6.99-6.89 (m, 3H),
6.81 (m, 1H), 5.04 (s, 2H), 3.93 (s, 5H), 2.78-2.71 (m, 4H), 2.42
(s, 3H) ppm.
Example 36
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-6-[(methylamino)methyl]-7-(pip-
erazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00150##
[0808] Intermediate 25A (150 mg, 279 .mu.mol) in THF (3 ml) was
treated with acetic acid (32 .mu.l, 559 .mu.mol), a 2 M solution of
methylamine in THF (698 .mu.l, 1.39 mmol) and sodium
triacetoxyborohydride (296 mg, 1.39 mmol). The mixture was stirred
at 60.degree. C. for 2 h and then evaporated. The residue was
dissolved in a 4 M solution of hydrogen chloride in 1,4-dioxane
(1.87 ml) and stirred at rt for 2 h. After evaporation, the residue
was purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). Lyophilization from a 4 M solution of
hydrogen chloride in 1,4-dioxane afforded 79 mg (49% of th.) of the
title compound.
[0809] LC-MS (method 2): R.sub.t=0.56 min; MS (ESIpos): m/z=452
(M+H).sup.+
[0810] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.17 (s, 1H), 7.57
(s, 1H), 7.46 (s, 1H), 7.01 (s, 1H), 4.52 (s, 2H), 4.46 (s, 2H),
4.04 (s, 3H), 3.48-3.39 (m, 4H), 3.25-3.15 (m, 4H), 2.59 (s, 3H),
2.51 (s, 3H) ppm.
Example 37
6-[(Dimethylamino)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(p-
iperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00151##
[0812] A solution of Intermediate 25A (150 mg, 279 .mu.mol) in THF
(3 ml) was treated with acetic acid (32 .mu.l, 559 .mu.mol), a 2 M
solution of dimethylamine in THF (698 .mu.l, 1.39 mmol) and sodium
triacetoxyborohydride (296 mg, 1.39 mmol). The mixture was stirred
at 60.degree. C. for 2 h and then evaporated. The residue was
dissolved in a 4 M solution of hydrogen chloride in 1,4-dioxane
(1.88 ml) and stirred at rt for 2 h. After evaporation, the residue
was purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). Lyophilization from a 4 M solution of
hydrogen chloride in 1,4-dioxane afforded 83 mg (50% of th.) of the
title compound.
[0813] LC-MS (method 4): R.sub.t=0.50 min; MS (ESIpos): m/z=466
(M+H).sup.+
[0814] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.17 (s, 1H), 7.57
(s, 1H), 7.45 (s, 1H), 7.01 (s, 1H), 4.57 (s, 2H), 4.50 (s, 2H),
4.04 (s, 3H), 3.47-3.36 (m, 4H), 3.21-3.12 (m, 4H), 2.80 (s, 6H),
2.51 (s, 3H) ppm.
Example 38
6-[(Ethylamino)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(pipe-
razin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00152##
[0816] A solution of Intermediate 25A (60 mg, 111.8 .mu.mol) in THF
(1 ml) was treated with a 2 M solution of ethylamine in THF (83
.mu.l, 167 .mu.mol), sodium triacetoxyborohydride (118 mg, 559
.mu.mol) and acetic acid (83 .mu.l, 167 .mu.mol). The mixture was
stirred at 60.degree. C. for 90 min. Further amounts of 2 M
ethylamine solution (83 .mu.l, 167 .mu.mol) and sodium
triacetoxyborohydride (71 mg, 335 .mu.mol) were added, and stirring
at 60.degree. C. was continued for another 2 h. After evaporation,
the residue was dissolved in a 4 M solution of hydrogen chloride in
1,4-dioxane (3 ml) and stirred at rt overnight. The mixture was
evaporated, and the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic acid)
and lyophilized from a 4 M solution of hydrogen chloride in
1,4-dioxane. Re-purification by preparative RP-HPLC (Sunfire C18,
20% acetonitrile/80% 0.02% aq. TFA) and re-lyophilization from a 4
M solution of hydrogen chloride in 1,4-dioxane afforded 19 mg (29%
of th.) of the title compound.
[0817] LC-MS (method 2): R.sub.t=0.56 min; MS (ESIpos): m/z=466
(M+H).sup.+
[0818] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.14 (s, 1H), 7.56
(s, 1H), 7.45 (s, 1H), 7.01 (s, 1H), 4.45 (s, 4H), 4.04 (s, 3H),
3.45-3.32 (m, 4H), 3.18-3.05 (m, 4H), 2.99 (q, 2H), 2.51 (s, 3H),
1.10 (t, 3H) ppm.
Example 39
2-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-yl-
methyl)pyrrolo[2,1-f]-[1,2,4]triazin-6-yl]methyl}amino)ethanol
trihydrochloride
##STR00153##
[0820] A solution of Intermediate 25A (150 mg, 279 .mu.mol) in THF
(3 ml) was treated with 2-aminoethanol (84 .mu.l, 1.39 mmol),
sodium triacetoxyborohydride (296 mg, 1.39 mmol) and acetic acid
(32 .mu.l, 559 mol). The mixture was stirred at 60.degree. C. for 2
h and then evaporated. The residue was dissolved in a 4 M solution
of hydrogen chloride in 1,4-dioxane (1.87 ml) and stirred at rt for
2 h. After evaporation, the residue was purified by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. TFA).
Lyophilization from a 4 M solution of hydrogen chloride in
1,4-dioxane afforded 140 mg (80% of th.) of the title compound.
[0821] LC-MS (method 2): R.sub.t=0.54 min; MS (ESIpos): m/z=482
(M+H).sup.+
[0822] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.15 (s, 1H), 7.56
(s, 1H), 7.45 (s, 1H), 7.00 (s, 1H), 4.47 (s, 2H), 4.37 (s, 2H),
4.04 (s, 3H), 3.79-3.71 (m, 2H), 3.42-3.34 (m, 4H), 3.19-3.12 (m,
2H), 3.06-2.97 (m, 4H), 2.51 (s, 3H) ppm.
General Procedure for Reductive Amination Reactions with
Intermediate 25A (GP1):
[0823] A 0.1 M solution of Intermediate 25A in THF was treated with
5 eq. of the respective amine component, 5 eq. of sodium
triacetoxyborohydride and 2 eq. of acetic acid. The resulting
mixture was stirred at 60.degree. C. for 2 h and then evaporated. A
0.15 M solution of the residue thus obtained in a 4 M solution of
hydrogen chloride in 1,4-dioxane was stirred at rt for 1-2 h. After
evaporation, the residue was purified as described below.
Example 40
rac-1-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-
-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}piperidin-3-ol
trihydrochloride
##STR00154##
[0825] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with 3-hydroxypiperidine (141 mg, 1.39 mmol). Purification
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 178 mg (quant.) of the
title compound.
[0826] LC-MS (method 2): R.sub.t=0.54 min; MS (ESIpos): m/z=522
(M+H).sup.+
[0827] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.16 (s, 1H), 7.55
(s, 1H), 7.45 (s, 1H), 7.01 (s, 1H), 4.66-4.16 (m, 5H), 4.04 (s,
3H), 3.55-2.71 (m, 11H), 2.51 (s, 3H), 1.96-1.42 (m, 4H), 1.64-1.50
(m, 1H) ppm.
Example 41
1-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}piperidin-4-ol
trihydrochloride
##STR00155##
[0829] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with 4-hydroxypiperidine (141 mg, 1.39 mmol). Purification
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 162 mg (91% of th.) of
the title compound.
[0830] LC-MS (method 4): R.sub.t=0.48 min; MS (ESIpos): m/z=522
(M+H).sup.+
[0831] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.16 (s, 1H), 7.58
(br. s, 1H), 7.46 (s, 1H), 7.01 (s, 1H), 4.67-4.48 (m, 4H), 4.04
(s, 4H), 3.59-2.78 (m, 12H), 2.51 (s, 3H), 2.14-1.38 (m, 4H)
ppm.
Example 42
rac-1-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-
-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}pyrrolidin-3-ol
trihydrochloride
##STR00156##
[0833] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with 3-hydroxypyrrolidine (113 .mu.l, 1.39 mmol).
Purification by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 148 mg (85% of th.) of
the title compound.
[0834] LC-MS (method 4): R.sub.t=0.46 min; MS (ESIpos): m/z=508
(M+H).sup.+
[0835] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.18 (s, 1H), 7.58
(s, 1H), 7.46 (s, 1H), 7.01 (s, 1H), 4.71-4.30 (m, 5H), 4.04 (s,
3H), 3.70-2.95 (m, 12H), 2.51 (s, 3H), 2.32-2.14 (m, 1H), 2.03-1.87
(m, 1H) ppm.
Example 43
6-[(Diethylamino)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(pi-
perazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00157##
[0837] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with diethylamine (144 .mu.l, 1.39 mmol). Purification by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 127 mg (69% of th.) of
the title compound.
[0838] LC-MS (method 2): R.sub.t=0.55 min; MS (ESIpos): m/z=494
(M+H).sup.+
[0839] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.19 (s, 1H), 7.61
(s, 1H), 7.47 (s, 1H), 7.01 (s, 1H), 4.59 (s, 2H), 4.51 (s, 2H),
4.04 (s, 3H), 3.47-3.35 (m, 4H), 3.29-3.00 (m, 8H), 2.51 (s, 3H),
1.07 (t, 6H) ppm.
Example 44
6-[(Cyclobutylamino)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7--
(piperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00158##
[0841] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with cyclobutylamine (119 .mu.l, 1.39 mmol). Purification
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 127 mg (72% of th.) of
the title compound.
[0842] LC-MS (method 2): R.sub.t=0.58 min; MS (ESIpos): m/z=492
(M+H).sup.+
[0843] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.16 (s, 1H), 7.59
(s, 1H), 7.47 (s, 1H), 7.00 (s, 1H), 4.40 (s, 2H), 4.37 (s, 2H),
4.04 (s, 3H), 3.69-3.58 (m, 1H), 3.44-3.33 (m, 4H), 3.21-3.12 (m,
4H), 2.51 (s, 3H), 2.02-1.87 (m, 4H), 1.81-1.62 (m, 2H) ppm.
Example 45
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmethyl)-6-(py-
rrolidin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00159##
[0845] According to GP1, Intermediate 25A (150 mg, 279 .mu.mol) was
reacted with pyrrolidine (116 .mu.l, 1.39 mmol). Purification by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 112 mg (64% of th.) of
the title compound.
[0846] LC-MS (method 2): R.sub.t=0.54 min; MS (ESIpos): m/z=492
(M+H).sup.+
[0847] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.19 (s, 1H), 7.60
(s, 1H), 7.46 (s, 1H), 7.01 (s, 1H), 4.70-4.61 (m, 4H), 4.04 (s,
3H), 3.55-3.40 (m, 6H), 3.35-3.26 (m, 4H), 3.02-2.87 (m, 2H), 2.51
(s, 3H), 1.97-1.83 (m, 4H) ppm.
Example 46
6-[(Cyclopropylamino)methyl]-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-
-(piperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00160##
[0849] According to GP1, Intermediate 25A (200 mg, 372 .mu.mol) was
reacted with cyclopropylamine (129 .mu.l, 1.86 mmol). Purification
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 140 mg (62% of th.) of
the title compound.
[0850] LC-MS (method 5): R.sub.t=1.42 min; MS (ESIpos): m/z=478
(M+H).sup.+
[0851] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.16 (s, 1H), 7.56
(s, 1H), 7.46 (s, 1H), 7.01 (s, 1H), 4.57 (s, 2H), 4.47 (s, 2H),
4.04 (s, 3H), 3.46-3.34 (m, 4H), 3.19-3.07 (m, 4H), 2.61-2.54 (m,
1H), 2.51 (s, 3H), 0.75-0.64 (m, 4H) ppm.
Example 47
6-{[(Cyclopropylmethyl)amino]methyl}-5-(7-methoxy-5-methyl-1-benzothiophen-
-2-yl)-7-(piperazin-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00161##
[0853] According to GP1, Intermediate 25A (200 mg, 372 .mu.mol) was
reacted with cyclopropylmethylamine (161 .mu.l, 1.86 mmol).
Purification by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) and lyophilization from a 4 M solution
of hydrogen chloride in 1,4-dioxane afforded 163 mg (69% of th.) of
the title compound.
[0854] LC-MS (method 2): R.sub.t=0.60 min; MS (ESIpos): m/z=492
(M+H).sup.+
[0855] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.14 (s, 1H), 7.57
(s, 1H), 7.45 (s, 1H), 7.00 (s, 1H), 4.50 (s, 2H), 4.44 (s, 2H),
4.04 (s, 3H), 3.42-3.34 (m, 4H), 3.16-3.07 (m, 4H), 2.86 (d, 2H),
2.50 (s, 3H), 0.89-0.76 (m, 1H), 0.55-0.45 (m, 2H), 0.23-0.14 (m,
2H) ppm.
Example 48
N-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f]-[1,2,4]triazin-6-yl]methyl}glycine
trihydrochloride
##STR00162##
[0857] A solution of Intermediate 25A (161 mg, 300 .mu.mol) in THF
(3.2 ml) was treated with 2-aminoacetic acid (112 mg, 1.5 mmol),
sodium triacetoxyborohydride (317 mg, 1.5 mmol) and acetic acid (34
.mu.l, 600 .mu.mol). The resulting mixture was stirred at
60.degree. C. for 2 h and then evaporated. The residue was purified
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). The product thus obtained was dissolved
in a 4 M solution of hydrogen chloride in 1,4-dioxane (2 ml) and
stirred at rt for 1 h. After evaporation, the residue was purified
by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). Subsequent lyophilization from a 4 M
solution of hydrogen chloride in 1,4-dioxane afforded 18 mg (9% of
th.) of the title compound.
[0858] LC-MS (method 2): R.sub.t=0.63 min; MS (ESIpos): m/z=496
(M+H).sup.+
[0859] .sup.1H-NMR (400 MHz, D.sub.2O): .delta.=8.14 (s, 1H), 7.54
(s, 1H), 7.45 (s, 1H), 6.99 (s, 1H), 4.42 (s, 2H), 4.36 (s, 2H),
4.03 (s, 3H), 3.64 (s, 2H), 3.42-3.33 (m, 4H), 3.07-2.96 (m, 4H),
2.50 (s, 3H) ppm.
Example 49
4-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}piperazin-2-one
trihydrochloride
##STR00163##
[0861] A solution of Intermediate 27A (220 mg, 354 .mu.mol) in a 4
M solution of hydrogen chloride in 1,4-dioxane (2 ml) was stirred
at rt for 2 h. Then, the mixture was evaporated leaving 235 mg of
the crude product which was used in the next step without further
purification.
