U.S. patent application number 17/372717 was filed with the patent office on 2022-01-06 for hpk1 inhibitors and methods of using same.
The applicant listed for this patent is UNIVERSITY HEALTH NETWORK. Invention is credited to Yunhui Lang, Radoslaw Laufer, Sze-Wan Li, Yong Liu, Grace Ng, Narendra Kumar B. Patel, Heinz W. Pauls, Peter Brent Sampson.
Application Number | 20220002313 17/372717 |
Document ID | / |
Family ID | |
Filed Date | 2022-01-06 |
United States Patent
Application |
20220002313 |
Kind Code |
A1 |
Sampson; Peter Brent ; et
al. |
January 6, 2022 |
HPK1 INHIBITORS AND METHODS OF USING SAME
Abstract
Thienopyridinone compounds of Formula (I) and pharmaceutically
acceptable salts thereof are described. In these compounds, one of
X.sub.1; X.sub.2, and X.sub.3 is S and the other two are each
independently CR, wherein R and all other variables are as defined
herein. The compounds are shown to inhibit HPK1 kinase activity and
to have in vivo antitumor activity. The compounds can be
effectively combined with pharmaceutically acceptable carriers and
also with other immunomodulatory approaches, such as a checkpoint
inhibition or inhibitors of tryptophan oxidation. Formula (I).
##STR00001##
Inventors: |
Sampson; Peter Brent;
(Oakville, CA) ; Patel; Narendra Kumar B.;
(Brampton, CA) ; Pauls; Heinz W.; (Oakville,
CA) ; Li; Sze-Wan; (Toronto, CA) ; Ng;
Grace; (Markham, CA) ; Laufer; Radoslaw;
(Oakville, CA) ; Liu; Yong; (Oakville, CA)
; Lang; Yunhui; (Markham, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
UNIVERSITY HEALTH NETWORK |
Toronto |
|
CA |
|
|
Appl. No.: |
17/372717 |
Filed: |
July 12, 2021 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
16705458 |
Dec 6, 2019 |
11059832 |
|
|
17372717 |
|
|
|
|
15738286 |
Dec 20, 2017 |
10501474 |
|
|
PCT/CA2016/050734 |
Jun 23, 2016 |
|
|
|
16705458 |
|
|
|
|
62184348 |
Jun 25, 2015 |
|
|
|
International
Class: |
C07D 495/04 20060101
C07D495/04; A61K 31/496 20060101 A61K031/496; A61K 39/395 20060101
A61K039/395; C07K 16/28 20060101 C07K016/28; A61K 31/4355 20060101
A61K031/4355; A61K 31/5355 20060101 A61K031/5355; A61K 31/551
20060101 A61K031/551; A61P 35/00 20060101 A61P035/00; A61K 31/5377
20060101 A61K031/5377; A61K 45/06 20060101 A61K045/06 |
Claims
1. A compound represented by Formula (I-C): ##STR00125## or a
pharmaceutically acceptable salt thereof, wherein: R is H, --F,
--Cl, --Br, --OH, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)haloalkyl, --(C.sub.1-C.sub.4)alkoxy,
--(C.sub.1-C.sub.4)alkylene-OH or 4-7 membered monocyclic
heterocyclyl optionally substituted with 1-3 groups selected from
--F, --Cl, --Br, --OH, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)haloalkyl, --(C.sub.1-C.sub.4)alkoxy, or
--CO.sub.2--(C.sub.1-C.sub.4)alkyl; R.sup.1 is --NR.sup.aR.sup.b or
--OR.sup.a1; R.sup.a for each occurrence is independently --H,
--(C.sub.1-C.sub.6)alkyl,
--(CH.sub.2).sub.n--(C.sub.3-C.sub.7)cycloalkyl,
--(CH.sub.2).sub.n-3-7 membered monocyclic heterocyclyl,
--(CH.sub.2).sub.n-bridged (C.sub.6-C.sub.12)cycloalkyl, optionally
substituted --(CH.sub.2).sub.n-5-10 membered heteroaryl; or
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl, wherein
--(C.sub.1-C.sub.6)alkyl,
--(CH.sub.2).sub.n--(C.sub.3-C.sub.7)cycloalkyl,
--(CH.sub.2).sub.n-3-7 membered monocyclic heterocyclyl,
--(CH.sub.2).sub.n-bridged (C.sub.6-C.sub.12)cycloalkyl,
--(CH.sub.2).sub.n-5-10 membered heteroaryl, or
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl, is
optionally substituted with 1-3 groups selected from --F, --Cl,
--Br, --CN, --NH.sub.2, --OH, oxo, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)haloalkyl, --(C.sub.1-C.sub.4)alkoxy,
--(C.sub.1-C.sub.4)haloalkoxy, --(C.sub.1-C.sub.4)alkylene-OH, or
--(C.sub.1-C.sub.4)alkylene-NH.sub.2; R.sup.b for each occurrence
is independently --H or --(C.sub.1-C.sub.6)alkyl; or, R.sup.a and
R.sup.b, together with the nitrogen to which they are attached,
form --(C.sub.3-C.sub.10)heterocyclyl; R.sup.a1 for each occurrence
is independently --H, (C.sub.1-C.sub.6)alkyl,
(C.sub.3-C.sub.10)cycloalkyl, 3-10 membered heterocyclyl,
(C.sub.6-C.sub.10)aryl, or 3-10 membered heteroaryl; R.sub.4 and
R.sub.5, together with the nitrogen to which they are attached,
form 4-7 membered monocyclic heterocyclyl or 6-12 membered bridged
heterocyclyl, wherein the 4-7 membered monocyclic heterocyclyl or
6-12 membered bridged heterocyclyl is optionally substituted with
1-3 groups selected from --F, --Cl, --Br, --CN, --NH.sub.2, --OH,
oxo, --(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl,
--(C.sub.1-C.sub.4)alkoxy, --(C.sub.1-C.sub.4)haloalkoxy,
--(C.sub.1-C.sub.4)alkylene-OH, or
--(C.sub.1-C.sub.4)alkylene-NH.sub.2; R.sub.6 for each occurrence
is independently --F, --Cl, --Br, --CN, --NH.sub.2, --OH,
--(C.sub.1-C.sub.6)alkyl, --(C.sub.1-C.sub.6)haloalkyl,
--(C.sub.2-C.sub.6)alkenyl, --(C.sub.2-C.sub.6)alkynyl,
(C.sub.3-C.sub.6)cycloalkyl, --(C.sub.1-C.sub.6)alkoxy,
--(C.sub.1-C.sub.6)haloalkoxy, --(C.sub.1-C.sub.6)alkylene-OH, or
--(C.sub.1-C.sub.6)alkylene-NH.sub.2; m is 0, 1, 2, or 3; and n is
0, 1, or 2.
2-6. (canceled)
7. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein the compound is represented by formula (II-C):
##STR00126##
8-9. (canceled)
10. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein the compound is represented by formula (III-C):
##STR00127##
11-13. (canceled)
14. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R.sub.4 and R.sub.5, together with the nitrogen to
which they are attached, form --N-alkyl-piperazinyl or morpholinyl,
wherein the piperazinyl or morpholinyl is optionally substituted
with 1-2 groups selected from --F, --Cl, --Br, --OH,
--(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl, or
--(C.sub.1-C.sub.4)alkoxy.
15. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R.sup.a for each occurrence is independently --H,
--(CH.sub.2).sub.n--(C.sub.3-C.sub.6)cycloalkyl,
--(CH.sub.2).sub.n-3-6 membered monocyclic heterocyclyl, wherein
the --(CH.sub.2).sub.n--(C.sub.3-C.sub.6)cycloalkyl or
--(CH.sub.2).sub.n-3-6 membered monocyclic heterocyclyl is
optionally substituted with 1-3 groups selected from --F, --Cl,
--Br, --CN, --NH.sub.2, --OH, --(C.sub.1-C.sub.4)alkyl, or
--(C.sub.1-C.sub.4)alkoxy; and n is 0 or 1.
16. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R is H, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)alkoxy, N-piperazinyl optionally substituted
with --CO.sub.2--(C.sub.1-C.sub.4)alkyl.
17. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R is H.
18. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R.sub.4 and R.sub.5, together with the nitrogen to
which they are attached, form --N-methyl-piperazinyl or
morpholinyl, both of which are optionally substituted with one or
two methyl.
19. The compound of claim 1 or a pharmaceutically acceptable salt
thereof, wherein R.sup.a for each occurrence is independently --H;
--(C.sub.3-C.sub.6)cycloalkyl optionally substituted with --OH;
--(CH.sub.2).sub.n-tetrahydro-2H-pyran; morpholinyl; piperidinyl
optionally substituted with --F, --OH or methyl; or
tetrahydrofuran; and n is 0 or 1.
20. A pharmaceutical composition comprising a compound of claim 1
or a pharmaceutically acceptable salt thereof and a
pharmaceutically acceptable carrier or diluent.
21. A method for treating a subject with cancer, comprising:
administering to the subject an effective amount of a compound of
claim 1, or a pharmaceutically acceptable salt thereof.
22. A method of treating a subject with cancer, comprising
administering to the subject an effective amount of a compound of
claim 1, or a pharmaceutically acceptable salt thereof, and an
effective second anti-cancer treatment.
23. A method of treating a subject with cancer, comprising
administering to the subject an effective amount of a compound of
claim 1, or a pharmaceutically acceptable salt thereof, and an
effective amount of an immunomodulatory agent such as a checkpoint
inhibitor or an inhibitor of tryptophan oxidation.
Description
RELATED APPLICATIONS
[0001] This application claims the benefit of U.S. Provisional
Application No. 62/184,348, filed Jun. 25, 2015. The entire
teachings of the aforementioned application are incorporated herein
by reference.