[0862] LC-MS (method 4): R.sub.t=0.60 min; MS (ESIpos): m/z=521
(M+H).sup.+.
Example 50
[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin-4-ylmeth-
yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methanol
##STR00164##
[0864] A solution of Intermediate 20A (500 mg, purity 87%, 1.28
mmol) and 4-methylenemorpholin-4-ium chloride (347 mg, 2.56 mmol)
in DMF (28 ml) was stirred at 70.degree. C. for 1.5 h. The mixture
was diluted with ethyl acetate and washed with sat. aq. sodium
hydrogencarbonate solution and sat. aq. sodium chloride solution.
The organic layer was dried with magnesium sulfate and evaporated
under reduced pressure yielding 710 mg (purity 78%, 99% of th.) of
the title compound.
[0865] LC-MS (method 2): R.sub.t=0.65 min; MS (ESIpos): m/z=440
(M+H).sup.+.
Example 51
(3S)-3-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin-
-4-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}amino)pyrrolidin-2-on-
e
##STR00165##
[0867] A solution of Intermediate 33A (65 mg, 149 .mu.mol) in
methanol (2 ml) was treated with (3S)-3-aminopyrrolidin-2-one (45
mg, 446 .mu.mol), sodium cyanoborohydride (47 mg, 743 .mu.mol) and
acetic acid (26 .mu.l, 446 .mu.mol). After stirring at 60.degree.
C. for 16 h, the resulting mixture was separated by preparative
RP-HPLC (Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA).
The product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 49 mg (63% of th.) of the
title compound.
[0868] LC-MS (method 2): R.sub.t=0.71 min; MS (ESIpos): m/z=522
(M+H).sup.+
[0869] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96 (s, 1H),
7.66 (s, 1H), 7.5-8.1 (br. s, 1H), 7.44 (s, 1H), 7.31 (s, 1H), 6.84
(s, 1H), 5.4-6.0 (br. s, 1H), 3.95 (s, 3H), 3.91 (s, 2H), 3.75 (d,
2H), 3.50-3.66 (m, 4H), 3.06-3.19 (m, 3H), 2.94-3.05 (m, 1H), 2.61
(t, 1H), 2.45 (t, 3H), 2.38-2.44 (m, 4H), 1.89-1.99 (m, 1H),
1.49-1.60 (m, 1H) ppm.
Example 52
4-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin-4-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}piperazin-2-one
##STR00166##
[0871] A solution of Intermediate 33A (65 mg, 149 .mu.mol) in
methanol (2 ml) was treated with 2-oxopiperazine (45 mg, 446
.mu.mol), sodium cyanoborohydride (47 mg, 743 .mu.mol) and acetic
acid (26 .mu.l, 446 .mu.mol). After stirring at 60.degree. C. for
16 h, the resulting mixture was separated by preparative RP-HPLC
(Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA). The
product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 41 mg (53% of th.) of the
title compound.
[0872] LC-MS (method 2): R.sub.t=0.70 min; MS (ESIpos): m/z=522
(M+H).sup.+
[0873] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.97 (s, 1H),
7.67 (br. s, 1H), 7.55-8.05 (br. s, 1H), 7.38 (s, 1H), 7.30 (s,
1H), 6.84 (s, 1H), 5.29-5.88 (br. s, 1H), 3.95 (s, 3H), 3.92 (s,
2H), 3.58 (s, 2H), 3.51-3.56 (m, 4H), 3.00-3.06 (m, 2H), 2.85 (s,
2H), 2.45 (s, 3H), 2.43-2.48 (m, 5H) ppm.
Example 53
rac-1-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin--
4-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}amino)propan-2-ol
##STR00167##
[0875] A solution of Intermediate 33A (64 mg, 146 .mu.mol) in
methanol (2 ml) was treated with rac-1-aminopropan-2-ol (33 mg, 439
.mu.mol), sodium cyanoborohydride (46 mg, 731 .mu.mol) and acetic
acid (25 .mu.l, 439 .mu.mol). After stirring at 60.degree. C. for
16 h, the resulting mixture was separated by preparative RP-HPLC
(Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA). The
product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 42 mg (57% of th.) of the
title compound.
[0876] LC-MS (method 5): R.sub.t=1.63 min; MS (ESIpos): m/z=497
(M+H).sup.+
[0877] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96 (s, 1H),
7.47-7.92 (br. s, 1H), 7.39 (s, 1H), 7.30 (s, 1H), 6.84 (s, 1H),
5.36-5.92 (br. s, 1H), 4.36 (d, 1H), 3.95 (s, 3H), 3.90 (s, 2H),
3.64-3.75 (m, 2H), 3.49-3.61 (m, 5H), 2.45 (s, 3H), 2.39-2.47 (m,
4H), 2.31-2.38 (m, 2H), 0.97 (d, 3H) ppm.
Example 54
1-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(morpholin-4-yl-
methyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}amino)-2-methylpropan-2-ol
##STR00168##
[0879] A solution of Intermediate 33A (80 mg, 183 .mu.mol) in
methanol (2 ml) was treated with 1-amino-2-methylpropan-2-ol (34
mg, 274 .mu.mol), sodium cyanoborohydride (57 mg, 914 .mu.mol) and
acetic acid (21 .mu.l, 366 .mu.mol). After stirring at 60.degree.
C. for 16 h, the resulting mixture was separated by preparative
RP-HPLC (Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA).
The product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 29 mg (31% of th.) of the
title compound.
[0880] LC-MS (method 2): R.sub.t=0.65 min; MS (ESIpos): m/z=511
(M+H).sup.+
[0881] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96 (s, 1H),
7.50-8.02 (br. s, 1H), 7.42 (s, 1H), 7.28 (s, 1H), 6.84 (s, 1H),
5.4-6.0 (br. s, 1H), 4.11 (s, 1H), 3.95 (s, 3H), 3.90 (s, 2H), 3.70
(d, 2H), 3.51-3.58 (m, 4H), 2.45 (s, 3H), 2.39-2.46 (m, 4H), 2.33
(d, 2H), 1.83-1.92 (m, 1H), 1.03 (s, 6H) ppm.
Example 55
1-(4-{[4-Amino-6-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-y-
l)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}piperazin-1-yl)ethanone
##STR00169##
[0883] A solution of Intermediate 20A (345 mg, 1.01 mmol) in acetic
acid (5 ml) was treated with 37% aq. formaldehyde solution (91
.mu.l, 1.22 mmol) and 1-acetylpiperazine (160 mg, 1.22 mmol). The
mixture was stirred at 60.degree. C. for 6 h and then evaporated.
The residue was dissolved in a mixture of THF/1 M aq. lithium
hydroxide solution (1:1, 10 ml) and stirred at rt for 2 h. The
mixture was then combined with the reaction mixture of a 100 mg
test run, and sat. aq. ammonium chloride solution was added. The
mixture was extracted with ethyl acetate, and the organic phase was
washed with sat. aq. sodium chloride solution, dried over magnesium
sulfate and evaporated under reduced pressure yielding 678 mg
(purity 87%, 94% of th.) of the title compound.
[0884] LC-MS (method 2): R.sub.t=0.71 min; MS (ESIpos): m/z=481
(M+H).sup.+.
Example 56
(3R)-3-[({7-[(4-Acetylpiperazin-1-yl)methyl]-4-amino-5-(7-methoxy-5-methyl-
-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl}methyl)amino]pyrro-
lidin-2-one
##STR00170##
[0886] A solution of Intermediate 31A (80 mg, 167 .mu.mol) in
methanol (1.4 ml) was treated with (3R)-3-aminopyrrolidin-2-one (21
mg, 251 .mu.mol), sodium cyanoborohydride (52 mg, 836 .mu.mol) and
acetic acid (19 .mu.l, 334 .mu.mol). After stirring at 60.degree.
C. for 16 h, the resulting mixture was separated by preparative
RP-HPLC (Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA).
The product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 46 mg (49% of th.) of the
title compound.
[0887] LC-MS (method 2): R.sub.t=0.70 min; MS (ESIpos): m/z=563
(M+H).sup.+
[0888] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.42 (s, 1H),
8.05 (s, 1H), 7.93-8.24 (br. s, 1H), 7.51 (s, 1H), 7.34 (s, 1H),
6.88 (s, 1H), 5.75-6.07 (br. s, 1H), 4.21-4.37 (m, 2H), 4.19 (s,
2H), 4.10 (t, 1H), 3.96 (s, 3H), 3.56-3.66 (m, 2H), 3.44-3.54 (m,
4H), 3.11-3.27 (m, 4H), 2.47 (s, 4H), 2.16-2.25 (m, 1H), 2.00 (s,
3H), 1.86-1.95 (m, 1H) ppm.
Example 57
1-(4-{[4-Amino-6-{[(2-hydroxy-2-methylpropyl)amino]methyl}-5-(7-methoxy-5--
methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-7-yl]methyl}piper-
azin-1-yl)ethanone
##STR00171##
[0890] A solution of Intermediate 31A (80 mg, 183 .mu.mol) in
methanol (2 ml) was treated with 1-amino-2-methylpropan-2-ol (31
mg, 251 .mu.mol), sodium cyanoborohydride (53 mg, 836 .mu.mol) and
acetic acid (19 .mu.l, 334 .mu.mol). After stirring at 60.degree.
C. for 16 h, the resulting mixture was separated by preparative
RP-HPLC (Reprosil C18, gradient 20-40% acetonitrile/0.2% aq. TFA).
The product thus obtained was dissolved in methanol and filtered
through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated yielding 29 mg (31% of th.) of the
title compound.
[0891] LC-MS (method 2): R.sub.t=0.71 min; MS (ESIpos): m/z=552
(M+H).sup.+
[0892] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96 (s, 1H),
7.59-8.02 (br. s, 1H), 7.42 (s, 1H), 7.28 (s, 1H), 6.84 (s, 1H),
5.45-5.93 (br. s, 1H), 4.13 (br. s, 1H), 3.95 (s, 3H), 3.93 (s,
2H), 3.71 (s, 2H), 3.37-3.44 (m, 4H), 2.45 (s, 3H), 2.32-2.43 (m,
6H), 1.97 (s, 3H), 1.03 (s, 6H) ppm.
Example 58
4-({4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-6-[(3-oxopiperazin-
-1-yl)methyl]pyrrolo[2,1-f][1,2,4]triazin-7-yl}methyl)piperazine-1-carbald-
ehyde formiate
##STR00172##
[0894] Acetic acid anhydride (498 .mu.l, 5.17 mmol) and formic acid
(237 .mu.l, 6.28 mmol) were stirred first 2 h at 50.degree. C. and
then overnight at rt. Subsequently, the mixture was diluted with
dichloromethane (5.1 ml), and 1.16 ml of this solution were added
to a solution of Example 49 (233 mg, 370 .mu.mol) in pyridine (89
.mu.l). After stirring at rt for 2 h, the mixture was diluted with
methanol and then evaporated. The residue was purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) affording 77 mg (35% of th.) of
title compound.
[0895] LC-MS (method 4): R.sub.t=0.62 min; MS (ESIpos): m/z=549
(M+H).sup.+
[0896] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.14
(s, 1H), 7.97 (s, 2H), 7.67 (s, 1H), 7.38 (s, 1H), 7.30 (s, 1H),
6.84 (s, 1H), 3.96 (s, 2H), 3.95 (s, 3H), 3.58 (s, 2H), 3.04 (br.
t, 2H), 2.86 (s, 2H), 2.45 (s, 3H), 2.42 (br. t, 2H) ppm.
Example 59
4-({7-[(4-Acetylpiperazin-1-yl)methyl]-4-amino-5-(7-methoxy-5-methyl-1-ben-
zothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl}methyl)piperazin-2-one
##STR00173##
[0898] A suspension of Example 49 (310 mg, 522 .mu.mol) in
dichloromethane/THF (2.5:1, 9.64 ml) was treated with sodium
carbonate (442 mg, 4.17 mmol) and stirred at rt for 30 min. Acetyl
chloride (74 .mu.l, 1.04 mmol) was added, and the resulting mixture
was stirred at rt for 2 h. After quenching with methanol, the
mixture was evaporated, and the residue was purified by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) affording 133 mg (45% of th.) of the title compound.
[0899] LC-MS (method 2): R.sub.t=0.69 min; MS (ESIpos): m/z=563
(M+H).sup.+
[0900] .sup.1H-NMR (400 MHz, methanol-d.sub.4): .delta.=7.89 (s,
1H), 7.30 (s, 1H), 7.28 (s, 1H), 6.79 (s, 1H), 4.13 (s, 2H), 3.98
(s, 3H), 3.74 (s, 2H), 3.59 (t, 2H), 3.54 (t, 2H), 3.18 (t, 2H),
3.02 (s, 2H), 2.68-2.55 (m, 6H), 2.48 (s, 3H), 2.08 (s, 3H)
ppm.
Example 60
Methyl
4-amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1--
ylcarbonyl)pyrrolo[2,1-f][1,2,4]triazine-6-carboxylate
bis(formiate)
##STR00174##
[0902] To a solution of Intermediate 28A (15 mg, 26 .mu.mol) in a
THF/methanol mixture (5:1, 180 .mu.l), cooled to 0.degree. C., was
added (trimethylsilyl)diazomethane (2 M solution in hexane, 15
.mu.l, 32 .mu.mol). The resulting mixture was slowly warmed to rt
over 30 min and then evaporated. The residue was dissolved in a 4 M
solution of hydrogen chloride in 1,4-dioxane (0.5 ml) and stirred
at rt for 1 h. After evaporation, the residue was purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) affording 4.8 mg (35% of th.) of
the title compound.
[0903] LC-MS (method 2): R.sub.t=0.66 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0904] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.17
(s, 2H), 8.38-8.23 (br. s, 1H), 8.04 (s, 1H), 7.43 (s, 1H), 7.31
(s, 1H), 6.86 (s, 1H), 5.71-5.58 (br. s, 1H), 3.96 (s, 3H), 3.60
(s, 3H, overlap with water peak), 2.46 (s, 3H) ppm.
Example 61
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-6-(1,3-oxazol-5-yl)-7-(piperaz-
in-1-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine
trihydrochloride
##STR00175##
[0906] A solution of Intermediate 25A (100 mg, 0.19 mmol) in
methanol (3.33 ml) was treated with
(4-toluenesulfonyl)methylisocyanide (36 mg, 0.19 mmol) and
potassium carbonate (25 mg, 186 mol). The mixture was refluxed for
6 h. It was then combined with the reactions mixtures from three 30
mg test runs and evaporated. The residue was dissolved in a 4 M
solution of hydrogen chloride in 1,4-dioxane (10 ml) and stirred at
rt for 2 h. After evaporation, the residue was purified by two-fold
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). Further purification by column
chromatography on silica gel (dichloromethane/methanol 5:1 with
0.5% aq. ammonia) and lyophilization from a 4 M solution of
hydrogen chloride in 1,4-dioxane afforded 75 mg (34% of th.) of the
title compound.
[0907] LC-MS (method 4): R.sub.t=0.69 min; MS (ESIpos): m/z=476
(M+H).sup.+
[0908] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.40
(s, 1H), 8.20 (s, 1H), 7.46 (s, 1H), 7.35 (s, 1H), 6.96 (s, 1H),
6.89 (s, 1H), 3.95 (s, 3H), 2.46 (s, 3H) ppm.
Example 62
6-(Aminomethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-
-ylmethyl)pyrrolo[2,1-f][1,2,4]triazin-4-amine trihydrochloride
##STR00176##
[0910] A suspension of Intermediate 29A (76 mg, 134 .mu.mol) and
10% Pd/C (60 mg) in a 0.5 M solution of hydrogen chloride in
methanol (20 ml) was stirred at rt for 3 h under 1 atm of hydrogen.
The mixture was then filtered through kieselguhr, the filtrate was
evaporated, and the residue was purified by preparative RP-HPLC
(Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. TFA). The
product thus obtained was lyophilized from a 4 M solution of
hydrogen chloride in 1,4-dioxane affording 34 mg (46% of th.) of
the title compound.
[0911] LC-MS (method 2): R.sub.t=0.55 min; MS (ESIpos): m/z=438
(M+H).sup.+
[0912] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.24
(br. s, 3H), 8.12 (s, 1H), 7.52 (s, 1H), 7.34 (s, 1H), 6.89 (s,
1H), 3.96 (s, 3H), 3.76-3.63 (m, 1H), 3.52-3.42 (m, 1H), 2.47 (s,
3H) ppm.
Example 63
N-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}acetamide
bis(trifluoroacetate)
##STR00177##
[0914] A solution of Intermediate 30A (210 mg, 362 .mu.mol) in
dichloromethane (26 ml) was treated with trifluoroacetic acid (5.2
ml) and stirred at rt for 1 h. After evaporation at rt, the residue
was purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA) affording 163 mg (63% of th.) of the
title compound.
[0915] LC-MS (method 3): R.sub.t=2.42 min; MS (ESIpos): m/z=480
(M+H).sup.+
[0916] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=9.00-8.75 (br. s, 1H), 8.30-8.16 (br. s, 1H), 8.08 (s, 1H),
7.38 (s, 1H), 7.33 (s, 1H), 6.87 (s, 1H), 4.32 (br. d, 2H), 3.96
(s, 3H), 3.40-2.98 (m, 8H), 2.46 (s, 3H), 1.77 (s, 3H) ppm.
Example 64
N-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylm-
ethyl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}acetamide
dihydrochloride
##STR00178##
[0918] A solution of Intermediate 30A (80 mg, 0.14 mmol) in
dichloromethane (10 ml) and trifluoroacetic acid (2 ml) was stirred
at rt for 1 h and then evaporated. Purification by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid) and lyophilization from a mixture of methanol and 4 M
solution of hydrogen chloride in 1,4-dioxane afforded 39 mg (50% of
th.) of the title compound.
[0919] LC-MS (method 5): R.sub.t=1.56 min; MS (ESIpos): m/z=480
(M+H).sup.+
[0920] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=8.12
(s, 1H), 7.38 (s, 1H), 7.33 (s, 1H), 6.87 (s, 1H), 4.37 (br. d,
2H), 3.96 (s, 3H), 3.85-3.20 (m, 8H, overlap with water peak), 2.46
(s, 3H), 1.76 (s, 3H) ppm.
Example 65
N-({4-Amino-7-[(4-formylpiperazin-1-yl)methyl]-5-(7-methoxy-5-methyl-1-ben-
zothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl}methyl)acetamide
formiate
##STR00179##
[0922] Acetic acid anhydride (304 .mu.l, 3.16 mmol) and formic acid
(145 .mu.l, 3.16 mmol) were stirred first 2 h at 50.degree. C. and
then overnight at rt. Subsequently, the mixture was diluted with
dichloromethane (3.1 ml), and 663 .mu.l of this solution were added
to a solution of Example 63 (160 mg, 226 .mu.mol) in pyridine (54
.mu.l). The mixture was stirred at rt for 2 h, then diluted with
methanol (1 ml), and stirring was continued at 40.degree. C. for
another 2 h. After evaporation, the residue was purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) affording 74 mg (61% of th.) of
title compound.
[0923] LC-MS (method 4): R.sub.t=0.65 min; MS (ESIpos): m/z=508
(M+H).sup.+
[0924] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.13 (s, 1H),
8.06-7.89 (m, 3H), 7.38 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.28
(d, 2H), 4.01-3.88 (m, 5H), 2.47-2.34 (m, 7H), 1.75 (s, 3H)
ppm.
Example 66
N-({7-[(4-Acetylpiperazin-1-yl)methyl]-4-amino-5-(7-methoxy-5-methyl-1-ben-
zothiophen-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl}methyl)acetamide
##STR00180##
[0926] A solution of Example 63 (110 mg, 229 .mu.mol) and acetyl
chloride (32 .mu.l, 458 .mu.mol) in THF/dichloromethane (1:2, 3 ml)
was treated with sodium carbonate (194 mg, 1.83 mmol) and stirred
at rt overnight. Then, the mixture was diluted with methanol (2 ml)
and water (1 ml) and stirred at rt for 1 h. After evaporation, the
residue was purified by preparative RP-HPLC (Reprosil C18, gradient
10-95% acetonitrile/0.1% aq. formic acid). Lyophilization from
1,4-dioxane and re-purification by column chromatography on silica
gel (dichloromethane/methanol 50:1.fwdarw.100% methanol) afforded
35 mg (28% of th.) of the title compound.
[0927] LC-MS (method 2): R.sub.t=0.70 min; MS (ESIpos): m/z=522
(M+H).sup.+
[0928] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.04-7.98 (m,
2H), 7.38 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.27 (d, 2H), 3.95
(s, 3H), 3.91 (s, 2H), 3.44-3.36 (m, 4H), 2.48-2.35 (m, 7H), 1.98
(s, 3H), 1.75 (s, 3H) ppm.
Example 67
N-({4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-[(3-oxopiperazin-
-1-yl)methyl]-pyrrolo[2,1-f][1,2,4]triazin-6-yl}methyl)acetamide
##STR00181##
[0930] A suspension of Intermediate 32A (35 mg, 73 .mu.mol) in
methanol (4.1 ml) was treated with acetic anhydride (13 .mu.l, 146
.mu.mol) and 10% Pd/C (41 mg) and stirred at rt for 1 h under 1 atm
of hydrogen. Filtration over kieselguhr and evaporation of the
filtrate afforded 30 mg (79% of th.) of the title compound.
[0931] LC-MS (method 5): R.sub.t=1.65 min; MS (ESIpos): m/z=494
(M+H).sup.+
[0932] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 2H),
7.75 (br. s, 1H), 7.39 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.27
(s, 1H), 4.26 (s, 1H), 3.95 (s, 5H), 3.19-3.08 (m, 2H), 3.05-2.96
(m, 2H), 2.66-2.57 (m, 2H), 2.45 (s, 3H), 1.72 (s, 3H) ppm.
Example 68
4-Amino-6-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazine-7-carbonitrile
##STR00182##
[0934] A solution of Intermediate 38A (1 g, 2.08 mmol) in THF (10
ml) was treated with a 1 M solution of tetra-n-butylammonium
fluoride in THF (12 ml, 12 mmol). The mixture was stirred at rt
overnight and then evaporated. The residue was taken up in water
and extracted three times with ethyl acetate. The combined organic
phases were dried over magnesium sulfate and evaporated. The
residue was triturated in tert-butyl methyl ether, and the solid
was filtered off affording 680 mg (78% of th.) of the title
compound.
[0935] LC-MS (method 2): R.sub.t=0.95 min; MS (ESIpos): m/z=366
(M+H).sup.+
[0936] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.61-8.29 (br.
s, 1H), 8.18 (s, 1H), 7.43 (s, 1H), 7.32 (s, 1H), 6.87 (s, 1H),
6.32-6.03 (br. s, 1H), 5.46 (t, 1H), 4.55 (d, 2H), 3.96 (s, 3H),
2.45 (s, 3H) ppm.
Example 69
4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazine-7-carbonitrile
##STR00183##
[0938] A solution of Example 68 (60 mg, 164 .mu.mol) in
dichloromethane (5 ml) was treated with thionyl chloride (18 .mu.l,
246 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in methanol (2 ml) and
treated with DIPEA (57 .mu.l, 328 .mu.mol). The mixture was stirred
first 2 h at 60.degree. C., then refluxed overnight and finally
heated to 150.degree. C. for 30 min in a microwave device. After
this, the mixture was evaporated, and the residue was purified by
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). The product thus obtained was dissolved
in methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated affording 18
mg (28% of th.) of the title compound.
[0939] LC-MS (method 4): R.sub.t=1.12 min; MS (ESIpos): m/z=379
(M+H).sup.+
[0940] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.61-8.40 (br.
s, 1H), 8.21 (s, 1H), 7.42 (s, 1H), 7.33 (s, 1H), 6.87 (s, 1H),
6.34-6.09 (br. s, 1H), 4.47 (s, 2H), 3.96 (s, 3H), 3.26 (s, 3H),
2.45 (s, 3H) ppm.
Example 70
4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrol-
o[2,1-f][1,2,4]-triazine-7-carbonitrile
##STR00184##
[0942] A solution of Example 68 (60 mg, 164 .mu.mol) in
dichloromethane (5 ml) was treated with thionyl chloride (18 .mu.l,
246 .mu.mol). The mixture was stirred at rt for 15 min and then
evaporated. The residue was dissolved in ethanol (2 ml) and treated
with DIPEA (57 .mu.l, 328 .mu.mol). The mixture was stirred at
60.degree. C. overnight and then heated to 150.degree. C. for 30
min in a microwave device. After this, the mixture was evaporated,
and the residue was purified by preparative RP-HPLC (Reprosil C18,
gradient 10-95% acetonitrile/0.1% aq. TFA). The product thus
obtained was dissolved in ethanol (2 ml) and again heated to
150.degree. C. for 30 min in the microwave oven. DIPEA (57 .mu.l,
328 .mu.mol) was added, and heating to 150.degree. C. in the
microwave oven was continued for further 30 min. After evaporation,
the residue was purified by column chromatography on silica gel
(dichloromethane/methanol 95:5) affording 16 mg (23% of th.) of the
title compound.
[0943] LC-MS (method 2): R.sub.t=1.18 min; MS (ESIpos): m/z=393
(M+H).sup.+
[0944] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.61-8.38 (br.
s, 1H), 8.20 (s, 1H), 7.42 (s, 1H), 7.33 (s, 1H), 6.87 (s, 1H),
6.33-6.08 (br. s, 1H), 4.51 (s, 2H), 3.96 (s, 3H), 3.45 (q, 2H),
2.45 (s, 3H), 1.10 (t, 3H) ppm.
Example 71
4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-6-[(3-oxopiperazin-1-y-
l)methyl]pyrrolo[2,1-f][1,2,4]triazine-7-carbonitrile
##STR00185##
[0946] A solution of Intermediate 39A (20 mg, 55 .mu.mol) in THF
(0.73 ml) was treated with acetic acid (6 .mu.l, 110 .mu.mol),
2-oxopiperazine (27 mg, 275 .mu.mol) and sodium
triacetoxyborohydride (58 mg, 275 .mu.mol). After stirring at rt
for 3 h, further amounts of THF (1 ml), acetic acid (6 .mu.l, 110
.mu.mol), 2-oxopiperazine (27 mg, 275 .mu.mol) and sodium
triacetoxyborohydride (58 mg, 275 .mu.mol) were added, and stirring
at rt was continued overnight. After evaporation, the residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). The product thus obtained was dissolved
in methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated affording 13
mg (52% of th.) of the title compound.
[0947] LC-MS (method 2): R.sub.t=0.86 min; MS (ESIpos): m/z=447
(M+H).sup.+
[0948] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.60-8.31 (br.
s, 1H), 8.19 (s, 1H), 7.74 (s, 1H), 7.44 (s, 1H), 7.33 (s, 1H),
6.87 (s, 1H), 6.25-5.98 (br. s, 1H), 3.96 (s, 3H), 3.62 (s, 2H),
3.13-3.07 (m, 2H), 2.91 (s, 2H), 2.45 (s, 3H) ppm.
Example 72
N,N'-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f][1-
,2,4]triazine-6,7-diyl]bis(methylene)}diacetamide
##STR00186##
[0950] A suspension of Intermediate 40A (crude, 85 mg), 10% Pd/C
(115 mg) and acetic anhydride (40 .mu.l, 435 .mu.mol) in methanol
(12 ml) was stirred at rt under 1 atm of hydrogen. After 4 h,
further amounts of 10% Pd/C (115 mg) and acetic anhydride (40
.mu.l, 435 .mu.mol) were added, and stirring at rt under 1 atm of
hydrogen was continued for 2 h. The resulting mixture was filtered
through kieselguhr, the filtrate was evaporated, and the residue
was purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. TFA). The product thus obtained was dissolved
in methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated affording 18
mg (18% of th.) of the title compound.
[0951] LC-MS (method 2): R.sub.t=0.73 min; MS (ESIpos): m/z=452
(M+H).sup.+
[0952] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.29 (br. t,
1H), 8.04 (br. t, 1H), 8.00 (s, 1H), 7.34 (s, 1H), 7.31 (s, 1H),
6.85 (s, 1H), 4.58 (d, 2H), 4.26 (d, 2H), 3.95 (s, 3H), 2.45 (s,
3H), 1.81 (s, 3H), 1.74 (s, 3H) ppm.
Example 73
2-[4-Amino-6-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-7-yl]propan-2-ol
##STR00187##
[0954] A flask containing a suspension of Intermediate 43A (180 mg,
598 .mu.mol), Intermediate 6A (299 mg, 897 .mu.mol) and caesium
fluoride (454 mg, 2.99 mmol) in THF/water (10:1, 11 ml) was
degassed under reduced pressure and then refilled with argon.
4-(Di-tert-butylphosphino)-N,N-dimethylaniline-dichloropalladium
(2:1; 13 mg, 18 .mu.mol) was added. The resulting mixture was
degassed again and stirred under argon at 50.degree. C. for 16 h.
After this, the reaction mixture was separated by preparative
RP-HPLC (Reprosil C18, gradient 30-50% acetonitrile/0.2% aq. TFA).
The product fractions were diluted with a 7 M solution of ammonia
in methanol and then concentrated under reduced pressure. The
precipitate was filtered off, washed with water and dried in vacuo
yielding 99 mg (42% of th.) of the title compound.