BACKGROUND
[0002] Hematopoietic progenitor kinase 1 (HPK1) is a hematopoietic
cell-restricted Ste20 serine/threonine kinase. HPK1 kinase activity
can be induced by activation signals generated by various different
cell surface receptors found in hematopoietic cells upon ligand
engagement. Ligand engagement or antibody-mediated crosslinking of
T cell receptors (TCR), B cell antigen receptor (BCR) (Liou et al.,
2000, Immunity 12:399), transforming growth factor .beta. receptor
(TGF-.beta.R) (Wang et al., 1997. J. Biol. Chem. 272:22771; Zhou et
al., 1999, J. Biol. Chem. 274:13133), erythropoietin receptor
(EPOR) (Nagata et al., 1999, Blood 93:3347), and Fas (Chen et al.,
1999, Oncogene 18:7370) can induce HPK1 kinase activity. Each
receptor utilizes unique, but sometimes overlapping, signaling
mechanisms to activate HPK1. HPK1 acts as a down-modulator of T and
B cell functions through the AP-1, NFKB, Erk2, and Fos pathways;
for example, HPK1 has been implicated as a negative regulator of
signal transduction in T-cells through phosphorylation and
activation of the T-cell receptor adaptor protein SLP-76 (Di
Bartolo et al., 2007, J. Exp. Med. 204:681), which leads to
subsequent downregulation of the AP-1 and Erk2 pathways. In
B-cells, HPK1 downregulates B-cell receptor (BCR) signaling through
phosphorylation of the SLP-76 paralog BLINK (Wang et al., 2012, J
Biol. Chem. 287:11037).
[0003] Thus, HPK1 is now viewed as a possible target for
therapeutic intervention. For example, it has been reported that
HPK1 can be a novel target for cancer immunotherapy (Sawasdikosol
et al., Immunol Res. 2012 December; 54(1-3):262-5). Specifically,
targeted disruption of HPK1 alleles confers T cells with an
elevated Th1 cytokine production in response to TCR engagement.
HPK1 (-/-) T cells proliferate more rapidly than the
haplotype-matched wild-type counterpart and are resistant to
prostaglandin E2 (PGE(2))-mediated suppression. Most strikingly,
mice that received adoptive transfer of HPK1 (-/-) T cells became
resistant to lung tumor growth. Also, the loss of HPK1 from
dendritic cells (DCs) endows them with superior antigen
presentation ability, enabling HPK1 (-/-) DCs to elicit a more
potent anti-tumor immune response when used as cancer vaccine.
[0004] When evaluating if a small-molecule inhibitor of HPK1 would
capture the phenotype of mice with targeted disruption of the gene,
it is important to consider the non-catalytic roles of the protein.
In particular, while full-length HPK1 can promote TCR-mediated
activation of the nuclear factor kappa-light-chain-enhancer of
activated B cells (NF-.kappa.B) pathway, the catalytically inactive
cleavage product HPK1-C can suppress NF-.kappa.B activation upon
TCR restimulation, leading to activation-induced cell death (AICD)
(Brenner et al., EMBO J. 2005, 24:4279). Taking together the
catalytic and non-catalytic roles of HPK1, it is possible that
blocking the HPK1 kinase activity with a small-molecule inhibitor
may promote activation of B- and T-cells, leading to superior
anti-tumor immunity, while also facilitating AICD, helping to
maintain peripheral immune tolerance. The exact effects of an HPK1
inhibitor would be borne out by testing in mouse models of cancer,
such as syngeneic tumor xenografts. Given that HPK1 is not
expressed in any major organs, outside the hematopoietic system, it
is less likely that an inhibitor of HPK1 kinase activity would
cause any serious side effects.
[0005] In view of the above, there is a need in the art for novel
compounds that can inhibit HPK1.
SUMMARY OF THE INVENTION
[0006] Applicant has now discovered that certain thienopyridinone
compounds are HPK1 inhibitors (see Example B). They also have
inhibitory activities against FLT3 and LCK (see Example C).
Additionally, it has been demonstrated that certain
thienopyridinone compounds as HPK1 inhibitors alone, and in
combination with anti-PD-1 antibodies are effective in pre-clinical
models with certain cancer cell types (see Example E). The
particular combination therapies disclosed herein demonstrate
surprising biological activity with significant anticancer effects.
Specifically, with the combination of HPK1 inhibitors and anti-PD-1
antibodies, significant responses following PD-1/PD-L1 blockade
have now been demonstrated in CT26.WT colon carcinoma. Based on
these discoveries, thienopyridinone compounds, pharmaceutical
compositions thereof, and methods of using the same are disclosed
herein.
[0007] One embodiment of the invention is a compound represented by
Structural Formula
##STR00002##
or a pharmaceutically acceptable salt thereof. Values for each of
the variables are provided below.
[0008] Another embodiment of the invention is a pharmaceutical
composition comprising a pharmaceutically acceptable carrier or
diluent and a compound represented by Structural Formula (I)
described above or a pharmaceutically acceptable salt thereof.
[0009] Another embodiment of the invention is a method of treating
a subject with a disease which can be regulated by HPK1 comprising
administering to the subject an effective amount of a compound of
Structural Formula (I) or a pharmaceutically acceptable salt
thereof.
[0010] Another embodiment of the invention is a method of
inhibiting HPK1 activity in a subject in need of inhibition of HPK1
activity, comprising administering to the subject an effective
amount of a compound represented by Structural Formula (I) or a
pharmaceutically acceptable salt thereof.
[0011] Another embodiment of the invention is a compound
represented by Structural Formula (I) or a pharmaceutically
acceptable salt thereof for use in therapy. In some embodiments,
the therapy is for treating a subject with cancer. Alternatively,
the therapy is for inhibiting HPK1 activity in a subject in need of
inhibition of HPK1 activity.
[0012] Another embodiment of the invention is the use of a compound
represented by Structural Formula (I) or a pharmaceutically
acceptable salt thereof for the manufacture of a medicament for
treating a subject with cancer.
[0013] Another embodiment of the invention the use of a compound
represented by Structural Formulas (I) or a pharmaceutically
acceptable salt thereof for the manufacture of a medicament for
inhibiting HPK1 activity in a subject in need of inhibition of HPK1
activity. The present invention is also directed to a method of
treating a subject with cancer, comprising administering to the
subject an effective amount of a HPK1 inhibitor (e.g., a compound
represented by Structural Formula (I)), or a pharmaceutically
acceptable salt thereof, and an effective second anti-cancer
treatment (e.g., a chemotherapeutic agent, a targeted therapeutic
agent, radiation or surgery). In one example, the second
anti-cancer treatment is a PD-1 inhibitor.
[0014] The present invention is also directed to a method of
treating a subject with cancer, comprising administering to the
subject an effective amount of a HPK1 inhibitor (e.g., a compound
represented by Structural Formula (I)), or a pharmaceutically
acceptable salt thereof, and an effective amount of an
immunomodulatory agent such as a checkpoint inhibitor (e.g.,
anti-PD-1 antibody, anti-CTLA4 antibody or anti-PD-L1 antibody) or
an inhibitor of tryptophan oxidation (e.g. IDO1, IDO2 or TDO2
inhibitor). In one example, the immunomodulatory agent is anti-PD-1
antibody.
[0015] In an embodiment, the present invention further provides the
use of a HPK1 inhibitor (e.g., a compound represented by Structural
Formula (I), or a pharmaceutically acceptable salt thereof), for
the manufacture of a medicament for the treatment of a subject with
cancer, in combination with a PD-1 inhibitor such as nivolumab,
pembrolizumab, pidilizumab, BMS 936559, MPDL3280A, MSB0010718C or
MED14736. Preferably, the PD-1 inhibitor is nivolumab.
Alternatively, the PD-1 inhibitor is pembrolizumab. In one
embodiment, the PD-1 inhibitor is anti-PD1 antibody.
[0016] In one alternative, the HPK1 inhibitor is administered with
an effective amount of one or more other anti-cancer therapies, and
preferably in combination with PD-1 inhibitor.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] FIG. 1 shows the inhibitory effect of compound example A30
against SLP-76 serine 376 phosphorylation in .alpha.-CD3 stimulated
Jurkat E6.1 cells.
[0018] FIG. 2 is a graph illustrating the tumour growth inhibition
percentage following administration of compound A1 alone and in
combination with an anti-PD1 antibody.
[0019] FIG. 3 shows the effect of compound example A30 in the EAE
disease progression model.
DETAILED DESCRIPTION OF THE INVENTION
[0020] In a first embodiment, the invention is directed to a
compound represented by Formula (I):
##STR00003##
or a pharmaceutically acceptable salt thereof, wherein:
[0021] one of X.sub.1, X.sub.2, and X.sub.3 is S, the other two are
each independently CR, wherein R is --H, --F, --Cl, --Br, --CN,
--NH.sub.2, --OH, optionally substituted (C.sub.1-C.sub.6)alkyl,
optionally substituted (C.sub.1-C.sub.6)alkoxy, optionally
substituted --(CH.sub.2).sub.n--(C.sub.3-C.sub.10)cycloalkyl,
optionally substituted --(CH.sub.2).sub.n-3-7 membered monocyclic
heterocyclyl, optionally substituted --(CH.sub.2).sub.nphenyl,
optionally substituted --(CH.sub.2).sub.n-5-7 membered monocyclic
heteroaryl, optionally substituted --(CH.sub.2).sub.n-bridged
(C.sub.6-C.sub.12)cycloalkyl, optionally substituted
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl, optionally
substituted --(CH.sub.2).sub.n-7-12 membered bicyclic heteroaryl,
or optionally substituted --(CH.sub.2).sub.n-7-12 membered bicyclic
heteroaryl;
[0022] Y is a bond, --CH.sub.2--, --C(.dbd.O)--;
[0023] R.sub.1 is --NR.sup.aR.sup.b or --OR.sup.a1;
[0024] R.sup.a for each occurrence is independently --H, optionally
substituted (C.sub.1-C.sub.6)alkyl, optionally substituted
--(CH.sub.2).sub.n(C.sub.3-C.sub.10)cycloalkyl, optionally
substituted --(CH.sub.2).sub.n-3-10 membered heterocyclyl,
optionally substituted --(CH.sub.2).sub.n(C.sub.6-C.sub.10)aryl,
optionally substituted --(CH.sub.2).sub.n-5-10 membered heteroaryl,
optionally substituted --(CH.sub.2).sub.n-bridged
(C.sub.6-C.sub.12)cycloalkyl, or optionally substituted
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl;
[0025] R.sup.b for each occurrence is independently --H or
--(C.sub.1-C.sub.6)alkyl; or,
[0026] R.sup.a and R.sup.b, together with the nitrogen to which
they are attached, form optionally substituted
--(C.sub.3-C.sub.10)heterocyclyl;
[0027] R.sup.a1 for each occurrence is independently --H,
optionally substituted (C.sub.1-C.sub.6)alkyl, optionally
substituted (C.sub.3-C.sub.10)cycloalkyl, optionally substituted
3-10 membered heterocyclyl, optionally substituted
(C.sub.6-C.sub.10)aryl, or optionally substituted 3-10 membered
heteroaryl; or
[0028] R.sub.2 and R.sub.3 are each independently --H or
--(C.sub.1-C.sub.6)alkyl;
[0029] R.sub.4 and R.sub.5 are each independently --H, optionally
substituted (C.sub.1-C.sub.6)alkyl, optionally substituted
(C.sub.3-C.sub.10)cycloalkyl, optionally substituted 3-10 membered
heterocyclyl, optionally substituted (C.sub.6-C.sub.10)aryl,
optionally substituted 5-10 membered heteroaryl, optionally
substituted bridged (C.sub.6-C.sub.12)cycloalkyl, or optionally
substituted 6-12 membered bridged heterocyclyl; or
[0030] R.sub.4 and R.sub.5, together with the nitrogen to which
they are attached, form optionally substituted 4-10 membered
heterocyclyl, optionally substituted 5-10 membered heteroaryl, or
optionally substituted 6-12 membered bridged heterocyclyl;
[0031] R.sub.6 for each occurrence is independently --F, --Cl,
--Br, --CN, --NH.sub.2, --OH, --(C.sub.1-C.sub.6)alkyl,
--(C.sub.1-C.sub.6)haloalkyl, --(C.sub.2-C.sub.6)alkenyl,
--(C.sub.2-C.sub.6)alkynyl, (C.sub.3-C.sub.6)cycloalkyl,
--(C.sub.1-C.sub.6)alkoxy, --(C.sub.1-C.sub.6)haloalkoxy,
--(C.sub.1-C.sub.6)alkylene-OH, or
--(C.sub.1-C.sub.6)alkylene-NH.sub.2;
[0032] m is 0, 1, 2, or 3; and
[0033] n is 0, 1, or 2.
[0034] In a second embodiment, the invention provides a compound
represented by structural formula (I-A)-(I-C), (II-A)-(II-C), or
(III-A)-(III-C):
##STR00004## ##STR00005##
or a pharmaceutically acceptable salt thereof. Values for the
variables in Structural Formulae (I-A)-(I-C), (II-A)-(II-C), and
(III-A)-(III-C) are as described for Structural Formula (I).
[0035] In a third embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sub.4 and R.sub.5, together with the
nitrogen to which they are attached, form 4-7 membered monocyclic
heterocyclyl or 6-12 membered bridged heterocyclyl, wherein the 4-7
membered monocyclic heterocyclyl or 6-12 membered bridged
heterocyclyl is optionally substituted with 1-3 groups selected
from --F, --Cl, --Br, --CN, --NH.sub.2, --OH, oxo,
--(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl,
--(C.sub.1-C.sub.4)alkoxy, --(C.sub.1-C.sub.4)haloalkoxy,
--(C.sub.1-C.sub.4)alkylene-OH, or
--(C.sub.1-C.sub.4)alkylene-NH.sub.2. Values for the remainder of
the variables are as described for Structural Formula (I).
[0036] In a fourth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sup.a for each occurrence is
independently --H, --(C.sub.1-C.sub.6)alkyl,
--(CH.sub.2).sub.n--(C.sub.3-C.sub.7)cycloalkyl,
--(CH.sub.2).sub.n-4-7 membered monocyclic heterocyclyl,
--(CH.sub.2).sub.n-bridged (C.sub.6-C.sub.12)cycloalkyl, optionally
substituted --(CH.sub.2).sub.n-5-10 membered heteroaryl; or
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl, wherein
--(C.sub.1-C.sub.6)alkyl,
--(CH.sub.2).sub.n--(C.sub.3-C.sub.7)cycloalkyl,
--(CH.sub.2).sub.n-4-7 membered monocyclic heterocyclyl,
--(CH.sub.2).sub.n-bridged (C.sub.6-C.sub.12)cycloalkyl,
--(CH.sub.2).sub.n-5-10 membered heteroaryl, or
--(CH.sub.2).sub.n-6-12 membered bridged heterocyclyl, is
optionally substituted with 1-3 groups selected from --F, --Cl,
--Br, --CN, --NH.sub.2, --OH, oxo, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)haloalkyl, --(C.sub.1-C.sub.4)alkoxy,
--(C.sub.1-C.sub.4)haloalkoxy, --(C.sub.1-C.sub.4)alkylene-OH, or
--(C.sub.1-C.sub.4)alkylene-NH.sub.2, and values for the remainder
of the variables are as described above for Structural Formula (I)
or in the third embodiment.
[0037] In a fifth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R is H, --F, --Cl, --Br, --OH,
--(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl,
--(C.sub.1-C.sub.4)alkoxy, --(C.sub.1-C.sub.4)alkylene-OH or 4-7
membered monocyclic heterocyclyl optionally substituted with 1-3
groups selected from --F, --Cl, --Br, --OH,
--(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl, or
--(C.sub.1-C.sub.4)alkoxy, and values for the remainder of the
variables are as described above for Structural Formula (I) or in
the third or fourth embodiment.
[0038] In a sixth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sub.4 and R.sub.5, together with the
nitrogen to which they are attached, form --N-alkyl-piperazinyl or
morpholinyl, wherein the piperazinyl or morpholinyl is optionally
substituted with 1-2 groups selected from --F, --Cl, --Br, --OH,
--(C.sub.1-C.sub.4)alkyl, --(C.sub.1-C.sub.4)haloalkyl, or
--(C.sub.1-C.sub.4)alkoxy, and values for the remainder of the
variables are as described above for Structural Formula (I), or in
the third, fourth, or fifth embodiment.
[0039] In a seventh embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sup.a for each occurrence is
independently --H, --(CH.sub.2).sub.n--(C.sub.3-C.sub.6)cycloalkyl,
--(CH.sub.2).sub.n-3-6 membered heterocyclyl, wherein the
--(CH.sub.2).sub.n--(C.sub.3-C.sub.6)cycloalkyl or
--(CH.sub.2).sub.n-3-6 membered heterocyclyl is optionally
substituted with 1-3 groups selected from --F, --Cl, --Br, --CN,
--NH.sub.2, --OH, --(C.sub.1-C.sub.4)alkyl, or
--(C.sub.1-C.sub.4)alkoxy; and n is 0 or 1, and values for the
remainder of the variables are as described above for Structural
Formula (I), or in the third, fourth, fifth, or sixth
embodiment.
[0040] In an eighth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R is H, --(C.sub.1-C.sub.4)alkyl,
--(C.sub.1-C.sub.4)alkoxy, N-piperazinyl optionally substituted
with --CO.sub.2--(C.sub.1-C.sub.4)alkyl, and values for the
remainder of the variables are as described above for Structural
Formula (I), or in the third, fourth, fifth, sixth, or seventh
embodiment. Alternatively, R is H.
[0041] In a ninth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sub.4 and R.sub.5, together with the
nitrogen to which they are attached, form --N-methyl-piperazinyl or
morpholinyl, both of which are optionally substituted with one or
two methyl, and values for the remainder of the variables are as
described above for Structural Formula (I), or in the third,
fourth, fifth, sixth, seventh, or eighth embodiment.
[0042] In a tenth embodiment, the invention provides a compound
represented by structural formula (I), (I-A)-(I-C), (II-A)-(II-C),
or (III-A)-(III-C), wherein R.sup.a for each occurrence is
independently --H; --(C.sub.3-C.sub.6)cycloalkyl optionally
substituted with --OH; --(CH.sub.2).sub.n-tetrahydro-2H-pyran;
morpholinyl; piperidinyl optionally substituted with --F, --OH or
methyl; or tetrahydrofuran; and n is 0 or 1, and values for the
remainder of the variables are as described above for Structural
Formula (I), or in the third, fourth, fifth, sixth, seventh, eighth
or ninth embodiment.
[0043] The invention also includes the compounds depicted by
structure and/or described by name in the Exemplification. The
invention includes both the neutral form (free base) of these
compounds as well as pharmaceutically acceptable salts thereof.
Treatments with and/or uses of these compounds includes the neutral
form of these compounds as well as pharmaceutically acceptable
salts thereof.
[0044] The term "alkyl" used alone or as part of a larger moiety,
such as "alkoxy" or "haloalkyl" and the like, means saturated
aliphatic straight-chain or branched monovalent hydrocarbon
radical. Unless otherwise specified, an alkyl group typically has
1-6 carbon atoms, i.e. (C.sub.1-C.sub.6)alkyl. As used herein, a
"(C.sub.1-C.sub.6)alkyl" group means a radical having from 1 to 6
carbon atoms in a linear or branched arrangement. Examples include
methyl, ethyl, n-propyl, iso-propyl etc.
[0045] "Alkoxy" means an alkyl radical attached through an oxygen
linking atom, represented by --O-alkyl. For example,
"(C.sub.1-C.sub.4)alkoxy" includes methoxy, ethoxy, propoxy, and
butoxy.
[0046] The terms "haloalkyl" and "haloalkoxy" means alkyl or
alkoxy, as the case may be, substituted with one or more halogen
atoms. The term "halogen" means F, Cl, Br or I. Preferably the
halogen in a haloalkyl or haloalkoxy is F.
[0047] "Alkenyl" means branched or straight-chain monovalent
hydrocarbon radical containing at least one double bond. Alkenyl
may be mono or polyunsaturated, and may exist in the E or Z
configuration. Unless otherwise specified, an alkenyl group
typically has 2-6 carbon atoms, i.e. (C.sub.2-C.sub.6)alkenyl. For
example, "(C.sub.2-C.sub.6)alkenyl" means a radical having from 2-6
carbon atoms in a linear or branched arrangement.