[0955] LC-MS (method 2): R.sub.t=0.89 min; MS (ESIpos): m/z=399
(M+H).sup.+
[0956] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.96 (s, 1H),
7.51-8.02 (br. s, 1H), 7.37 (s, 1H), 7.31 (s, 1H), 6.86 (s, 1H),
5.99 (s, 1H), 5.23-5.82 (br. s, 1H), 5.04 (t, 1H), 4.53 (d, 2H),
3.96 (s, 3H), 2.45 (s, 3H), 1.73 (s, 6H) ppm.
Example 74
4-{[4-Amino-7-(2-hydroxypropan-2-yl)-5-(7-methoxy-5-methyl-1-benzothiophen-
-2-yl)pyrrolo[2,1-f][1,2,4]triazin-6-yl]methyl}piperazin-2-one
##STR00188##
[0958] A solution of Intermediate 44A (50 mg, 73% purity, 92
.mu.mol) in methanol (4 ml) was treated with 2-oxopiperazine (28
mg, 276 .mu.mol), sodium cyanoborohydride (23 mg, 368 .mu.mol) and
acetic acid (21 .mu.l, 368 .mu.mol). The mixture was stirred first
18 h at 60.degree. C. and then 3 days at rt. After this, the
mixture was separated by preparative RP-HPLC (Reprosil C18,
gradient 20-40% acetonitrile/0.2% aq. TFA). The product fractions
were diluted with a 7 M solution of ammonia in methanol and
evaporated under reduced pressure. The residue was dissolved in
ethyl acetate and washed with water. The organic phase was dried
with sodium sulfate and concentrated under reduced pressure. The
residue was dissolved in 1,4-dioxane and lyophilized yielding 24 mg
(52% of th.) of the title compound.
[0959] LC-MS (method 5): R.sub.t=1.85 min; MS (ESIpos): m/z=481
(M+H).sup.+
[0960] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.73-8.01 (br. s, 1H), 7.41 (s, 1H), 7.33 (s, 1H), 6.87 (s, 1H),
5.2-5.7 (br. s, 1H), 3.95 (s, 3H), 3.57 (s, 2H), 3.14 (br. s, 2H),
2.60 (br. s, 4H), 2.46 (s, 3H), 1.70 (br. s, 6H) ppm.
Example 75
[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-methylpyrrolo[2,1-f-
][1,2,4]triazin-6-yl]-methanol
##STR00189##
[0962] Under an argon atmosphere, a flask was charged with 165 mg
(0.64 mmol) of Intermediate 53A, 143 mg (0.64 mmol) of Intermediate
5A, 25 mg (0.03 mmol)
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; see S. L. Buchwald et al., J. Am.
Chem. Soc. 132 (40), 14073-14075 (2010)) and 409 mg (1.93 mmol)
potassium phosphate. Then, 7 ml of a degassed mixture of
1,4-dioxane and water (5:1) were added, and the solution was
stirred at 70.degree. C. for 1 h. Another portion of Intermediate
5A (142 mg, 0.64 mmol) and of
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (10 mg, 0.012 mmol) were added, and
stirring was continued for 1 h. After this, the reaction mixture
was partially evaporated under reduced pressure, water was added,
and the mixture was extracted with ethyl acetate. The combined
organic phases were evaporated under reduced pressure, and the
residue was triturated with acetonitrile. The precipitate was
filtered off and dried in vacuo yielding 180 mg (92% purity by
LC-MS, 73% of th.) of the title compound. From the filtrate a
second batch (41 mg, 18% of th.) was obtained by preparative
RP-HPLC (Reprosil C18, gradient 10-95% acetonitrile/0.1% aq. formic
acid). Total yield: 91% of th.
[0963] LC-MS (method 5): R.sub.t=1.92 min; MS (ESIpos): m/z=355
(M+H).sup.+
[0964] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.95 (s, 1H),
7.35 (s, 1H), 7.30 (s, 1H), 6.83 (s, 1H), 4.82 (t, 1H), 4.44 (d,
2H), 3.95 (s, 3H), 2.45 (s, 3H) ppm.
Example 76
4-{[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-methylpyrrolo[2,-
1-f][1,2,4]triazin-6-yl]methyl}piperazin-2-one
##STR00190##
[0966] A suspension of 55 mg (0.16 mmol) of Intermediate 54A in THF
(1.5 ml) was treated with 78 mg (0.78 mmol) piperazin-2-one, 18
.mu.l (0.31 mmol) acetic acid and 166 mg (0.78 mmol)
triacetoxyborohydride. The mixture was stirred at ambient
temperature overnight. Then, 1.5 ml water were added, and most of
the THF solvent was evaporated under reduced pressure. The
remaining mixture was diluted with more water, and the precipitated
solid was filtered off and dried (35 mg). This material was further
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid). The product fractions were
adjusted to pH 9 with solid potassium carbonate and partially
concentrated under reduced pressure. The precipitated solid was
filtered off and dried at 45.degree. C. in vacuo to afford 16 mg
(24% of th.) of the title compound.
[0967] LC-MS (method 5): R.sub.t=1.74 min; MS (ESIpos): m/z=437
(M+H).sup.+
[0968] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.95 (s, 1H),
7.66 (br. s, 1H), 7.35 (s, 1H), 7.30 (s, 1H), 6.83 (s, 1H), 3.95
(s, 3H), 3.50 (s, 2H), 3.05 (m, 2H), 2.84 (s, 2H), 2.45 (m, 5H)
ppm.
Example 77
1-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-methylpyrrolo[2-
,1-f][1,2,4]triazin-6-yl]methyl}amino)-2-methylpropan-2-ol
formiate
##STR00191##
[0970] A suspension of 55 mg (0.16 mmol) of Intermediate 54A in THF
(1.5 ml) was treated with 98 mg (0.78 mmol)
1-amino-2-methylpropan-2-ol hydrochloride, 39 mg (0.47 mmol) sodium
acetate and 166 mg (0.78 mmol) sodium triacetoxyborohydride. The
mixture was stirred at ambient temperature overnight. Then, 1.5 ml
water were added, and the mixture was evaporated under reduced
pressure. The residue was purified by preparative RP-HPLC (Reprosil
C18, gradient 10-95% acetonitrile/0.1% aq. formic acid) to afford
38 mg (52% of th.) of the title compound.
[0971] LC-MS (method 5): R.sub.t=1.68 min; MS (ESIpos): m/z=426
(M+H).sup.+
[0972] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.19 (s, 1H),
7.94 (s, 1H), 7.38 (s, 1H), 7.29 (s, 1H), 6.84 (s, 1H), 3.95 (s,
3H), 2.45 (s, 3H), 2.34 (s, 2H), 1.02 (s, 6H) ppm.
Example 78
1-({[4-Amino-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-methylpyrrolo[2-
,1-f][1,2,4]triazin-6-yl]methyl}amino)-2-methylpropan-2-ol
##STR00192##
[0974] A solution of 29 mg (0.06 mmol) of Example 77 in 3 ml of
methanol was run through a Stratospheres SPE PL-HCO.sub.3 MP-resin
cartridge, preconditioned with 2 ml of methanol. The cartridge was
washed with 4 ml of methanol, and the eluate was evaporated to
afford 21.5 mg (82% of th.) of the title compound.
[0975] LC-MS (method 4): R.sub.t=0.68 min; MS (ESIpos): m/z=426
(M+H).sup.+
[0976] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.94 (s, 1H),
7.37 (s, 1H), 7.28 (s, 1H), 6.83 (s, 1H), 4.07 (s, 1H), 3.95 (s,
3H), 3.63-3.75 (m, 2H), 2.45 (s, 3H), 2.32 (m, 3H), 1.42-1.56 (m,
1H), 1.02 (s, 6H) ppm.
Example 79
[4-Amino-7-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-f-
][1,2,4]triazin-6-yl]-methanol
##STR00193##
[0978] A solution of Intermediate 48A (117 mg, 0.24 mmol) in 5 ml
THF was treated with 5 ml conc. hydrochloric acid and stirred at
ambient temperature overnight. Then, 12 ml of 5 M aq. sodium
hydroxide solution as well as ethyl acetate were added, the layers
were separated, and the aqueous layer was extracted twice with
ethyl acetate. The combined organic phases were washed with sat.
aq. sodium chloride solution, dried and evaporated. The residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) to afford 30 mg (34% of th.) of
the title compound.
[0979] LC-MS (method 2): R.sub.t=0.90 min; MS (ESIpos): m/z=375
(M+H).sup.+
[0980] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.04 (s, 1H),
7.43 (s, 1H), 7.32 (s, 1H), 6.85 (s, 1H), 5.02 (t, 1H), 4.43 (d,
2H), 3.96 (s, 3H), 2.45 (s, 3H) ppm. 9.2 mg (10% of th.) of
7-chloro-6-(chloromethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrr-
olo[2,1-f][1,2,4]triazin-4-amine were isolated as a by-product (cf
Intermediate 50A).
Example 80
4-{[4-Amino-7-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2,-
1-f][1,2,4]triazin-6-yl]methyl}piperazin-2-one
##STR00194##
[0982] The title compound was prepared according to the procedure
of Example 76 starting from 40 mg (0.11 mmol) of Intermediate 49A.
Yield: 27 mg (55% of th.).
[0983] LC-MS (method 2): R.sub.t=0.86 min; MS (ESIpos): m/z=457
(M+H).sup.+
[0984] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.05 (s, 1H),
7.68 (br. s, 1H), 7.44 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 3.96
(s, 3H), 3.52 (s, 2H), 3.05 (br. s, 2H), 2.86 (s, 2H), 2.45 (s, 3H)
ppm.
Example 81
1-({[4-Amino-7-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2-
,1-f][1,2,4]triazin-6-yl]methyl}amino)-2-methylpropan-2-ol
formiate
##STR00195##
[0986] The title compound was prepared according to the procedure
of Example 77 starting from 40 mg (0.11 mmol) of Intermediate 49A.
Yield: 24 mg (45% of th.).
[0987] LC-MS (method 2): R.sub.t=0.73 min; MS (ESIpos): m/z=446
(M+H).sup.+
[0988] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.15 (s, 1H),
8.04 (s, 1H), 7.45 (s, 1H), 7.30 (s, 1H), 6.85 (s, 1H), 3.96 (s,
3H), 3.73 (s, 2H), 2.45 (s, 3H), 2.32 (s, 2H), 1.00 (s, 6H)
ppm.
Example 82
1-({[4-Amino-7-chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrrolo[2-
,1-f][1,2,4]triazin-6-yl]methyl}amino)-2-methylpropan-2-ol
##STR00196##
[0990] The title compound was prepared from Example 81 according to
the procedure of Example 78.
[0991] LC-MS (method 2): R.sub.t=0.72 min; MS (ESIpos): m/z=446
(M+H).sup.+
[0992] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.04 (s, 1H),
7.45 (s, 1H), 7.30 (s, 1H), 6.85 (s, 1H), 4.09 (br. s, 1H), 3.95
(s, 3H), 3.71 (s, 2H), 2.45 (s, 3H), 2.31 (s, 2H), 1.44-1.64 (m,
1H), 1.00 (s, 6H) ppm.
Example 83
7-Chloro-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazin-4-amine
##STR00197##
[0994] 8.2 mg (0.02 mmol) of Intermediate 50A were suspended in 1
ml of ethanol, treated with 41 .mu.l (0.11 mmol) of a 2.68 M
solution of sodium ethanolate in ethanol and refluxed for 1 min.
The clear solution was then evaporated, and the crude product was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) to afford 5 mg (56% of th.) of
the title compound.
[0995] LC-MS (method 4): R.sub.t=1.25 min; MS (ESIpos): m/z=402
(M+H).sup.+
[0996] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.06 (s, 1H),
7.40 (s, 1H), 7.32 (s, 1H), 6.86 (s, 1H), 4.40 (s, 2H), 3.96 (s,
3H), 3.40 (q, 2H), 2.45 (s, 3H), 1.06 (t, 3H) ppm.
Example 84
5-(7-Methoxy-5-methyl-1-benzothiophen-2-yl)-6-methyl-7-(piperazin-1-ylmeth-
yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine formiate
##STR00198##
[0998] A solution of Intermediate 57A (100 mg, 191 .mu.mol) in a 4
M solution of hydrogen chloride in 1,4-dioxane (2 ml) was stirred
at rt for 3 h and then evaporated. Purification by two-fold
preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. formic acid) afforded 71 mg (67% of th.) of
the title compound.
[0999] LC-MS (method 2): R.sub.t=0.81 min; MS (ESIpos): m/z=423
(M+H).sup.+
[1000] .sup.1H-NMR (400 MHz, CDCl.sub.3): .delta.=8.46 (br. s, 1H),
7.89 (s, 1H), 7.27 (s, 1H, overlap with CHCl.sub.3 peak), 7.18 (s,
1H), 6.67 (s, 1H), 4.01 (s, 3H), 3.97 (s, 2H), 3.18-3.09 (m, 4H),
2.82-2.73 (m, 4H), 2.50 (s, 3H), 2.24 (s, 3H) ppm.
Example 85
6-Chloro-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-7-(piperazin-1-ylmeth-
yl)pyrrolo[2,1-f]-[1,2,4]triazin-4-amine trihydrochloride
##STR00199##
[1002] Intermediate 60A (65 mg, 0.12 mmol) was stirred in 1 ml of a
4 M solution of hydrogen chloride in 1,4-dioxane for 2 h at rt. The
suspension was evaporated to dryness, and the crude product was
purified by preparative RP-HPLC (Reprosil C18, gradient 10-95%
acetonitrile/0.1% aq. hydrochloric acid) affording 49 mg (74% of
th.) of the title compound.
[1003] LC-MS (method 2): R.sub.t=0.81 min; MS (ESIpos): m/z=443
(M+H).sup.+
[1004] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al. .delta.=9.56
(br. s, 1H), 8.45 (br. s, 1H), 8.19 (s, 1H), 7.44 (s, 1H), 7.36 (s,
1H), 6.89 (s, 1H), 6.30 (br. s, 1H), 4.63 (br. s, 1H), 3.40 (br. s,
8H), 2.46 (s, 3H) ppm.
Example 86
[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazin-7-yl]methanol
##STR00200##
[1006] The title compound (360 mg) was obtained as a side product
in the preparation of Example 5 by preparation method 1.
[1007] LC-MS (method 2): R.sub.t=0.99 min; MS (ESIpos): m/z=399
(M+H).sup.+
[1008] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.09-7.72 (br.
s, 1H), 7.99 (s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H),
6.05-5.48 (br. s, 1H), 5.04 (br. s, 1H), 4.81 (d, 2H), 4.47 (s,
2H), 3.96 (s, 3H), 3.39 (q, 3H), 2.45 (s, 3H), 1.05 (t, 3H)
ppm.