[0048] "Alkynyl" means branched or straight-chain monovalent
hydrocarbon radical containing at least one triple bond. Unless
otherwise specified, an alkynyl group typically has 2-6 carbon
atoms, i.e. (C.sub.2-C.sub.6)alkynyl. For example,
"(C.sub.2-C.sub.6)alkynyl" means a radical having from 2-6 carbon
atoms in a linear or branched arrangement.
[0049] "Cycloalkyl" means a saturated aliphatic cyclic hydrocarbon
radical, typically containing from 3-8 ring carbon atoms, i.e.,
(C.sub.3-C.sub.8)cycloalkyl. (C.sub.3-C.sub.8)cycloalkyl includes,
but is not limited to, cyclopropyl, cyclobutyl, cyclopentyl,
cyclohexyl, cycloheptyl and cyclooctyl.
[0050] As used herein, the term "bridged" used alone or as part of
a larger moiety as in "bridged cycloalkyl" or "bridged
heterocyclyl" refers to a ring system which includes two rings that
share at least three adjacent ring atoms. Bridged cycloalkyl
typically contains 6-12 ring carbon atoms. Bridged heterocyclyl
typically have 6-12 ring atoms selected from carbon and at least
one (typically 1 to 4, more typically 1 or 2) heteroatom (e.g.,
oxygen, nitrogen or sulfur).
[0051] The term "aryl" used alone or as part of a larger moiety as
in "arylalkyl", "arylalkoxy", or "aryloxyalkyl", means a
carbocyclic aromatic ring. It also includes a phenyl ring fused
with a cycloalkyl group. The term "aryl" may be used
interchangeably with the terms "aryl ring" "carbocyclic aromatic
ring", "aryl group" and "carbocyclic aromatic group". An aryl group
typically has six to fourteen ring atoms. Examples includes phenyl,
naphthyl, anthracenyl, 1,2-dihydronaphthyl,
1,2,3,4-tetrahydronaphthyl, fluorenyl, indanyl, indenyl and the
like. A "substituted aryl group" is substituted at any one or more
substitutable ring atom, which is a ring carbon atom bonded to a
hydrogen.
[0052] The term "heteroaryl", "heteroaromatic", "heteroaryl ring",
"heteroaryl group", "heteroaromatic ring", and "heteroaromatic
group", are used interchangeably herein. "Heteroaryl" when used
alone or as part of a larger moiety as in "heteroarylalkyl" or
"heteroarylalkoxy", refers to aromatic ring groups having five to
fourteen ring atoms selected from carbon and at least one
(typically 1 to 4, more typically 1 or 2) heteroatoms (e.g.,
oxygen, nitrogen or sulfur). "Heteroaryl" includes monocyclic rings
and polycyclic rings in which a monocyclic heteroaromatic ring is
fused to one or more other aryl, heterocyclyl or heteroaromatic
rings. As such, "5-14 membered heteroaryl" includes monocyclic,
bicyclic or tricyclic ring systems.
[0053] Examples of monocyclic 5-6 membered heteroaryl groups
include furanyl (e.g., 2-furanyl, 3-furanyl), imidazolyl (e.g.,
N-imidazolyl, 2-imidazolyl, 4-imidazolyl, 5-imidazolyl), isoxazolyl
(e.g., 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl), oxadiazolyl
(e.g., 2-oxadiazolyl, 5-oxadiazolyl), oxazolyl (e.g., 2-oxazolyl,
4-oxazolyl, 5-oxazolyl), pyrazolyl (e.g., 3-pyrazolyl,
4-pyrazolyl), pyrrolyl (e.g., 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl),
pyridyl (e.g., 2-pyridyl, 3-pyridyl, 4-pyridyl), pyrimidinyl (e.g.,
2-pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl), pyridazinyl (e.g.,
3-pyridazinyl), thiazolyl (e.g., 2-thiazolyl, 4-thiazolyl,
5-thiazolyl), isothiazolyl, triazolyl (e.g., 2-triazolyl,
5-triazolyl), tetrazolyl (e.g., tetrazolyl), and thienyl (e.g.,
2-thienyl, 3-thienyl). Examples of polycyclic aromatic heteroaryl
groups include carbazolyl, benzimidazolyl, benzothienyl,
benzofuranyl, isobenzofuranyl, indolyl, benzotriazolyl,
benzothiazolyl, benzoxazolyl, quinolinyl, isoquinolinyl, indazolyl,
isoindolyl, acridinyl, or benzisoxazolyl. A "substituted heteroaryl
group" is substituted at any one or more substitutable ring atom,
which is a ring carbon or ring nitrogen atom bonded to a
hydrogen.
[0054] "Heterocyclyl" means a saturated or unsaturated non-aromatic
3-12 membered ring radical optionally containing one or more double
bonds. It can be monocyclic, bicyclic, tricyclic, or fused. The
heterocycloalkyl contains 1 to 4 heteroatoms, which may be the same
or different, selected from N, O or S. The heterocyclyl ring
optionally contains one or more double bonds and/or is optionally
fused with one or more aromatic rings (e.g., phenyl ring). The term
"heterocyclyl" is intended to include all the possible isomeric
forms. Examples of heterocycloalkyl include, but are not limited
to, azetidinyl, morpholinyl, thiomorpholinyl, pyrrolidinonyl,
pyrrolidinyl, piperidinyl, piperazinyl, hydantoinyl,
valerolactamyl, oxiranyl, oxetanyl, dihydroimidazole,
dihydrofuranyl, dihydropyranyl, dihydropyridinyl,
dihydropyrimidinyl, dihydrothienyl, dihydrothiophenyl,
dihydrothiopyranyl, tetrahydroimidazole, tetrahydrofuranyl,
tetrahydropyranyl, tetrahydrothienyl, tetrahydropyridinyl,
tetrahydropyrimidinyl, tetrahydrothiophenyl, and
tetrahydrothiopyranyl. Examples of polycyclic heterocycloalkyl
groups include dihydroindolyl, dihydroisoindolyl,
dihydrobenzimidazolyl, dihydrobenzothienyl, dihydrobenzofuranyl,
dihydroisobenzofuranyl, dihydrobenzotriazolyl,
dihydrobenzothiazolyl, dihydrobenzoxazolyl, dihydroquinolinyl,
tetrahydroquinolinyl, dihydroisoquinolinyl,
tetrahydroisoquinolinyl, dihydroindazolyl, dihydroacridinyl,
tetrahydroacridinyl, dihydrobenzisoxazolyl, chroman, chromene,
isochroman and isochromene.
[0055] Certain of the compounds described herein may exist in
various stereoisomeric or tautomeric forms. Stereoisomers are
compounds which differ only in their spatial arrangement. When a
disclosed compound is named or depicted by structure without
indicating stereochemistry, it is understood that the name or
structure encompasses all possible stereoisomers, geometric
isomers, including essentially pure stereo or geometric isomers, as
well as combination thereof.
[0056] In certain instances tautomeric forms of the disclosed
compounds exist, such as the tautomeric structures shown below:
##STR00006##
[0057] It is to be understood that when a compound herein is
represented by a structural formula or designated by a chemical
name herein, all other tautomeric forms which may exist for the
compound are encompassed by the structural formula.
[0058] Certain of the disclosed compounds may exist in various
stereoisomeric forms. Stereoisomers are compounds that differ only
in their spatial arrangement. Enantiomers are pairs of
stereoisomers whose mirror images are not superimposable, most
commonly because they contain an asymmetrically substituted carbon
atom that acts as a chiral center. "Enantiomer" means one of a pair
of molecules that are mirror images of each other and are not
superimposable. Diastereomers are stereoisomers that contain two or
more asymmetrically substituted carbon atoms. "Geometric isomers"
are stereoisomers that differ in the orientation of substituent
atoms in relationship to a carbon-carbon double bond, to a
carbocyclyl ring, or to a bridged bicyclic system.
[0059] When a geometric isomer is depicted by name or structure, it
is to be understood that the geometric isomeric purity of the named
or depicted geometric isomer is at least 60%, 70%, 80%, 90%, 99% or
99.9% pure by weight. Geometric isomeric purity is determined by
dividing the weight of the named or depicted geometric isomer in
the mixture by the total weight of all of the geometric isomers in
the mixture.
[0060] When the stereochemistry of a disclosed compound is named or
depicted by structure, the named or depicted stereoisomer is at
least 60%, 70%, 80%, 90%, 99% or 99.9% by weight pure relative to
all of the other stereoisomers. Percent by weight pure relative to
all of the other stereoisomers is the ratio of the weight of one
stereoisomer over the weight of the other stereoisomers. When a
single enantiomer is named or depicted by structure, the depicted
or named enantiomer is at least 60%, 70%, 80%, 90%, 99% or 99.9% by
weight optically pure (also referred to as "enantiomerically
pure"). Percent optical purity by weight is the ratio of the weight
of the enantiomer over the weight of the enantiomer plus the weight
of its optical isomer.
[0061] When the stereochemistry of a disclosed compound is named or
depicted by structure, and the named or depicted structure
encompasses more than one stereoisomer (e.g., as in a
diastereomeric pair), it is to be understood that one of the
encompassed stereoisomers or any mixture of the encompassed
stereoisomers are included. It is to be further understood that the
stereoisomeric purity of the named or depicted stereoisomers at
least 60%, 70%, 80%, 90%, 99% or 99.9% by weight pure relative to
all of the other stereoisomers. The stereoisomeric purity in this
case is determined by dividing the total weight in the mixture of
the stereoisomers encompassed by the name or structure by the total
weight in the mixture of all of the stereoisomers.
[0062] When a disclosed compound is named or depicted by structure
without indicating the stereochemistry, and the compound has one
chiral center, it is to be understood that the name or structure
encompasses one enantiomer of compound free from the corresponding
optical isomer, a racemic mixture of the compound and mixtures
enriched in one enantiomer relative to its corresponding optical
isomer.
[0063] When a disclosed compound is named or depicted by structure
without indicating the stereochemistry and e.g., the compound has
at least two chiral centers, it is to be understood that the name
or structure encompasses one stereoisomer free of other
stereoisomers, mixtures of stereoisomers, and mixtures of
stereoisomers in which one or more stereoisomers is enriched
relative to the other stereoisomer(s). For example, the name or
structure may encompass one stereoisomer free of other
diastereomers, mixtures of stereoisomers, and mixtures of
stereoisomers in which one or more diastereomers is enriched
relative to the other diastereomer(s).
[0064] Enantiomeric and diastereomeric mixtures can be resolved
into their component enantiomers or stereoisomers by well-known
methods, such as chiral-phase gas chromatography, chiral-phase high
performance liquid chromatography, crystallizing the compound as a
chiral salt complex, or crystallizing the compound in a chiral
solvent.
[0065] Enantiomers and diastereomers can also be obtained from
diastereomerically- or enantiomerically-pure intermediates,
reagents, and catalysts by well-known asymmetric synthetic
methods.
[0066] Included in the present teachings are pharmaceutically
acceptable salts of the compounds disclosed herein. The disclosed
compounds have basic amine groups and therefore can form
pharmaceutically acceptable salts with pharmaceutically acceptable
acid(s). Suitable pharmaceutically acceptable acid addition salts
of the compounds described herein include salts of inorganic acids
(such as hydrochloric acid, hydrobromic, phosphoric,
metaphosphoric, nitric, and sulfuric acids) and of organic acids
(such as acetic acid, benzenesulfonic, benzoic, ethanesulfonic,
methanesulfonic, succinic, and trifluoroacetic acid acids).
Compounds of the present teachings with acidic groups such as
carboxylic acids can form pharmaceutically acceptable salts with
pharmaceutically acceptable base(s). Suitable pharmaceutically
acceptable basic salts include ammonium salts, alkali metal salts
(such as sodium and potassium salts) and alkaline earth metal salts
(such as magnesium and calcium salts). Compounds with a quaternary
ammonium group also contain a counteranion such as chloride,
bromide, iodide, acetate, perchlorate and the like. Other examples
of such salts include hydrochlorides, hydrobromides, sulfates,
methanesulfonates, nitrates, acetates, succinates, benzoates and
salts with amino acids such as glutamic acid.
[0067] Compounds described herein can inhibit HPK1. Thus,
generally, compounds described herein are useful in the treatment
of diseases or conditions associated with such kinases.
[0068] In one embodiment, the compounds described herein are HPK1
inhibitors, and are useful for treating diseases, such as cancer,
associated with such kinase(s).
[0069] Another aspect of the present teachings relates to a method
of treating a subject with cancer comprising administering to the
subject an effective amount of a compound described herein. In one
embodiment, the compounds described herein inhibit the growth of a
tumor.
[0070] Cancers that can be treated (including reduction in the
likelihood of recurrence) by the methods of the present teachings
include breast cancer, colorectal cancer, lung cancer, ovarian
cancer, uterine cancer, prostate cancer, leukemias, lymphomas,
brain cancer (including glioblastoma multiforme and neuroblastoma),
head and neck cancer, pancreatic cancer, melanoma, hepatocellular
carcinoma, renal cancer, and soft tissue sarcomas. In one
embodiment, the cancer is breast cancer, colon cancer, and ovarian
cancer. In one embodiment, the cancer is selected from leukemia,
acute myeloid leukemia, chronic myelogenous leukemia, breast
cancer, brain cancer, colon cancer, colorectal cancer, head and
neck cancer, hepatocellular carcinoma, lung adenocarcinoma,
metastatic melanoma, pancreatic cancer, prostate cancer, ovarian
cancer and renal cancer. In one embodiment, the cancer is lung
cancer, colon cancer, brain cancer, neuroblastoma, prostate cancer,
melanoma, glioblastoma multiforme or ovarian cancer. In another
embodiment, the cancer is lung cancer, breast cancer, colon cancer,
brain cancer, neuroblastoma, prostate cancer, melanoma,
glioblastoma multiforme or ovarian cancer. In yet another
embodiment, the cancer is breast cancer, colon cancer and lung
cancer. In another embodiment, the cancer is a breast cancer. In
yet another embodiment, the cancer is a basal sub-type breast
cancer or a luminal B sub-type breast cancer. In yet another
embodiment, the cancer is a basal sub-type breast cancer. In yet
another embodiment, the basal sub-type breast cancer is ER
(estrogen receptor), HER2 and PR (progesterone receptor) negative
breast cancer. In yet another embodiment, the cancer is a soft
tissue cancer. A "soft tissue cancer" is an art-recognized term
that encompasses tumors derived from any soft tissue of the body.
Such soft tissue connects, supports, or surrounds various
structures and organs of the body, including, but not limited to,
smooth muscle, skeletal muscle, tendons, fibrous tissues, fatty
tissue, blood and lymph vessels, perivascular tissue, nerves,
mesenchymal cells and synovial tissues. Thus, soft tissue cancers
can be of fat tissue, muscle tissue, nerve tissue, joint tissue,
blood vessels, lymph vessels, and fibrous tissues. Soft tissue
cancers can be benign or malignant. Generally, malignant soft
tissue cancers are referred to as sarcomas, or soft tissue
sarcomas. There are many types of soft tissue tumors, including
lipoma, lipoblastoma, hibemoma, liposarcoma, leiomyoma,
leiomyosarcoma, rhabdomyoma, rhabdomyosarcoma, neurofibroma,
schwannoma (neurilemoma), neuroma, malignant schwannoma,
neurofibrosarcoma, neurogenic sarcoma, nodular tenosynovitis,
synovial sarcoma, hemangioma, glomus tumor, hemangiopericytoma,
hemangioendothelioma, angiosarcoma, Kaposi sarcoma, lymphangioma,
fibroma, elastofibroma, superficial fibromatosis, fibrous
histiocytoma, fibrosarcoma, fibromatosis, dermatofibrosarcoma
protuberans (DFSP), malignant fibrous histiocytoma (MFH), myxoma,
granular cell tumor, malignant mesenchymomas, alveolar soft-part
sarcoma, epithelioid sarcoma, clear cell sarcoma, and desmoplastic
small cell tumor. In a particular embodiment, the soft tissue
cancer is a sarcoma selected from the group consisting of a
fibrosarcoma, a gastrointestinal sarcoma, a leiomyosarcoma, a
dedifferentiated liposarcoma, a pleomorphic liposarcoma, a
malignant fibrous histiocytoma, a round cell sarcoma, and a
synovial sarcoma.
[0071] The present teachings also provide methods of treating a
subject with a disease comprising administering to the subject an
effective amount of a compound represented by Structural Formula
(I) in combination with an effective immunomodulatory therapy (also
referred as immunotherapy). Immunotherapy is the treatment of
disease by using an immunomodulatory agent to induce, enhance, or
suppress an immune response. Immunotherapies designed to elicit or
amplify an immune response are classified as activation
immunotherapies, while immunotherapies that reduce or suppress are
classified as suppression immunotherapies. The disease described
herein is a cancer.
[0072] Immunomodulatory therapies, used alone or in combination
approaches, include i) immune checkpoint blockade inhibitors,
including but not limited to anti-CTLA4 (cytotoxic
T-lymphocyte-associated protein 4) antibodies (e.g. Ipilimumab),
agents that disrupt the PD-1/PD-L1 and PD-L2 interaction, e.g.
Nivolumab (Opdivo--Bristol Myers Squibb), Pembrolizumab (Keytruda,
KM-3475, Merck), Pidilizumab (CT-011, Cure Tech), BMS 936559 (BMS)
and MPDL3280A (Roche); and other immune response inhibitory
receptors e.g. anti-CD47; ii) cell based therapies (including, but
not limited to, dendritic cell therapy (e.g. Sipuleucel T
(Provenge) and adoptive T-cell therapies, iii) vaccination
strategies; iv) Adoptive T-cell therapy; v) agents that prevent
metabolic inhibition of the immune response, including inhibitors
of indoleamine 2,3-dioxygenase (e.g. INCB024360 (Incyte),
1-methyl-D-tryptophan, indoximod (NewLink Genetics)) or arginase;
and vi) cytokine-based therapy, e.g., interferons (in particular
type I interferon) and interleukins (e.g. interleukin-2).
[0073] In one embodiment, the immunomodulatory agent used for the
immunomodulatory therapy is a PD-1 inhibitor, for example, an
anti-PD1 antibody.
[0074] Programmed cell death protein 1, also known as PD-1 and
CD279 (cluster of differentiation 279), is a protein that in humans
is encoded by the PDCD1 gene. PD-1 is a cell surface receptor that
belongs to the immunoglobulin superfamily and is expressed on T
cells and pro-B cells. PD-1 binds two ligands, PD-L1 and PD-L2,
both of which are members of the B7 family.
[0075] PD-1 and its ligands play an important role in down
regulating the immune system by preventing the activation of
T-cells, which in turn reduces autoimmunity and promotes
self-tolerance. The inhibitory effect of PD-1 is accomplished
through a dual mechanism of promoting apoptosis (programmed cell
death) in antigen specific T-cells in lymph nodes while
simultaneously reducing apoptosis in regulatory T cells (suppressor
T cells).
[0076] The PD-1 inhibitor used in the present invention includes,
but is not limited to, nivolumab, pembrolizumab, pidilizumab, BMS
936559, MPDL3280A, MSB0010718C or MED14736. Among them, BMS 936559,
MPDL3280A, MSB0010718C, and MED14736 bind ligand PD-L1, all of
which are antibodies. Both nivolumab and pembrolizumab are approved
by the Food and Drug Administration for treatment of unresectable
or metastatic melanoma which no longer responds to other drugs.
[0077] Vaccination strategies include anti-microbial immunotherapy,
which includes vaccination, involves activating the immune system
to respond to an infectious agent.
[0078] Adoptive T-cell therapy uses T cell-based cytotoxic
responses to attack cancer cells. T cells that have a natural or
genetically engineered reactivity to a patient's cancer are
generated in vitro and then transferred back into the cancer
patient. One study using autologous tumor-infiltrating lymphocytes
was an effective treatment for patients with metastatic melanoma.