Example 87
1-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-7-yl]methyl}imidazolidin-2-one
##STR00201##
[1010] A suspension of Intermediate 63A (100 mg, 0.221 mmol) in THF
(5 ml) was treated with imidazolidin-2-one (57 mg, 0.662 mmol) and
N,N-diisopropylethylamine (153 .mu.l, 0.926 mmol), and the mixture
was heated to 150.degree. C. for 90 min in a microwave oven. After
this, the reaction mixture was purified by preparative RP-HPLC
(Reprosil C18, gradient 40-60% acetonitrile/0.2% aq.
trifluoroacetic acid). The product thus obtained was dissolved in
methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated. The product
was purified once again by preparative thin layer chromatography
over silica gel (cyclohexane/ethyl acetate 3:1) affording 24 mg
(22% of th.) of the title compound.
[1011] LC-MS (method 2): R.sub.t=0.99 min; MS (ESIpos): m/z=467
(M+H).sup.+
[1012] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.21-7.65 (br.
s, 1H), 7.99 (s, 1H), 7.36 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H),
6.38 (s, 1H), 6.04-5.56 (br. s, 1H), 4.65 (s, 2H), 4.46 (s, 2H),
3.95 (s, 3H), 3.38 (q, 2H), 3.31-3.10 (m, 4H), 2.45 (s, 3H), 1.06
(t, 3H) ppm.
Example 88
4-{[4-Amino-5-(7-methoxy-1-benzothiophen-2-yl)-6-(methoxymethyl)pyrrolo[2,-
1-f][1,2,4]triazin-7-yl]methyl}piperazin-2-one
##STR00202##
[1014] To a solution of Intermediate 62A (72.6 mg, 197 .mu.mol),
(7-methoxy-1-benzothiophen-2-yl)boronic acid (45 mg, 216 .mu.mol)
and caesium fluoride (149 mg, 983 .mu.mol) in degassed THF/water
(10:1, 2.2 ml) under argon was added
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 7.7 mg, 9.8 .mu.mol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010)).
The resulting mixture was degassed again and stirred under argon at
60.degree. C. for 3 h. After this, the mixture was separated by
preparative RP-HPLC (Reprosil C18, gradient 20-40%
acetonitrile/0.1% aq. TFA). The product fractions were combined and
evaporated to dryness. The residue was dissolved in methanol and
filtered through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 31 mg (35% of th.) of the
title compound.
[1015] LC-MS (method 2): R.sub.t=0.71 min; MS (ESIpos): m/z=453
(M+H).sup.+
[1016] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.72 (s, 1H), 7.61-8.23 (br. s, 1H), 7.51 (d, 1H), 7.46 (s, 1H),
7.40 (t, 1H), 6.99 (d, 1H), 5.57-6.11 (br. s, 1H), 4.42 (s, 2H),
3.98 (s, 5H), 3.20 (s, 3H), 3.11 (br. m, 2H), 3.01 (s, 2H),
2.62-2.67 (m, 2H) ppm.
Example 89
4-{[4-Amino-6-(methoxymethyl)-5-(5-methyl-1-benzothiophen-2-yl)pyrrolo[2,1-
-f][1,2,4]triazin-7-yl]methyl}piperazin-2-one
##STR00203##
[1018] To a solution of Intermediate 62A (50 mg, 135 .mu.mol),
(5-methyl-1-benzothiophen-2-yl)boronic acid (28.6 mg, 149 .mu.mol)
and caesium fluoride (103 mg, 677 .mu.mol) in degassed THF/water
(10:1, 4.4 ml) under argon was added
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 5.3 mg, 6.8 .mu.mol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010)).
The resulting mixture was degassed again and stirred under argon at
60.degree. C. for 16 h. After this, the mixture was separated by
preparative RP-HPLC (Reprosil C18, gradient 30-50%
acetonitrile/0.1% aq. TFA). The product fractions were combined and
evaporated to dryness. The residue was dissolved in methanol and
filtered through an anion exchange cartridge (StratoSpheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 27 mg (45% of th.) of the
title compound.
[1019] LC-MS (method 2): R.sub.t=0.79 min; MS (ESIpos): m/z=437
(M+H).sup.+
[1020] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.89 (d, 1H), 7.73 (s, 1H), 7.71 (s, 1H), 7.58-8.18 (br. s, 1H),
7.40 (s, 1H), 7.24 (dd, 1H), 5.58-6.03 (m, 1H), 4.42 (s, 2H), 3.97
(s, 2H), 3.21 (s, 3H), 3.11 (br. t, 2H), 3.01 (s, 2H), 2.64 (t,
2H), 2.44 (s, 3H) ppm.
Example 90
1-[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyr-
rolo[2,1-f][1,2,4]-triazin-7-yl]ethanol
##STR00204##
[1022] Under argon, a 1 M solution of methylmagnesium bromide in
THF (630 .mu.l, 630 .mu.mol) was added dropwise at rt to a solution
of Intermediate 17A (100 mg, 252 .mu.mol) in THF (10 ml). The
mixture was stirred at rt for 3 h and then treated with another
portion of methylmagnesium bromide in THF (177 .mu.l, 177 .mu.mol).
The reaction mixture was stirred for further 16 h, then quenched
with sat. aq. ammonium chloride solution and extracted with ethyl
acetate. The combined organic layers were washed with brine, dried
with magnesium sulfate, filtered and evaporated. The residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 40-60%
acetonitrile/0.1% aq. TFA). The product fractions were diluted with
sat. aq. sodium hydrogencarbonate solution and extracted with ethyl
acetate. The organic phase was washed with sat. aq. sodium chloride
solution, dried with magnesium sulfate, filtered and evaporated
yielding 39 mg (36% of th.) of the title compound.
[1023] LC-MS (method 4): R.sub.t=1.07 min; MS (ESIpos): m/z=413
(M+H).sup.+
[1024] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.95 (s, 1H),
7.56-8.07 (br. s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H),
5.46-5.54 (m, 1H), 5.37-5.95 (br. s, 1H), 5.26 (d, 1H), 4.67 (d,
1H), 4.38 (d, 1H), 3.95 (s, 3H), 3.38 (q, 2H), 2.45 (s, 3H), 1.52
(d, 3H), 1.05 (t, 3H) ppm.
Example 91
[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)pyrro-
lo[2,1-f][1,2,4]-triazin-7-yl](cyclopropyl)methanol
##STR00205##
[1026] Under argon, a 0.5 M solution of cyclopropylmagnesium
bromide in THF (1.26 ml, 630 .mu.mol) was added dropwise at rt to a
solution of Intermediate 17A (100 mg, 252 .mu.mol) in THF (5 ml).
The mixture was stirred at rt for 1 h, then quenched with sat. aq.
ammonium chloride solution and extracted with ethyl acetate. The
combined organic layers were washed with brine, dried with
magnesium sulfate, filtered and evaporated. The residue was
purified by preparative RP-HPLC (Reprosil C18, gradient 50-70%
acetonitrile/0.1% aq. TFA). The product fractions were diluted with
sat. aq. sodium hydrogencarbonate solution and extracted with ethyl
acetate. The organic phase was washed with sat. aq. sodium chloride
solution, dried with magnesium sulfate, filtered and evaporated
yielding 10 mg (10% of th.) of the title compound.
[1027] LC-MS (method 2): R.sub.t=1.13 min; MS (ESIpos): m/z=439
(M+H).sup.+
[1028] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.94 (s, 1H),
7.68-8.04 (br. s, 1H), 7.36 (s, 1H), 7.31 (s, 1H), 6.84 (s, 1H),
5.47-5.89 (br. s, 1H), 5.30 (d, 1H), 4.64 (d, 1H), 4.59-4.67 (m,
1H), 4.39 (d, 1H), 3.96 (s, 3H), 3.38 (q, 2H), 2.45 (s, 3H),
1.54-1.64 (m, 1H), 1.04 (t, 3H), 0.51-0.59 (m, 1H), 0.40-0.47 (m,
1H), 0.28-0.38 (m, 2H) ppm.
Example 92
(3S)-3-({[4-Amino-6-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen--
2-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
##STR00206##
[1030] A suspension of Intermediate 13A (100 mg, 0.233 mmol) in THF
(2 ml) was treated with (S)-3-aminopyrrolidin-2-one (35 mg, 0.349
mmol), sodium triacetoxyborohydride (148 mg, 0.698 mmol) and acetic
acid (26.6 .mu.l, 0.465 mmol). The resulting mixture was stirred at
rt for 3 h and then directly purified by preparative RP-HPLC
(Reprosil C18, gradient 40-60% acetonitrile/0.2% aq.
trifluoroacetic acid). The product fractions were combined and
evaporated to dryness. The residue was dissolved in methanol and
filtered through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 56 mg (51% of th.) of the
title compound.
[1031] LC-MS (method 4): R.sub.t=0.72 min; MS (ESIpos): m/z=467
(M+H).sup.+
[1032] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.01 (s, 1H),
7.76 (s, 1H), 7.35 (s, 1H), 7.31 (s, 1H), 6.85 (s, 1H), 4.48 (d,
2H), 4.43 (d, 2H), 4.24-4.02 (m, 2H), 3.96 (s, 3H), 3.26-2.99 (m,
6H), 2.45 (s, 3H), 2.40-2.27 (m, 1H), 1.82-1.64 (m, 1H) ppm.
Example 93
(3S)-3-({[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-
-yl)pyrrolo[2,1-f]-[1,2,4]triazin-7-yl]methyl}amino)pyrrolidin-2-one
##STR00207##
[1034] A suspension of Intermediate 17A (100 mg, 0.252 mmol) in THF
(2 ml) was treated with (S)-3-aminopyrrolidin-2-one (38 mg, 378
.mu.mol), sodium triacetoxyborohydride (160 mg, 757 .mu.mol) and
acetic acid (30 .mu.l, 504 .mu.mol). The resulting mixture was
stirred at rt for 3 h and then directly purified by preparative
RP-HPLC (Reprosil C18, gradient 30-50% acetonitrile/0.2% aq.
trifluoroacetic acid). The product fractions were combined and
evaporated to dryness. The residue was dissolved in methanol and
filtered through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 84 mg (69% of th.) of the
title compound.
[1035] LC-MS (method 4): R.sub.t=0.75 min; MS (ESIpos): m/z=481
(M+H).sup.+
[1036] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.00 (s, 1H),
7.75 (s, 1H), 7.67-8.11 (br. s, 1H), 7.35 (s, 1H), 7.31 (s, 1H),
6.85 (s, 1H), 5.5-6.0 (br. s, 1H), 4.44 (q, 2H), 4.14-4.21 (m, 1H),
4.02-4.11 (m, 1H), 3.38-3.45 (q, 2H), 3.16-3.24 (m, 1H), 3.05-3.16
(m, 2H), 2.45 (s, 3H), 2.31-2.40 (m, 1H), 1.67-1.79 (m, 1H), 1.08
(t, 3H) ppm.
General Procedure for the Preparation of Examples 94-105 in Table
I:
[1037] A 0.13 M suspension of Intermediate 17A in THF was treated
with 1.5 eq. of the respective amine component, 3 eq. of sodium
triacetoxyborohydride and 1.5 eq. of acetic acid. The resulting
mixture was stirred at 60.degree. C. for 3-20 h. After this,
purification was carried out according to the methods
indicated.
TABLE-US-00001 TABLE I Example Purification No. Structure method(s)
LC-MS data 94 ##STR00208## P1, P5 Method 4: R.sub.t = 1.22 min; MS
(ESIpos): m/z = 468 (M + H).sup.+ 95 ##STR00209## P6, P3 Method 2:
R.sub.t = 0.92 min; MS (ESIpos): m/z = 509 (M + H).sup.+ 96
##STR00210## P1, P5 Method 4: R.sub.t = 0.83 min; MS (ESIpos): m/z
= 452 (M + H).sup.+ 97 ##STR00211## P1, P5 Method 5: R.sub.t = 1.84
min; MS (ESIpos): m/z = 454 (M + H).sup.+ 98 ##STR00212## P1, P5
Method 5: R.sub.t = 1.82 min; MS (ESIpos): m/z = 482 (M + H).sup.+
99 ##STR00213## P1, P5 Method 5: R.sub.t = 2.21 min; MS (ESIpos):
m/z = 516 (M + H).sup.+ 100 ##STR00214## P1, P5 Method 2: R.sub.t =
0.86 min; MS (ESIpos): m/z = 507 (M + H).sup.+ 101 ##STR00215## P1,
P5 Method 2: R.sub.t = 0.81 min; MS (ESIpos): m/z = 468 (M +
H).sup.+ 102 ##STR00216## P1, P5, P4 Method 2: R.sub.t = 1.03 min;
MS (ESIpos): m/z = 474 (M + H).sup.+ 103 ##STR00217## P1, P5, P2
Method 2: R.sub.t = 1.05 min; MS (ESIpos): m/z = 460 (M + H).sup.+
104 ##STR00218## P1, P6 Method 2: R.sub.t = 0.91 min; MS (ESIpos):
m/z = 467 (M + H).sup.+ 105 ##STR00219## P1, P6 Method 2: R.sub.t =
0.89 min; MS (ESIpos): m/z = 495 (M + H).sup.+
Example 106
4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)-N-[(3-
R)-2-oxopyrrolidin-3-yl]pyrrolo[2,1-f][1,2,4]triazine-7-carboxamide
##STR00220##
[1039] A stirred solution of Intermediate 64A (50 mg, 121 .mu.mol)
in DMF (2 ml) was treated at rt with
N-[(1H-benzotriazol-1-yloxy)(dimethylamino)methylene]-N-methylmethanamini-
um tetrafluoroborate (TBTU) (43 mg, 133 .mu.mol) and DIPEA (53
.mu.l, 303 .mu.mol). After 15 min, (3R)-3-aminopyrrolidin-2-one (24
mg, 242 .mu.mol) was added, and the resulting mixture was stirred
at rt for further 2 h. After this, the mixture was separated by
preparative RP-HPLC (Reprosil C18, gradient 30-50%
acetonitrile/0.2% aq. trifluoroacetic acid). The product fractions
were combined and evaporated to dryness. The residue was dissolved
in methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated affording 36
mg (60% of th.) of the title compound.
[1040] LC-MS (method 2): R.sub.t=0.91 min; MS (ESIpos): m/z=495
(M+H).sup.+
[1041] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=9.50 (d, 1H),
8.21-8.45 (br. s, 1H), 8.17 (s, 1H), 7.97 (s, 1H), 7.41 (s, 1H),
7.33 (s, 1H), 6.86 (s, 1H), 5.84-6.09 (br. s, 1H), 4.74 (dd, 2H),
4.47-4.56 (m, 1H), 3.96 (s, 3H), 3.37 (q, 2H), 3.22-3.29 (m, 2H),
2.54-2.61 (m, 1H), 2.46 (s, 3H), 1.90-2.03 (m, 1H), 1.00 (t, 3H)
ppm.