This can be achieved by taking T cells that are found with the
tumor of the patient, which are trained to attack the cancerous
cells. These T cells are referred to as tumor-infiltrating
lymphocytes (TIL) are then encouraged to multiply in vitro using
high concentrations of IL-2, anti-CD3 and allo-reactive feeder
cells. These T cells are then transferred back into the patient
along with exogenous administration of IL-2 to further boost their
anti-cancer activity.
[0079] The present teachings also provide methods of treating a
subject with a cancer comprising administering to the subject an
effective amount of a compound represented by Structural Formula
(I) in combination with an effective anti-cancer therapy. In one
embodiment, the cancer is a metastatic cancer. A "metastatic
cancer" is a cancer that has spread from its primary site to other
parts of the body.
[0080] The anti-cancer therapy described herein includes
co-administration of an effective amount of a second anti-cancer
agent together with a disclosed HPK-1 inhibitor. An "anti-cancer
agent" is a compound, which when administered in an effective
amount to a subject with cancer, can achieve, partially or
substantially, one or more of the following: arresting the growth,
reducing the extent of a cancer (e.g., reducing size of a tumor),
inhibiting the growth rate of a cancer, and ameliorating or
improving a clinical symptom or indicator associated with a cancer
(such as tissue or serum components) or increasing longevity of the
subject.
[0081] The anti-cancer agents suitable for use in the methods
described herein include any anti-cancer agents that have been
approved for the treatment of cancer. In one embodiment, the
anti-cancer agent includes, but is not limited to, a targeted
antibody, an angiogenesis inhibitor, an alkylating agent, an
antimetabolite, a vinca alkaloid, a taxane, a podophyllotoxin, a
topoisomerase inhibitor, a hormonal antineoplastic agent and other
antineoplastic agents. In one embodiment, the anti-cancer agent is
a PD-1 inhibitor, for example, an anti-PD1 antibody.
[0082] In one embodiment, the anti-cancer agents that can be used
in methods described herein include, but are not limited to,
paclitaxel, docetaxel, 5-fluorouracil, trastuzumab, lapatinib,
bevacizumab, letrozole, goserelin, tamoxifen, cetuximab,
panitumumab, gemcitabine, capecitabine, irinotecan, oxaliplatin,
carboplatin, cisplatin, doxorubicin, epirubicin, cyclophosphamide,
methotrexate, vinblastine, vincristine, melphalan, cytarabine,
etoposide, daunorubicin, bleomycin, mitomycin and adriamycin and a
combination thereof.
[0083] In one embodiment, the anti-cancer agent and the compound
represented by Structural Formula (I) are administered
contemporaneously. When administered contemporaneously, the
anti-cancer agent and the compound can be administered in the same
formulation or in different formulations. Alternatively, the
compound and the additional anti-cancer agent are administered
separately at different times.
[0084] As used herein, "treating a subject with a cancer" includes
achieving, partially or substantially, one or more of the
following: arresting the growth, reducing the extent of the cancer
(e.g., reducing size of a tumor), inhibiting the growth rate of the
cancer, ameliorating or improving a clinical symptom or indicator
associated with the cancer (such as tissue or serum components) or
increasing longevity of the subject; and reducing the likelihood of
recurrence of the cancer.
[0085] The term an "effective amount" means an amount when
administered to the subject which results in beneficial or desired
results, including clinical results, e.g., inhibits, suppresses or
reduces the cancer (e.g., as determined by clinical symptoms or the
amount of cancer cells) in a subject as compared to a control.
[0086] Generally, an effective amount of a compound taught herein
varies depending upon various factors, such as the given drug or
compound, the pharmaceutical formulation, the route of
administration, the type of disease or disorder, the identity of
the subject or host being treated, and the like, but can
nevertheless be routinely determined by one skilled in the art. An
effective amount of a compound of the present teachings may be
readily determined by one of ordinary skill by routine methods
known in the art.
[0087] In an embodiment, an effective amount of a compound taught
herein ranges from about 0.1 to about 1000 mg/kg body weight,
alternatively about 1 to about 500 mg/kg body weight. In another
embodiment, an effective amount of a compound taught herein ranges
from about 0.5 to about 5000 mg/m.sup.2, alternatively about from 5
to about 2500 mg/m.sup.2, and in another alternative from about 50
to about 1000 mg/m.sup.2. The skilled artisan will appreciate that
certain factors may influence the dosage required to effectively
treat a subject suffering from cancer or reduce the likelihood of
recurrence of a cancer. These factors include, but are not limited
to, the severity of the disease or disorder, previous treatments,
the general health and/or age of the subject and other diseases
present.
[0088] A "subject" is a mammal, preferably a human, but can also be
an animal in need of veterinary treatment, e.g., companion animals
(e.g., dogs, cats, and the like), farm animals (e.g., cows, sheep,
pigs, horses, and the like) and laboratory animals (e.g., rats,
mice, guinea pigs, and the like).
[0089] The compounds taught herein can be administered to a patient
in a variety of forms depending on the selected route of
administration, as will be understood by those skilled in the art.
The compounds of the present teachings may be administered, for
example, by oral, parenteral, buccal, sublingual, nasal, rectal,
patch, pump or transdermal administration and the pharmaceutical
compositions formulated accordingly. Parenteral administration
includes intravenous, intraperitoneal, subcutaneous, intramuscular,
transepithelial, nasal, intrapulmonary, intrathecal, rectal and
topical modes of administration. Parenteral administration can be
by continuous infusion over a selected period of time.
[0090] The compounds taught herein can be suitably formulated into
pharmaceutical compositions for administration to a subject. The
pharmaceutical compositions of the present teachings optionally
include one or more pharmaceutically acceptable carriers and/or
diluents therefor, such as lactose, starch, cellulose and dextrose.
Other excipients, such as flavoring agents; sweeteners; and
preservatives, such as methyl, ethyl, propyl and butyl parabens,
can also be included. More complete listings of suitable excipients
can be found in the Handbook of Pharmaceutical Excipients (5.sup.th
Ed., Pharmaceutical Press (2005)). A person skilled in the art
would know how to prepare formulations suitable for various types
of administration routes. Conventional procedures and ingredients
for the selection and preparation of suitable formulations are
described, for example, in Remington's Pharmaceutical Sciences
(2003--20th edition) and in The United States Pharmacopeia: The
National Formulary (USP 24 NF19) published in 1999. The carriers,
diluents and/or excipients are "acceptable" in the sense of being
compatible with the other ingredients of the pharmaceutical
composition and not deleterious to the recipient thereof.
[0091] Typically, for oral therapeutic administration, a compound
of the present teachings may be incorporated with excipient and
used in the form of ingestible tablets, buccal tablets, troches,
capsules, elixirs, suspensions, syrups, wafers, and the like.
[0092] Typically for parenteral administration, solutions of a
compound of the present teachings can generally be prepared in
water suitably mixed with a surfactant such as
hydroxypropylcellulose. Dispersions can also be prepared in
glycerol, liquid polyethylene glycols, DMSO and mixtures thereof
with or without alcohol, and in oils. Under ordinary conditions of
storage and use, these preparations contain a preservative to
prevent the growth of microorganisms.
[0093] Typically, for injectable use, sterile aqueous solutions or
dispersion of, and sterile powders of, a compound described herein
for the extemporaneous preparation of sterile injectable solutions
or dispersions are appropriate.
[0094] For nasal administration, the compounds of the present
teachings can be formulated as aerosols, drops, gels and powders.
Aerosol formulations typically comprise a solution or fine
suspension of the active substance in a physiologically acceptable
aqueous or non-aqueous solvent and are usually presented in single
or multi-dose quantities in sterile form in a sealed container,
which can take the form of a cartridge or refill for use with an
atomizing device. Alternatively, the sealed container may be a
unitary dispensing device such as a single dose nasal inhaler or an
aerosol dispenser fitted with a metering valve which is intended
for disposal after use. Where the dosage form comprises an aerosol
dispenser, it will contain a propellant which can be a compressed
gas such as compressed air or an organic propellant such as
fluorochlorohydrocarbon. The aerosol dosage forms can also take the
form of a pump-atomizer.
[0095] For buccal or sublingual administration, the compounds of
the present teachings can be formulated with a carrier such as
sugar, acacia, tragacanth, or gelatin and glycerine, as tablets,
lozenges or pastilles.
[0096] For rectal administration, the compounds described herein
can be formulated in the form of suppositories containing a
conventional suppository base such as cocoa butter.
[0097] The compounds of invention may be prepared by methods known
to those skilled in the art, as illustrated by the general schemes
and procedures below and by the preparative examples that follow.
All starting materials are either commercially available or
prepared by methods known to those skilled in the art and the
procedures described below.
[0098] General synthetic approaches to the claims compounds are
provided in the exemplification below, as illustrated in Schemes 1
and 2.
EXEMPLIFICATION
Example A: Synthesis
General Methods
[0099] Commercially available starting materials, reagents, and
solvents were used as received. In general, anhydrous reactions
were performed under an inert atmosphere such as nitrogen or Argon.
PoraPak.RTM. Rxn CX refers to a commercial cation-exchange resin
available from Waters.
[0100] Microwave reactions were performed with a Biotage Initiator
microwave reactor. Reaction progress was generally monitored by
LCMS (Bruker Exquire 4000 or Waters Acquity UPLC system). Flash
column chromatographic purification of intermediates or final
products was performed using a Biotage Isolera with KP-SIL or
HP-SIL silica cartridges, or KP-NH basic modified silica and
corresponding samplets. Reverse-phase HPLC purification was
performed on a Varian PrepStar model SD-1 HPLC system with a Varian
Monochrom 10.mu. C-18 reverse-phase column using a gradient of 10%
MeOH/0.05% TFA-H.sub.2O to 90% MeOH/0.05% TFA in H.sub.2O over a
40-min period at a flow rate of 40 mL/min. Reverse phase
purification was also performed using a Biotage Isolera equipped
with a KP-C.sub.18-H column using a between 10-95% MeOH or
CH.sub.3CN/0.1% TFA in H.sub.2O. Proton NMRs were recorded on a
Bruker 400 MHz spectrometer, and mass spectra were obtained using a
Bruker Esquire 4000 spectrometer or Waters Acquity UPLC system.
[0101] Compound names were generated using the software built into
CambridgeSoft-PerkinElmer's ChemBioDraw Ultra version 12.0.
Abbreviations
[0102] aq aqueous [0103] anh anhydrous [0104] Ar argon [0105] Boc
tert-butoxycarbonyl [0106] br. broad [0107] calcd calculated [0108]
d doublet (only when used within 1H NMR spectra) [0109] DCM
dichloromethane [0110] de diastereomeric excess [0111] DIPEA
diisopropylethylamine [0112] DMF N,N-dimethylformamide [0113] DMSO
dimethylsulfoxide [0114] dppf 1,1'-bis(diphenylphosphino) ferrocene
[0115] equiv equivalent [0116] Flt3 fins-related tyrosine kinase 3
[0117] h hour [0118] HPK1 hematopoietic progenitor kinase 1 [0119]
HPLC high performance liquid chromatography [0120] IPA isopropanol
[0121] KHMDS potassium hexamethyldisilazide [0122] Lck
lymphocyte-specific protein tyrosine kinase [0123] LC-MS liquid
chromatography coupled to mass spectrometry [0124] LDA lithium
diisopropyllamide [0125] LiHMDS lithium hexamethyldisilazide [0126]
min minute [0127] m multiplet [0128] MeCN acetonitrile [0129] MS
ESI mass spectra, electrospray ionization [0130] NMR nuclear
magnetic resonance [0131] O/N overnight [0132] PMB
para-methoxybenzyl [0133] prep preparative [0134] rt room
temperature [0135] Rt retention time [0136] RP reverse phase [0137]
s singlet [0138] satd saturated [0139] t triplet [0140] temp.
temperature [0141] TFA trifluoroacetic acid [0142] THF
tetrahydrofuran
##STR00007##
##STR00008##
[0142] Preparation of Starting Materials
General Method A1 (Base-Induced Cyclization Using Benzimidazole
Ester)
[0143] A solution of aryl oxazine-2,4-dione (1 equiv), or aminoaryl
nitrile and substituted 1H-benzo[d]imidazol-2-yl)acetate (1-1.2
equiv) in THF was treated with KHMDS, LiHMDS, or LDA (3-5 equiv).
The reaction was stirred at 45.degree. C. for 4-24 h. The reaction
was then cooled to rt and quenched with satd aq NH.sub.4Cl. The
aqueous layer was extracted with EtOAc or DCM, and the combined
organic extracts were dried over MgSO.sub.4, filtered and
concentrated. Crude product was purified by column chromatography
or prep-HPLC to give the desired product.
General Method A2 (Two-Step, Base-Induced Cyclization Using
Benzimidazole Ester)
[0144] A solution of aryl oxazine-2,4-dione (1 equiv), or aminoaryl
nitrile and substituted 1H-benzo[d]imidazol-2-yl)acetate (1-1.2
equiv) in was treated with KHMDS, LiHMDS, KOBu.sup.t or LDA (3-5
equiv) at 45.degree. C. for 2-4 h. The reaction was then cooled to
rt and quenched with satd aq NH.sub.4Cl. The aqueous layer was
extracted with EtOAc or DCM, and the combined organic extracts were
dried over MgSO.sub.4, filtered and concentrated. The uncyclized
addition adduct was separated by column chromatography, dissolved
in THF and treated with KHMDS, LiHMDS, or LDA (3-5 equiv). The
reaction was stirred at 45.degree. C. for 1-4 h. The reaction was
then cooled to rt and quenched with satd aq NH.sub.4Cl. The aqueous
layer was extracted with EtOAc or DCM, and the combined organic
extracts were dried over MgSO.sub.4, filtered and concentrated.
Crude product was purified by column chromatography or prep-HPLC to
give the desired product.
General Method A3 (Two-Step, Base-Induced Cyclization Using
Benzimidazole Ester)
[0145] A solution of aminoaryl nitrile and substituted
1H-benzo[d]imidazol-2-yl)acetate (1 equiv) in THF was treated with
LiHMDS, or LDA (5 equiv) (step 1). The reaction was stirred at
35-40.degree. C. for 1-1.5 h. The reaction was then cooled to rt
and quenched with satd aq NH.sub.4Cl and concentrated. Crude
product was purified by prep-HPLC to give uncyclized intermediate
that was neutralized, dried and subjected to the conditions
described in general method A1 using LiHMDS (step 2).
General Method B (Triflate Formation)
[0146] A solution of benzimidazol-2-yl arylpyridinone derivate (1
equiv) and pyridine (20 equiv) in DCM was treated with Tf.sub.2O (8
equiv). The reaction was stirred at 0.degree. C. for 2-8 h. The
reaction was then quenched with satd aq NaHCO.sub.3. The aqueous
layer was extracted with DCM, and the combined organic extracts
were dried over MgSO.sub.4, filtered and concentrated. Crude
product was used in the next step without further purification.
General Method C (Amine Substitution)
[0147] A solution of benzoimidazol-2-yl arylpyridinone bistriflate
derivate (1 equiv) in MeCN, DCM, or DMF was treated with amine
(1.2-3 equiv). In the case where the amine is a salt (e.g. HCl),
the amine salt was dissolved in MeOH or DMF and passed through a
PoraPak Rxn CX ion exchange column to yield the free base which was
added to the reaction mixture. The reaction mixture was stirred at
rt or up to 45.degree. C. for 1-48 h. Solvent was removed and the
crude product was purified by column chromatography or prep-HPLC to
give the desired product.
General Method D (Global Deprotection)
[0148] A solution of protected benzoimidazol-2-yl arylpyridinone
derivate (1 equiv) in TFA/conc. HCl (7:1 v/v) was heated at
80-100.degree. C. for 3-24 h. Solvent was removed and the crude
product was purified by column chromatography (free base) or
prep-HPLC (TFA salt) to give the desired product. To generate the
desired product as a HCl salt, the free base was dissolved in MeOH
and 1 M HCl-Et.sub.2O (2-4 equiv) was added at rt. The solution was
stirred for 5 min and azeotroped twice with MeOH.
General Method E (PMB-Protection)
[0149] A solution of thiaisatoic anhydride (1 equiv),
1-(chloromethyl)-4-methoxybenzene (1-1.2 equiv), K.sub.2CO.sub.3
(1-1.2 equiv) and/or KI (1-1.2 equiv) in DMF was stirred at rt for
4-24 h. The reaction mixture was then slowly added to H.sub.2O,
precipitate was collected by vacuum filtration to give the
desired.
Intermediates
1H-thieno[3,4-d][1,3]oxazine-2,4-dione
##STR00009##
[0151] To a solution of
4-tert-butoxycarbonylamino-thiophene-3-carboxylic acid (2.5 g, 10.2
mmol) in PhMe (25 mL) was added oxalyl chloride (1.29 mL, 15.3
mmol) at rt. The reaction mixture was gradually heated to
95.degree. C. and stirred at 95.degree. C. for 1 h. After reaction
completion, the reaction was cooled to rt and filtered. The solid
was washed with hexanes (2.times.5 mL), and dried under vacuum to
afford the title compound as a cream solid (1.61 g, 93%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 11.57 (s, 1H), 8.64 (d, J=3.2
Hz, 1H), 6.89 (d, J=2.8 Hz, 1H); MS ESI [M+H].sup.+ 170.0, calcd
for [C.sub.6H.sub.3NO.sub.3S+H].sup.+ 169.9.
1-(4-methoxybenzyl)-1H-thieno[3,4-d][1,3]oxazine-2,4-dione
##STR00010##
[0153] According to general method E, to a solution of
1H-thieno[3,4-d][1,3]oxazine-2,4-dione (1.6 g, 9.45 mmol) in anh
DMF (20 mL), K.sub.2CO.sub.3 (1.56 g, 11.3 mmol) was added followed
by KI (0.62 g, 3.78 mmol) under stirring at rt. PMBCl (1.54 mL,
11.3 mmol) was added dropwise over 10 min and the reaction mixture
was stirred for a further 2 h. After reaction completion the
reaction mixture was poured into H.sub.2O (200 mL) to precipitate
the product which was filtered, washed with H.sub.2O and dried to
afford the title compound as an off-white solid (2.3 g, 84%).
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.35 (d, J=3.2 Hz, 1H),
7.31 (d, J=8.8 Hz, 2H), 6.89 (d, J=8.8 Hz, 2H), 6.62 (d, J=3.2 Hz,
1H), 5.08 (s, 2H), 3.80 (s, 3H); MS ESI [M+H].sup.+ 291.2, calcd
for [C.sub.14H.sub.11NO.sub.4S+H].sup.+ 290.0.
7-hydroxy-4-(4-methoxybenzyl)-6-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imi-
dazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one
##STR00011##
[0155] According to general method A1, to a solution of
1-(4-methoxybenzyl)-1H-thieno[3,2-d][1,3]oxazine-2,4-dione
[Tetrahedron (1999) 55 6167-6174] (2.89 g, 10 mmol), ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate [J.
Med. Chem. (2009), 52, 278-292] (3.02 g, 10 mmol), LiHMDS (1 M in
THF, 4 mL, 4 mmol) were used to generate the title compound as an
orange solid (2.65 g, 51%). .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 13.68 (br.s., 1H), 12.57 (s, 1H), 7.55 (dd, J=5.2, 2.0 Hz,
1H), 7.40-7.32 (m, 1H), 7.23 (d, J=8.8 Hz, 2H), 7.04-6.93 (m, 3H),
6.85 (d, J=8.8 Hz, 2H), 5.37 (s, 2H), 3.77 (s, 3H), 3.30-3.19 (m,
4H), 2.69-2.58 (m, 4H), 2.39 (s, 3H); MS ESI [M+H].sup.+ 502.1,
calcd for [C.sub.27H.sub.27N.sub.5O.sub.3S+H].sup.+ 502.2.