Example 107
4-{[4-Amino-6-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-7-yl]carbonyl}piperazin-2-one
##STR00221##
[1043] A stirred solution of Intermediate 64A (50 mg, 121 .mu.mol)
in DMF (2 ml) was treated at rt with TBTU (43 mg, 133 .mu.mol) and
DIPEA (53 .mu.l, 303 .mu.mol). After 15 min, piperazin-2-one (24
mg, 242 .mu.mol) was added, and the resulting mixture was stirred
at rt for further 16 h. After this, the mixture was separated by
preparative RP-HPLC (Reprosil C18, gradient 40-60%
acetonitrile/0.2% aq. trifluoroacetic acid). The product fractions
were combined and evaporated to dryness. The residue was dissolved
in methanol and filtered through an anion exchange cartridge
(Stratospheres SPE, PL-HCO.sub.3 MP-resin). The cartridge was
eluted with methanol, and the filtrate was evaporated affording 45
mg (68% of th.) of the title compound.
[1044] LC-MS (method 2): R.sub.t=0.92 min; MS (ESIpos): m/z=495
(M+H).sup.+
[1045] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=8.14 (br. s,
1H), 7.99-8.01 (m, 1H), 7.9-8.3 (br. s, 1H), 7.41 (d, 1H), 7.32 (s,
1H), 6.86 (s, 1H), 5.78-6.05 (br. s, 1H), 4.30-4.49 (m, 2H),
4.13-4.23 (m, 1H), 4.03-4.10 (m, 1H), 3.96 (s, 3H), 3.76-3.87 (m,
1H), 3.57-3.74 (m, 1H), 3.35-3.43 (m, 2H), 3.14-3.25 (m, 1H), 2.45
(s, 3H), 0.96-1.05 (m, 3H) ppm.
General Procedure for the Preparation of Examples 108-123 in Table
II:
[1046] A 0.13 M solution of Intermediate 64A in DMF was treated
with 1.1 eq. TBTU and 2.5 eq. DIPEA and stirred at rt for 15 min. 2
eq. of the respective amine were added, and the resulting mixture
was stirred at rt for further 18 h. After this, purification was
carried out according to the methods indicated.
TABLE-US-00002 TABLE II Example Purification No. Structure
method(s) LC-MS data 108 ##STR00222## P1 Method 2: R.sub.t = 1.02
min; MS (ESIpos): m/z = 456 (M + H).sup.+ 109 ##STR00223## P1
Method 4: R.sub.t = 0.97 min; MS (ESIpos): m/z = 412 (M + H).sup.+
110 ##STR00224## P1, P5 Method 2: R.sub.t = 1.01 min; MS (ESIpos):
m/z = 426 (M + H).sup.+ 111 ##STR00225## P1, P5 Method 4: R.sub.t =
1.11 min; MS (ESIpos): m/z = 440 (M + H).sup.+ 112 ##STR00226## P1
Method 4: R.sub.t = 1.07 min; MS (ESIpos): m/z = 482 (M + H).sup.+
113 ##STR00227## P1, P5 Method 2: R.sub.t = 0.84 min; MS (ESIpos):
m/z = 523 (M + H).sup.+ 114 ##STR00228## P1, P5 Method 2: R.sub.t =
0.96 min; MS (ESIpos): m/z = 440 (M + H).sup.+ 115 ##STR00229## P1,
P5 Method 2: R.sub.t = 1.09 min; MS (ESIpos): m/z = 452 (M +
H).sup.+ 116 ##STR00230## P1, P5 Method 4: R.sub.t = 0.95 min; MS
(ESIpos): m/z = 495 (M + H).sup.+ 117 ##STR00231## P1, P5 Method 4:
R.sub.t = 0.93 min; MS (ESIpos): m/z = 481 (M + H).sup.+ 118
##STR00232## P1, P5 Method 2: R.sub.t = 0.93 min; MS (ESIpos): m/z
= 482 (M + H).sup.+ 119 ##STR00233## P1, P5 Method 2: R.sub.t =
1.04 min; MS (ESIpos): m/z = 516 (M + H).sup.+ 120 ##STR00234## P1,
P5 Method 4: R.sub.t = 0.88 min; MS (ESIpos): m/z = 468 (M +
H).sup.+ 121 ##STR00235## P1, P5 Method 4: R.sub.t = 0.91 min; MS
(ESIpos): m/z = 496 (M + H).sup.+ 122 ##STR00236## P1, P5 Method 4:
R.sub.t = 1.09 min; MS (ESIpos): m/z = 488 (M + H).sup.+ 123
##STR00237## P1, P5 Method 4: R.sub.t = 0.98 min; MS (ESIpos): m/z
= 530 (M + H).sup.+
Example 124
4-{[4-Amino-5-(5,7-dimethoxy-1-benzothiophen-2-yl)-6-(methoxymethyl)pyrrol-
o[2,1-f][1,2,4]-triazin-7-yl]methyl}piperazin-2-one
##STR00238##
[1048] To a solution of Intermediate 62A (100 mg, 271 .mu.mol),
Intermediate 66A (77 mg, 325 .mu.mol) and caesium fluoride (206 mg,
1.35 mmol) in degassed THF/water (10:1; 5 ml) under argon was added
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 42.6 mg, 54 .mu.mol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010)).
The resulting mixture was degassed again and stirred under argon at
60.degree. C. for 6 h. Then, another portion of Intermediate 66A
(39 mg, 162 .mu.mol) was added, and stirring at 60.degree. C. was
continued for 10 h. After this, the reaction mixture was separated
by preparative RP-HPLC (Reprosil C18, gradient 30-50%
acetonitrile/0.1% aq. TFA). The product fractions were combined and
evaporated to dryness. The residue was dissolved in methanol and
filtered through an anion exchange cartridge (Stratospheres SPE,
PL-HCO.sub.3 MP-resin). The cartridge was eluted with methanol, and
the filtrate was evaporated affording 40 mg (28% of th.) of the
title compound.
[1049] LC-MS (method 2): R.sub.t=0.78 min; MS (ESIpos): m/z=483
(M+H).sup.+
[1050] .sup.1H-NMR (500 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.68 (br. s, 1H), 7.49-8.17 (br. s, 1H), 7.37 (s, 1H), 7.05 (d,
1H), 6.63 (d, 1H), 5.5-6.0 (br. s, 1H), 4.42 (s, 2H), 3.97 (s, 2H),
3.94 (s, 3H), 3.84 (s, 3H), 3.20 (s, 3H), 3.09-3.14 (m, 2H), 3.01
(s, 2H), 2.62-2.67 (m, 2H) ppm.
Example 125
4-{[4-Amino-7-(hydroxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)p-
yrrolo[2,1-f][1,2,4]-triazin-6-yl]methyl}piperazin-2-one
##STR00239##
[1052] To a suspension of Intermediate 70A (700 mg, 1.49 mmol),
Intermediate 5A (497 mg, 2.24 mmol) and caesium fluoride (1.36 g,
8.95 mmol) in degassed THF/water (2:1, 90 ml) under argon was added
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 117 mg, 0.149 mmol; see S. L.
Buchwald et al., J. Am. Chem. Soc. 132 (40), 14073-14075 (2010)).
The resulting mixture was degassed again and stirred under argon at
60.degree. C. overnight. Another portion of Intermediate 5A (231
mg, 1.04 mmol) and
(2'-aminobiphenyl-2-yl)(chloro)palladium-dicyclohexyl(2',4',6'-triisoprop-
ylbiphenyl-2-yl)phosphine (1:1; 117 mg, 0.149 mmol) were added, and
stirring was continued at 60.degree. C. for 3 h. The precipitate
was filtered off, washed with THF and dried in vacuo. The solid was
suspended in mixture of DMF and 1 M aq. trifluoroacetic acid and
filtered. The filtrate was separated by preparative RP-HPLC
(Reprosil C18, gradient 20-40% acetonitrile/0.2% aq.
trifluoroacetic acid). The product fractions were combined and then
alkalized by addition of sat. aq. sodium bicarbonate solution. The
solution was extracted with ethyl acetate, and the organic layer
was washed with brine, dried over magnesium sulfate and evaporated
to yield 88 mg of the title compound (13% of th.).
[1053] LC-MS (method 2): R.sub.t=0.70 min; MS (ESIpos): m/z=453
(M+H).sup.+
[1054] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): inter al.
.delta.=7.6-8.0 (br. s, 1H), 7.98 (s, 1H), 7.71 (s, 1H), 7.37 (s,
1H), 7.31 (s, 1H), 6.84 (s, 1H), 5.4-5.8 (br. s, 1H), 5.19 (t, 1H),
4.85 (d, 2H), 3.95 (s, 3H), 3.58 (s, 2H), 3.04 (br. m, 2H), 2.88
(s, 2H), 2.45 (s, 3H) ppm.
Example 126
4-{[4-Amino-7-(methoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)p-
yrrolo[2,1-f]-[1,2,4]triazin-6-yl]methyl}piperazin-2-one
##STR00240##
[1056] A suspension of Example 125 (88 mg, 194 .mu.mol) in
dichloromethane (5 ml) was treated with thionyl chloride (29 .mu.l,
389 .mu.mol), and the mixture was stirred at rt for 65 min. Another
portion of thionyl chloride (29 .mu.l, 389 .mu.mol) was added, and
stirring was continued for 1.5 h. An excess of methanol was then
added, followed by dropwise addition of a 5.4 M solution of sodium
methylate in methanol (84 mg, 1.56 mmol) until pH 8 was reached.
After stirring for 3 days, the volatiles were evaporated under
reduced pressure, and the residue was separated by preparative
RP-HPLC (Reprosil C18, gradient 20-40% acetonitrile/0.2% aq.
trifluoroacetic acid). The product fractions were evaporated to
yield 19 mg of the title compound (21% of th.).
[1057] LC-MS (method 2): R.sub.t=0.77 min; MS (ESIpos): m/z=466
(M+H).sup.+
[1058] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.75-8.11 (br. s, 1H), 7.68 (s, 1H), 7.38 (s, 1H), 7.31 (s, 1H),
6.84 (s, 1H), 5.51-5.82 (m, 1H), 4.77 (s, 2H), 3.95 (s, 3H), 3.55
(s, 2H), 3.31 (s, 3H), 3.04 (br. s, 2H), 2.85 (s, 2H), 2.42-2.48
(m, 5H) ppm.
Example 127
4-{[4-Amino-7-(ethoxymethyl)-5-(7-methoxy-5-methyl-1-benzothiophen-2-yl)py-
rrolo[2,1-f][1,2,4]-triazin-6-yl]methyl}piperazin-2-one
##STR00241##
[1060] A suspension of Example 125 (100 mg, 221 .mu.mol) in
dichloromethane (10 ml) was treated with thionyl chloride (161
.mu.l, 2.21 mmol), and the mixture was stirred at rt for 30 min.
Ethanol was added, and the volatiles were evaporated under reduced
pressure. The residue was dissolved in ethanol (10 ml), sodium
ethylate (30 mg, 442 .mu.mol) was added, and the mixture was
stirred at rt for 1 h. The mixture was then directly separated by
preparative RP-HPLC (Reprosil C18, gradient 20-40%
acetonitrile/0.2% aq. trifluoroacetic acid). The product fractions
were combined and alkalized by addition of sat. aq. sodium
bicarbonate solution. The solution was extracted with ethyl
acetate, and the organic layer was washed with brine, dried over
magnesium sulfate and evaporated to yield 57 mg of the title
compound (54% of th.).
[1061] LC-MS (method 2): R.sub.t=0.85 min; MS (ESIpos): m/z=480
(M+H).sup.+
[1062] .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta.=7.99 (s, 1H),
7.75-8.09 (br. s, 1H), 7.64-7.71 (m, 1H), 7.38 (s, 1H), 7.31 (s,
1H), 6.84 (s, 1H), 5.38-5.91 (br. s, 1H), 4.81 (s, 2H), 3.95 (s,
3H), 3.49-3.59 (m, 4H), 2.99-3.08 (m, 2H), 2.86 (s, 2H), 1.12 (t,
3H) ppm.
B. EVALUATION OF BIOLOGICAL ACTIVITY
Abbreviations and Acronyms
[1063] Ahx 6-aminohexanoic acid ATP adenosine triphosphate BSA
bovine serum albumin CREB cAMP-response element-binding protein
DMSO dimethylsulfoxide EDTA ethylenediaminetetraacetic acid EGTA
ethyleneglycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic acid
FBS fetal bovine serum FGF fibroblast growth factor FGFR fibroblast
growth factor receptor GFP green fluorescent protein GST
glutathione S-transferase HEPES
4-(2-hydroxyethyl)piperazine-1-ethansulfonic acid HRTF homogeneous
time-resolved fluorescence MOPS 3-(N-morpholino)propanesulfonic
acid mTOR mammalian target of Rapamycin PBS phosphate buffered
saline PI3K phosphatidylinositol 3-kinase RTK receptor tyrosine
kinase SNP single nucleotide polymorphism TR-FRET time-resolved
fluorescence resonance energy transfer VEGF vascular endothelial
growth factor VEGFR vascular endothelial growth factor receptor
[1064] Demonstration of the activity of the compounds of the
present invention may be accomplished through in vitro, ex vivo,
and in vivo assays that are well known in the art. For example, to
demonstrate the activity of the compounds of the present invention,
the following assays may be used.
B-1. FGFR-1 High ATP Kinase Assay
[1065] FGFR-1 inhibitory activity at high ATP concentration of the
compounds of the present invention after their pre-incubation with
FGFR-1 was quantified employing the TR-FRET based FGFR-1 high ATP
assay as described in the following paragraphs:
[1066] A recombinant tagged FGFR-1 fusion protein [fusion of
glutathione-S-transferase (GST) (N-terminally), His6-tag, thrombin
cleavage site, and the intracellular part of human FGFR-1 from
amino acids G400 to R800 as in GenBank entry NM_015850], expressed
in SF9 insect cells using baculovirus expression system and
purified via glutathione-agarose affinity chromatography, was
purchased from Proqinase (product no. 0101-0000-1) and used as
enzyme. As substrate for the kinase reaction, the biotinylated
peptide biotin-Ahx-AAEEEYFFLFAKKK (C-terminus in amide form) was
used which can be purchased, e.g., from Biosyntan (Berlin-Buch,
Germany).
[1067] Usually, test compounds were tested on the same microtiter
plate at 11 different concentrations in the range of 20 .mu.M to
0.1 nM (e.g. 20 .mu.M, 5.9 .mu.M, 1.7 .mu.M, 0.51 .mu.M, 0.15
.mu.M, 44 nM, 13 nM, 3.8 nM, 1.1 nM, 0.33 nM, and 0.1 nM) in
duplicates for each concentration. The dilution series was prepared
separately prior to the assay as 100-fold concentrated stock
solutions in DMSO; exact concentrations could vary depending on the
pipettor used. For the assay, 50 nl of each stock solution of the
test compound in DMSO was pipetted into a black, low-volume
384-well microtiter plate (Greiner Bio-One, Frickenhausen,
Germany). 2 .mu.l of a solution of the above FGFR-1 fusion protein
in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate,
1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA),
0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was
incubated for 15 min at 22.degree. C. to allow pre-binding of the
test compound to the enzyme. Then, the kinase reaction was started
by the addition of 3 .mu.l of a solution of adenosine triphosphate
(ATP, 3.3 mM; final concentration in the 5 .mu.l assay volume=2 mM)
and substrate (0.16 .mu.M; final concentration in the 5 .mu.l assay
volume=0.1 .mu.M) in assay buffer, and the resulting mixture was
incubated for a reaction time of 15 min at 22.degree. C. The
concentration of FGFR-1 fusion protein was adjusted depending on
the activity of the enzyme lot and was chosen appropriately to have
the assay in the linear range (typical concentrations were in the
range of 0.05 .mu.g/ml). The reaction was stopped by the addition
of 5 .mu.l of a solution of HTRF detection reagents [25 nM
streptavidin-XL665 (Cis Biointernational) and 1 nM PT66-Eu-chelate,
an europium-chelate labelled anti-phosphotyrosine antibody
(Perkin-Elmer; PT66-Tb-cryptate from Cis Biointernational may be
used instead), in an aqueous EDTA solution (50 mM EDTA, 0.1% (w/v)
BSA in 50 mM HEPES/NaOH pH 7.5)].
[1068] The resulting mixture was incubated for 1 h at 22.degree. C.
to allow formation of the complex between the phosphorylated
biofinylated peptide and the detection reagents. Subsequently, the
amount of phosphorylated substrate was evaluated by measurement of
the resonance energy transfer from the Eu-chelate to the
streptavidin-XL665. For this, the fluorescence emissions at 620 nm
and 665 nm after excitation at 350 nm were measured in a TR-FRET
reader [e.g. Rubystar (BMG Labtechnologies, Offenburg, Germany) or
Viewlux (Perkin-Elmer)]. The ratio of the emissions at 665 nm and
at 620 nm was taken as the measure for the amount of phosphorylated
substrate. Data were nornmalised (enzyme reaction without
inhibitor=0% inhibition, all other assay components but no
enzyme=100% inhibition), and IC.sub.50 values were calculated by a
4-parameter fit using an in-house software.
[1069] IC.sub.50 values for individual compounds of the invention
from this assay are listed in Table 1A below:
TABLE-US-00003 TABLE 1A FGFR-1 Example (high ATP) No. IC.sub.50
[nM] 1 12.2 2 7.2 3 6.6 4 13.7 5 4.6 6 9.0 7 4.2 8 9.4 9 31.0 10
12.7 11 6.5 12 3.3 13 2.5 14 1.4 15 1.0 16 12.5 17 0.9 18 0.7 19
0.5 20 25.0 21 4.2 22 0.9 23 17.9 24 1.3 25 0.7 26 0.9 27 1.3 28
2.1 29 1.6 30 2.8 31 1.5 32 5.2 33 1.0 34 6.0 35 61.3 36 3.0 37
22.1 38 6.9 39 1.1 40 2.1 41 2.8 42 5.1 43 8.6 44 2.1 45 3.4 46 3.2
47 1.7 48 12.8 51 4.4 52 9.7 53 3.9 54 11.2 56 2.4 57 3.2 58 6.8 59
3.8 60 16.2 61 0.5 62 3.1 64 1.0 65 5.6 66 1.6 67 5.0 68 27.0 69
55.3 70 39.1 71 2.7 72 1.4 73 4.4 74 4.0 75 4.1 76 2.8 77 1.4 78
1.7 79 12.6 80 1.3 81 1.9 82 3.4 83 14.5 84 3.7 85 26.4 86 3.9 87
14.1 88 79.6 89 82.9 90 9.9 91 52.9 92 8.2 93 5.5 94 81.9 95 17.0
96 54.5 97 16.4 98 17.0 99 24.8 100 54.5 101 30.8 102 39.2 103 50.3
104 5.6 105 8.7 106 6.4 107 9.4 108 3.9 109 8.4 110 17.5 111 27.0
112 42.1 113 52.7 114 11.2 115 24.9 116 3.7 117 8.8 118 6.8 119 9.0
120 6.8 121 8.2 122 18.9 123 25.1 124 27.3 125 5.0 126 3.9 127
8.0
[1070] Selected
8-amino-1-(benzothiophen-2-yl)imidazo[1,5-a]pyrazine derivatives
and related compounds which were regarded to be representative of
closest prior art (see Int. Pat. Appl. WO 2007/061737-A2 and
example compounds described therein) were synthesized following the
published procedures and also tested in the FGFR-1 high ATP assay
for comparative purposes. IC.sub.50 values that were obtained for
these compounds are listed in Table 1B below:
TABLE-US-00004 TABLE 1B Example No. FGFR-1 in WO (high ATP)
Structure of comparative compound 2007/061737 IC.sub.50 [nM]
##STR00242## 4 12000 ##STR00243## 5 500 ##STR00244## 25 880
##STR00245## 120 985 ##STR00246## 205 20000 ##STR00247## 210 456
##STR00248## 233 4600
[1071] The IC.sub.50 values specified in Table 1A and 1B
demonstrate that the compounds of the present invention are about
five to a thousand times more potent in inhibiting FGFR-1 kinase
activity than the selected prior art compounds.
B-2. FGFR-3 Kinase Assay
[1072] FGFR-3 inhibitory activity of the compounds of the present
invention after their pre-incubation with FGFR-3 was quantified
employing the TR-FRET based FGFR-3 assay as described in the
following paragraphs:
[1073] A recombinant tagged FGFR-3 fusion protein [fusion of
glutathione-S-transferase (GST) (N-terminally), His6-tag, thrombin
cleavage site, and the intracellular part of human FGFR-3 from
amino acids R397 to T806 as in NCBI/Protein entry NP_000133.1],
expressed in SF9 insect cells using baculovirus expression system
and purified via glutathione-S-transferase affinity chromatography,
was purchased from Proqinase (product no. 1068-0000-1) and used as
enzyme. As substrate for the kinase reaction, the biotinylated
peptide biotin-Ahx-AAEEEYFFLFAKKK (C-terminus in amide form) was
used which can be purchased, e.g., from Biosyntan (Berlin-Buch,
Germany).
[1074] Usually, test compounds were tested on the same microtiter
plate at 11 different concentrations in the range of 20 .mu.M to
0.1 nM (e.g. 20 .mu.M, 5.9 .mu.M, 1.7 .mu.M, 0.51 .mu.M, 0.15
.mu.M, 44 nM, 13 nM, 3.8 nM, 1.1 nM, 0.33 nM, and 0.1 nM) in
duplicates for each concentration. The dilution series was prepared
separately prior to the assay as 100-fold concentrated stock
solutions in DMSO; exact concentrations could vary depending on the
pipettor used. For the assay, 50 nl of each stock solution of the
test compound in DMSO was pipetted into a black, low-volume
384-well microtiter plate (Greiner Bio-One, Frickenhausen,
Germany). 2 .mu.l of a solution of the above FGFR-3 fusion protein
in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate,
1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA),
0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was
incubated for 15 min at 22.degree. C. to allow pre-binding of the
test compound to the enzyme. Then, the kinase reaction was started
by the addition of 3 .mu.l of a solution of adenosine triphosphate
(ATP, 16.7 .mu.M; final concentration in the 5 .mu.l assay
volume=10 .mu.M) and substrate (0.8 .mu.M; final concentration in
the 5 .mu.l assay volume=0.5 .mu.M) in assay buffer, and the
resulting mixture was incubated for a reaction time of 60 min at
22.degree. C. The concentration of FGFR-3 fusion protein was
adjusted depending on the activity of the enzyme lot and was chosen
appropriately to have the assay in the linear range (typical
concentrations were in the range of 0.03 .mu.g/ml). The reaction
was stopped by the addition of 5 .mu.l of a solution of HTRF
detection reagents [100 nM streptavidin-XL665 (Cis
Biointernational) and 1 nM PT66-Tb-cryptate, a terbium-cryptate
labelled anti-phosphotyrosine antibody (Cis Biointernational;
PT66-Eu-chelate from Perkin-Elmer may be used instead), in an
aqueous EDTA solution (50 mM EDTA, 0.1% (w/v) BSA in 50 mM
HEPES/NaOH pH 7.5)].
[1075] The resulting mixture was incubated for 1 h at 22.degree. C.
to allow formation of the complex between the phosphorylated
biotinylated peptide and the detection reagents. Subsequently, the
amount of phosphorylated substrate was evaluated by measurement of
the resonance energy transfer from the Tb-chelate to the
streptavidin-XL665. For this, the fluorescence emissions at 620 nm
and 665 nm after excitation at 350 nm were measured in a TR-FRET
reader [e.g. Rubystar (BMG Labtechnologies, Offenburg, Germany) or
Viewlux (Perkin-Elmer)]. The ratio of the emissions at 665 nm and
at 620 nm was taken as the measure for the amount of phosphorylated
substrate. Data were normalised (enzyme reaction without
inhibitor=0% inhibition, all other assay components but no
enzyme=100% inhibition), and IC.sub.50 values were calculated by a
4-parameter fit using an in-house software.
[1076] IC.sub.50 values for individual compounds of the invention
from this assay are listed in Table 2A below:
TABLE-US-00005 TABLE 2A Example FGFR-3 No. IC.sub.50 [nM] 1 24.8 2
29.4 3 46.6 4 24.8 5 12.1 6 28.5 7 10.4 8 17.7 9 63.9 10 8.3 11
13.0 12 14.0 13 10.0 14 2.5 15 1.0 16 9.4 17 3.3 18 2.8 19 2.7 20
26.1 21 13.0 22 4.7 23 41.9 24 5.7 25 5.2 26 4.9 27 12.6 28 9.7 29
2.5 30 6.4 31 13.3 32 52.3 33 7.2 34 10.5 35 65.4 36 9.8 37 34.6 38
18.8 39 4.9 40 7.9 41 7.2 42 18.4 43 32.4 44 6.1 45 9.7 46 8.1 47
3.4 48 45.4 51 3.9 52 7.0 53 2.5 54 6.8 56 2.0 57 1.8 58 9.8 59 6.8
60 77.9 61 0.9 62 11.0 64 2.9 65 3.1 66 3.7 67 9.4 68 42.7 69 58.1
71 5.4 72 0.7 73 12.2 74 1.4 75 19.8 76 1.4 77 3.8 78 2.0 79 17.9
80 0.2 81 0.5 82 16.0 83 2.1 84 16.5 85 26.0 86 6.8 87 28.1 90 11.0
91 31.1 93 8.7 95 28.7 98 26.5 99 36.9 102 39.8 106 7.1 107 17.4
108 7.7 114 9.6 117 5.3 119 4.5 121 24.6
[1077] Selected
8-amino-1-(benzothiophen-2-yl)imidazo[1,5-a]pyrazine derivatives
and related compounds which were regarded to be representative of
closest prior art (see Int. Pat. Appl. WO 2007/061737-A2 and
example compounds described therein) were synthesized following the
published procedures and also tested in the FGFR-3 assay for
comparative purposes. IC.sub.50 values that were obtained for these
compounds are listed in Table 2B below:
TABLE-US-00006 TABLE 2B Example No. in WO FGFR-3 Structure of
comparative compound 2007/061737 IC.sub.50 [nM] ##STR00249## 4 2400
##STR00250## 5 250 ##STR00251## 25 1200 ##STR00252## 120 506
##STR00253## 205 20000 ##STR00254## 210 554 ##STR00255## 233
10000
[1078] The IC.sub.50 values specified in Table 2A and 2B
demonstrate that the compounds of the present invention are about
three to a thousand times more potent in inhibiting FGFR-3 kinase
activity than the selected prior art compounds.
B-3. FGFR-4 High ATP Kinase Assay
[1079] FGFR-4 inhibitory activity at high ATP concentration of the
compounds of the present invention after their pre-incubation with
FGFR-4 was quantified employing the TR-FRET based FGFR-4 high ATP
assay as described in the following paragraphs:
[1080] A recombinant tagged FGFR-4 fusion protein [fusion of
glutathione-S-transferase (GST) (N-terminally), His6-tag, thrombin
cleavage site, and the intracellular part of human FGFR-4 from
amino acids R391 to T802 as in GenBank entry NM_002011], expressed
in SF9 insect cells using baculovirus expression system and
purified via glutathione-agarose affinity chromatography, was
purchased from Proqinase (product no. 0127-0000-3) and used as
enzyme. As substrate for the kinase reaction, the biotinylated
peptide biotin-Ahx-AAEEEYFFLFAKKK (C-terminus in amide form) was
used which can be purchased, e.g., from Biosyntan (Berlin-Buch,
Germany).
[1081] Usually, test compounds were tested on the same microtiter
plate at 11 different concentrations in the range of 20 .mu.M to
0.1 nM (e.g. 20 .mu.M, 5.9 .mu.M, 1.7 .mu.M, 0.51 .mu.M, 0.15
.mu.M, 44 nM, 13 nM, 3.8 nM, 1.1 nM, 0.33 nM, and 0.1 nM) in
duplicates for each concentration. The dilution series was prepared
separately prior to the assay as 100-fold concentrated stock
solutions in DMSO; exact concentrations could vary depending on the
pipettor used. For the assay, 50 nl of each stock solution of the
test compound in DMSO was pipetted into a black, low-volume
384-well microtiter plate (Greiner Bio-One, Frickenhausen,
Germany). 2 .mu.l of a solution of the above FGFR-4 fusion protein
in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate,
1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA),
0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was
incubated for 15 min at 22.degree. C. to allow pre-binding of the
test compound to the enzyme. Then, the kinase reaction was started
by the addition of 3 .mu.l of a solution of adenosine triphosphate
(ATP, 3.3 mM; final concentration in the 5 .mu.l assay volume=2 mM)
and substrate (0.8 .mu.M; final concentration in the 5 .mu.l assay
volume=0.5 .mu.M) in assay buffer, and the resulting mixture was
incubated for a reaction time of 60 min at 22.degree. C. The
concentration of FGFR-4 fusion protein was adjusted depending on
the activity of the enzyme lot and was chosen appropriately to have
the assay in the linear range (typical concentrations were in the
range of 0.03 .mu.g/ml). The reaction was stopped by the addition
of 5 .mu.l of a solution of HTRF detection reagents [100 nM
streptavidin-XL665 (Cis Biointernational) and 1 nM
PT66-Tb-cryptate, a terbium-cryptate labelled anti-phosphotyrosine
antibody (Cis Biointernational; PT66-Eu-chelate from Perkin-Elmer
may be used instead), in an aqueous EDTA solution (50 mM EDTA, 0.1%
(w/v) BSA in 50 mM HEPES/NaOH pH 7.5)].
[1082] The resulting mixture was incubated for 1 h at 22.degree. C.
to allow formation of the complex between the phosphorylated
biotinylated peptide and the detection reagents. Subsequently, the
amount of phosphorylated substrate was evaluated by measurement of
the resonance energy transfer from the Tb-chelate to the
streptavidin-XL665. For this, the fluorescence emissions at 620 nm
and 665 nm after excitation at 350 nm were measured in a TR-FRET
reader [e.g. Rubystar (BMG Labtechnologies, Offenburg, Germany) or
Viewlux (Perkin-Elmer)]. The ratio of the emissions at 665 nm and
at 620 nm was taken as the measure for the amount of phosphorylated
substrate. Data were normalised (enzyme reaction without
inhibitor=0% inhibition, all other assay components but no
enzyme=100% inhibition), and IC.sub.50 values were calculated by a
4-parameter fit using an in-house software.
B-4. mTOR Kinase Assay (for Comparative Purposes)
[1083] mTOR inhibitory activity of the compounds of the present
invention was quantified employing the TR-FRET based mTOR assay as
described in the following paragraphs:
[1084] Recombinant fusion tagged mTOR protein
[glutathione-S-transferase (GST) fused to human mTOR amino acids
from 1360 to 2549], expressed in insect cells and purified by
glutathione-sepharose affinity chromatography, was purchased from
Invitrogen (Cat.-No. 4753) and used as enzyme. As substrate for the
kinase reaction, a recombinant fusion protein of GFP and 4E-BP1
(purchased from Invitrogen, Cat.-No. PV4759) was used.
[1085] Test compounds were dissolved in DMSO to generate 10 mM
stock solutions. These solutions were first 10-fold diluted by 100%
DMSO to get 1 mM solutions in 100% DMSO, then 100-fold diluted by
50% DMSO to get 10 .mu.M solutions in 50% DMSO.
[1086] For the assay, 0.5 .mu.l of a 10 .mu.M solution of the test
compound in 50% DMSO was pipetted into a black, low-volume 384-well
microtiter plate (Greiner Bio-One, Frickenhausen, Germany). 2 .mu.l
of a solution of the above mTOR fusion protein in aqueous assay
buffer [50 mM HEPES/NaOH pH 7.5, 5 mM magnesium chloride, 1.0 mM
dithiothreitol, 1 mM EGTA, 0.01% (v/v) Triton-X100, 0.01% (w/v)
bovine serum albumin (BSA)] was added, and the mixture was
incubated for 15 min at 22.degree. C. to allow pre-binding of the
test compound to the enzyme. Then, the kinase reaction was started
by the addition of 2.5 .mu.l of a solution of adenosine
triphosphate (ATP, 80 .mu.M; final concentration in the 5 .mu.l
assay volume=40 .mu.M) and substrate (0.6 .mu.M; final
concentration in the 5 .mu.l assay volume=0.3 .mu.M) in assay
buffer, and the resulting mixture was incubated for a reaction time
of 60 min at 22.degree. C. The concentration of mTOR fusion protein
was chosen appropriately to have the assay in the linear range (a
typical final concentration in the 5 .mu.l assay volume was 1.25
ng/l). The reaction was stopped by the addition of 5 .mu.l of 30 mM
EDTA (final concentration in the 10 .mu.l assay volume=15 mM) and 2
nM Tb-chelate labelled anti-4E-BP1 [pT46] phosphospecific antibody
[Invitrogen Cat.-No. PV4755] (final concentration in the 10 .mu.l
assay volume=1 nM) in FRET buffer.
[1087] The resulting mixture was incubated for 1 h at 22.degree. C.
to allow formation of the complex between the phosphorylated
substrate and the Tb-chelate labelled antibody. Subsequently, the
amount of phosphorylated substrate was evaluated by measurement of
the resonance energy transfer from the Tb-chelate to the GFP. For
this, the fluorescence emissions at 495 nm and 520 nm after
excitation at 340 nm was measured in an Envision 2104 multilabel
reader (Perkin-Elmer). The ratio of the emissions at 520 nm and at
495 nm was taken as the measure for the amount of phosphorylated
substrate. Data were normalised (enzyme reaction without
inhibitor=0% inhibition, all other assay components but no
enzyme=100% inhibition), and either mean values (if tested in
replicates at a single concentration) or IC.sub.50 values (by a
4-parameter fit using an in-house software) were calculated.
[1088] Mean inhibition values at 1 .mu.M for individual compounds
of the present invention are listed in Table 3 below:
TABLE-US-00007 TABLE 3 Example mTOR No. % inhibition @ 1 .mu.M 1
8.7 2 7.2 3 8.2 5 9.7 6 0.3 7 3.1 8 7.8 9 no inhib. effect detect.
10 6.0 11 33.2 12 25.8 13 14.9 14 15.7 16 36.9 17 6.7 18 no inhib.
effect detect. 19 17.6 20 31.3 21 5.0 22 17.6 23 11.6 24 6.3 25 3.5
26 6.8 27 5.9 28 no inhib. effect detect. 29 28.5 30 21.1 31 16.3
32 1.6 33 0.9 34 25.9 35 1.4 36 14.4 37 8.6 38 20.8 39 11.5 40 0.1
41 no inhib. effect detect. 42 no inhib. effect detect. 43 10.1 44
13.3 45 15.5 46 11.0 47 14.7 48 4.1 51 8.4 52 4.2 53 3.3 54 1.7 56
16.1 57 2.0 58 4.0 59 0.8 60 11.1 61 15.9 62 0.2 64 1.1 65 13.9 66
15.8 67 8.4 68 8.0 69 8.7 70 5.8 71 0.1 72 8.2 73 18.5 74 18.6 75
10.2 76 8.5 77 0.7 78 5.8 79 14.7 80 3.9 81 4.1 82 7.1 83 6.3 84
9.1 85 0.9 87 12.7 88 5.3 89 no inhib. effect detect. 93 12.5 98 no
inhib. effect detect. 99 5.3 103 no inhib. effect detect. 107 no
inhib. effect detect. 108 8.5 119 no inhib. effect detect. 121 no
inhib. effect detect. (no inhib. effect detect. = no inhibitory
effect detectable at 1 .mu.M).
[1089] The data in Table 3 show that the compounds of the present
invention only have a weak, if any, inhibitory effect on mTOR
kinase which is not considered to contribute to the pharmacological
activity observed with these compounds.
B-5. Inhibition of Growth Factor-Mediated Cell Proliferation
[1090] Human umbilical vein endothelial cells (HUVEC) were obtained
from Cellsystems (FC-0003) and grown in Vasculife VEGF complete
medium (Cellsystems, LL-1020) containing 2% fetal bovine serum
(FBS) at 37.degree. C. and 5% CO.sub.2. The cells were used for
proliferation assays up to passage 7.
[1091] The HUVEC cells were harvested using accutase (PAA, L11-007)
and seeded in columns 2 to 12 of 96-well plates (Falcon MICROTEST
tissue culture plate 96-well flat bottom, BD 353075, or
CLEAR-PLATE, black, 96-well, Greiner Bio-One, No. 655090) at a cell
density of 2500 cells/well in 100 .mu.l Vasculife VEGF complete
medium with column 1 remaining empty as blank. Cells were allowed
to incubate at 37.degree. C. and 5% CO.sub.2 for at least 6 h.
Then, the cells were washed once with PBS and starved overnight in
Vasculife basal medium (Cellsystems, LM-0002) containing heparin,
ascorbate and L-glutamine (components of the Vasculife Life Factors
Kit, Cellsystems, LL-1020) as well as 0.2% FBS.
[1092] After about 18 h, the starving medium was discarded, and the
cells were exposed for 72 h to 9 consecutive log or half-log
concentrations of test compound in the range of 10 .mu.M to 30
.mu.M and to 5, 10 or 20 ng/ml hFGF-2 (recombinant human FGF basic,
R&D Systems, 233-FB) in 100 .mu.l starving medium. 10 mM stock
solutions of test compounds in DMSO were diluted to 200.times.
final concentration in DMSO resulting in a final DMSO concentration
of 0.5% in all wells. Controls consisted of cells grown in starving
medium only and of cells grown in hFGF-2 containing starving medium
with 0.5% DMSO. To determine cell proliferation, 5 .mu.l Alamar
Blue solution (Biosource, DAL 1100) was added to each well (1:20
dilution), and the cells were allowed to incubate for further 4 h
at 37.degree. C. and 5% CO.sub.2 before measuring fluorescence (ex.
535 nm, em. 595 nm) with a Spectrafluor Plus Tecan plate reader
(XFLUOR4 version 4.20). In some experiments, an ATP Determination
Kit (BIAFFTN GmbH, LBR-T100) was used according to the
manufacturer's instructions. In each experiment, samples were
assayed in triplicate, and the standard deviations were determined.
GraphPad Prism 5 software was used to analyze the data and to
obtain IC.sub.50 values. All test compounds were assayed 2 to 10
times in independent experiments and similar results were
obtained.
[1093] The data listed in Table 4 below represent the IC.sub.50
values for representative compounds of the invention resulting from
the corresponding averaged pIC.sub.50 values:
TABLE-US-00008 TABLE 4 hFGF-2 mediated Example HUVEC proliferation,
No. IC.sub.50 [nM] 1 16.4 4 17.5 5 5.3 8 9.7 10 16.0 11 150.0 13
17.0 14 25.0 16 117.3 18 10.4 21 3.7 26 56.7 34 37.3 38 213.3 39
365.0 45 171.0 46 84.5 48 360.0 51 1.2 52 4.0 53 3.3 54 2.2 56 18.1
57 3.2 59 51.1 60 250.0 64 4.5 67 12.1 68 14.4 71 5.4 72 1.6 73 5.5
74 0.2 75 1.5 76 1.5 77 3.2 79 3.5 80 1.6 81 1.6 84 46.6 85
126.0
[1094] Most compounds of the present invention displayed about ten-
to hundred-fold reduced inhibitory activity in this proliferation
assay when vascular endothelial growth factor (VEGF-A.sub.165
isoform) was used as mediating growth factor (instead of FGF-2),
indicating a significant selectivity of these compounds for FGFR
versus VEGFR kinases.
B-6. Human Xenograft and Syngeneic Tumor Models
[1095] Different tumor models have been conducted in order to
profile compounds of the present invention in vivo. Human, rat or
mouse tumor cells were cultivated in vitro and implanted into
either immunodeficient or immunocompetent mice, or immunodeficient
rats. Treatment started after tumor establishment, and
tumor-bearing animals were treated with substances via different
routes (per os, intravenously, intraperitoneally or
subcutaneously). Substances were tested as mono-therapy or in
combination therapy with other pharmacological substances.
Treatment of the tumor-bearing animals was conducted until the
tumors reached an average size of 120 mm.sup.2. Tumors were
measured in two dimensions using a caliper, and tumor volume was
calculated according to the formula (length.times.width.sup.2)/2.
Substance efficacy was evaluated at the end of the experiment using
the T/C ratio [T=final tumor weight in the treated group; C=final
tumor weight in the control group]. Statistical significance of the
efficacy between control and treated groups was determined using
the ANOVA variance test. All animal studies were conducted
according to the German regulatory guidelines.
[1096] Although the invention has been disclosed with reference to
specific embodiments, it is apparent that other embodiments and
variations of the invention may be devised by others skilled in the
art without departing from the true spirit and scope of the
invention. The claims are intended to be construed to include all
such embodiments and equivalent variations.
C. EXAMPLES RELATING TO PHARMACEUTICAL COMPOSITIONS
[1097] Pharmaceutical compositions according to the present
invention can be illustrated as follows:
Sterile i.v. Solution:
[1098] A 5 mg/mL solution of the desired compound of the invention
can be made using sterile, injectable water, and the pH is adjusted
if necessary. The solution is diluted for administration to 1-2
mg/mL with sterile 5% dextrose and is administered as an i.v.
infusion over about 60 minutes.
Lyophilized Powder for i.v. Administration:
[1099] A sterile preparation can be prepared with (i) 100-1000 mg
of the desired compound of the invention as a lyophilized powder,
(ii) 32-327 mg/mL sodium citrate, and (iii) 300-3000 mg Dextran 40.
The formulation is reconstituted with sterile, injectable saline or
5% dextrose to a concentration of 10 to 20 mg/mL, which is further
diluted with saline or 5% dextrose to 0.2 to 0.4 mg/mL, and is
administered either as i.v. bolus or by i.v. infusion over 15-60
minutes.
Intramuscular Suspension:
[1100] The following solution or suspension can be prepared for
intramuscular injection:
[1101] 50 mg/mL of the desired, water-insoluble compound of the
invention; 5 mg/mL sodium carboxymethylcellulose; 4 mg/mL Tween 80;
9 mg/mL sodium chloride; 9 mg/mL benzyl alcohol.
Hard Shell Capsules:
[1102] A large number of unit capsules are prepared by filling
standard two-piece hard gelatin capsules each with 100 mg of the
desired, powdered compound of the invention, 150 mg of lactose, 50
mg of cellulose and 6 mg of magnesium stearate.
Soft Gelatin Capsules:
[1103] A mixture of the desired compound of the invention in a
digestible oil, such as soybean oil, cottonseed oil or olive oil,
is prepared and injected by means of a positive displacement pump
into molten gelatin to form soft gelatin capsules containing 100 mg
of the active ingredient. The capsules are washed and dried. The
desired compound of the invention can be dissolved in a mixture of
polyethylene glycol, glycerin and sorbitol to prepare a
water-miscible medicine mix.
Tablets: A large number of tablets are prepared by conventional
procedures so that the dosage unit is 100 mg of the desired
compound of the invention, 0.2 mg of colloidal silicon dioxide, 5
mg of magnesium stearate, 275 mg of microcrystalline cellulose, 11
mg of starch, and 98.8 mg of lactose. Appropriate aqueous and
non-aqueous coatings may be applied to increase palatability,
improve elegance and stability, or delay absorption.
Solution or Suspension for Topical Application to the Eye (Eye
Drops):
[1104] A sterile formulation can be prepared with 100 mg of the
desired compound of the invention as a lyophilized powder
reconstituted in 5 mL of sterile saline. As preservative,
benzalkonium chloride, thimerosal, phenylmercuric nitrate, or the
like may be used in a range of about 0.001% to 1% by weight.
* * * * *