4-hydroxy-7-(4-methoxybenzyl)-5-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imi-
dazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one
##STR00012##
[0157] According to general method A2, a solution of
1-(4-methoxybenzyl)-1H-thieno[2,3-d][1,3]oxazine-2,4-dione (0.40 g,
1.4 mmol), ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(0.46 g, 1.5 mmol), and LDA (1 M in THF, 6.2 mL, 4.5 mmol) were
used to generate the title compound as a brown solid (0.220 g,
32%). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 13.87 (br.s., 1H),
12.52 (s, 1H), 7.49 (dd, J=14.9 Hz, 1H), 7.40-7.24 (m, 3H),
7.03-6.64 (m, 5H), 5.28 (d, J=13.8 Hz, 2H), 3.76 (s, 3H), 3.21 (d,
J=18.8 Hz, 4H), 2.65 (m, d, J=19.1 Hz, 4H), 2.41 (s, 3H); MS ESI
[M+H].sup.+ 502.3, calcd for
[C.sub.27H.sub.27N.sub.5O.sub.3S+H].sup.+ 502.2.
TABLE-US-00001 7-hydroxy-4-(4- methoxybenzyl)-6-(6- morpholino-1H-
benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin-5(4H)- one ##STR00013##
325 mg (48%) Brown solid; free base Reagents (General Method A1):
1-(4-methoxybenzyl)-1H-thieno[3,2-d][1,3]oxazine-2,4-dione
[Tetrahedron (1999) 55 6167-6174] (0.4 g, 1.4 mmol), ethyl
2-(6-morpholino-1H- benzo[d]imidazol-2-yl)acetate [J.Med.Chem.
(2009), 52, 278-292] (0.4 g, 1.4 mmol), LiHMDS (5.5 mL, 5.5 mmol)
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 13.73-13.60 (m, 1H),
12.64-12.52 (m, 1H), 7.54 (d, J = 5.3 Hz, 1H), 7.42-7.29 (m, 2H),
7.21 (d, J = 7.8 Hz, 2H), 7.03-6.89 (m, 2H), 6.85 (d, J = 9.0 Hz,
2H), 5.37 (br. s, 2H), 3.83-3.98 (m, 4H), 3.77 (s, 3H), 3.23-3.09
(m, 4H); MS ESI [M + H].sup.+ 489.2, calcd for
[C.sub.26H.sub.24N.sub.4O.sub.4S + H].sup.+ 489.2. 4-hydroxy-7-(4-
methoxybenzyl)-5-(6- morpholino-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00014## 445 mg (24%) brown
solid; Free base Reagents (General Method A1):
1-(4-methoxybenzyl)-1H-thieno[2,3-d][1,3]oxazine-2,4-dione (0.40 g,
1.4 mmol), ethyl 2-(6-morpholino-1H-benzo[d]imidazol-2-yl)acetate
(0.4 g, 1.4 mmol), LDA (17 mL, 17 mmol). MS ESI [M + H].sup.+
489.2, calcd for [C.sub.26H.sub.24N.sub.4O.sub.4S + H].sup.+
489.1.
Ethyl
3-(4-((4-methoxybenzyl)amino)thiophen-3-yl)-2-(6-(4-methylpiperazin--
1-yl)-1H-benzo[d]imidazol-2-yl)-3-oxopropanoate
##STR00015##
[0159] To a solution of ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(2.58 g, 8.55 mmol) and 1-(4-methoxy
benzyl)-1H-thieno[3,4-d][1,3]oxazine-2,4-dione (2.46 g, 8.55 mmol)
in anh THF (48 mL), 1 M LDA (34 mL, 1 M in THF/hexane, 34 mmol) was
added dropwise at 40.degree. C. under Ar. The resulting brown
solution was stirred at 40.degree. C. for 1 h and then quenched
with aq NH.sub.4Cl (50 mL) at rt. The reaction mixture was diluted
with H.sub.2O (50 mL) and extracted with DCM (2.times.200 mL). The
combined organic layers were washed once with H.sub.2O, dried over
Na.sub.2SO.sub.4, and concentrated to give crude ester. The crude
product was purified by flash chromatography (gradient: EtOAc/hex
0-40%, followed by MeOH/DCM 0-25%) to give the title compound as a
light brown solid (3.05 g, 65%). MS ESI [M+H].sup.+ 548.2, calcd
for [C.sub.29H.sub.33N.sub.5O.sub.4S+H].sup.+ 548.2.
4-hydroxy-1-(4-methoxybenzyl)-3-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imi-
dazol-2-yl)thieno[3,4-b]pyridin-2(1H)-one
##STR00016##
[0161] Ethyl
3-(4-((4-methoxybenzyl)amino)thiophen-3-yl)-2-(6-(4-methylpiperazin-1-yl)-
-1H-benzo[d]imidazol-2-yl)-3-oxopropanoate (3.05 g, 5.57 mmol),
described above, was dissolved in anh THF (30 mL) at rt under Ar. A
solution of LDA (16.8 mL, 1 M in THF/hexane, 16.71 mmol) was added
dropwise at 40.degree. C. The resulting brown solution was stirred
at 40.degree. C. for 1 h and then quenched with aq NH.sub.4Cl (25
mL) at rt. The mixture was diluted with H.sub.2O (25 mL) and
extracted with DCM (2.times.250 mL). The combined organic layers
were washed once with H.sub.2O, dried over Na.sub.2SO.sub.4, and
concentrated to give crude product. The crude product was purified
by flash chromatography (gradient: MeOH/DCM 0-20%) to give the
title compound as a light brown solid (1.81 g, 65%). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 13.8-13.25 (m, 1H), 8.13 (d, J=3.6
Hz, 1H), 7.58 (d, J=8.8 Hz, 1H), 7.36-7.29 (m, 3H), 7.06-7.02 (m,
1H), 6.97 (d, J=3.6 Hz, 1H), 6.86 (d, J=8.4 Hz, 2H), 5.19 (s, 2H),
3.69 (s, 3H), 3.16 (br.s, 4H), 2.60 (br.s, 4H), 2.31 (s, 3H); a
signal due to OH group cannot be readily detected. MS ESI 502.1
[M+H].sup.+, calcd for [C.sub.27H.sub.27N.sub.5O.sub.3S+H].sup.+
502.2.
7-(4-methoxybenzyl)-5-(5 and/or
6)-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imid-
azol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate
##STR00017##
[0163] Synthesized according to general method B using
4-hydroxy-7-(4-methoxybenzyl)-5-(6-(4-methylpiper-azin-1-yl)-1H-benzo[d]i-
midazol-2-yl)thieno[2,3-b]py-ridin-6(7H)-one (0.22 g, 0.44 mmol),
Tf.sub.2O (0.60 mL, 3.5 mmol), and pyridine (0.72 mL, 8.8 mmol).
The title compounds obtained as an indeterminate mixture of
regioisomers, were used in the next step without purification. MS
ESI [M+H].sup.+ 766.1, calcd for
[C.sub.29H.sub.25F.sub.6N.sub.5O.sub.7S.sub.3+H].sup.+ 766.1.
1-(4-methoxybenzyl)-3-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]-imid-
azol-2-yl)-2-oxo-1,2-dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate
##STR00018##
[0165] According to general method B, a solution of
4-hydroxy-1-(4-methoxybenzyl)-3-(6-(4-methylpiperazin-1-yl)-1H-benzo
[d]imidazol-2-yl)thieno[3,4-b]pyridin-2(1H)-one (220 mg, 0.43 mmol)
and pyridine (708 mL, 8.76 mmol) in DCM (12 mL) was added Tf.sub.2O
(558 mL, 3.50 mmol) at -5.degree. C. The reaction was stirred
between -5 and 0.degree. C. for 1 h. The reaction was quenched with
satd aq NaHCO.sub.3. The aqueous layer was extracted with DCM, and
the combined organic extracts were dried over Na.sub.2SO.sub.4, and
concentrated under vacuum to give dark brown oil. The crude
product, obtained as an indeterminate mixture of regioisomers, was
used directly in the next step without further purification. MS ESI
[M+H].sup.+ 766.0, calcd for
[C.sub.29H.sub.25F.sub.6N.sub.5O.sub.7S.sub.3+H].sup.+ 766.1.
7-(4-methoxybenzyl)-5-(5 and/or
6-morpholino-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-ox-
o-6,7-dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate
##STR00019##
[0167] According to general method B, a solution of
4-hydroxy-7-(4-methoxybenzyl)-5-(6-morpholino-1H-benzo[d]imidazol-2-yl)-t-
hieno[2,3-b]pyridin-6(7H)-one (200 mg, 0.41 mmol) and pyridine
(0.66 mL, 8.2 mmol) in DCM (20 mL) was added Tf.sub.2O (0.55 mL,
3.28 mmol) at 0.degree. C. The reaction mixture was stirred at
0.degree. C. for 2 h and then quenched with satd aq NaHCO.sub.3.
The aqueous layer was extracted with DCM. The combined organic
extracts were dried over Na.sub.2SO.sub.4, and concentrated to give
the crude title compound (mixture of two regioisomers) as brown oil
which was used directly in the subsequent step without further
purification considering quantitative yield. MS ESI [M+H].sup.+
753.0, calcd for
[C.sub.28H.sub.22F.sub.6N.sub.4O.sub.8S.sub.3+H].sup.+ 752.9.
5-methyl-1H-thieno[2,3-d][1,3]oxazine-2,4-dione
##STR00020##
[0169] To a solution of KOH (0.49 g, 8.76 mmol) in H.sub.2O (20 mL)
was added methyl 2-amino-4-methyl-3-thiophene carboxylate (1.0 g,
5.84 mmol) at rt. The resulting reaction was heated to 90.degree.
C. for 2 h and then cooled to 0.degree. C. A solution of
triphosgene (0.866 g, 2.92 mmol) in PhMe (12 mL) was added dropwise
over 10 min. The resulting solution was gradually warmed to rt and
stirred for 2 h. The resulting solid was filtered, washed with
H.sub.2O and dried to afford the title compound as a light pink
solid (0.65 g, 61%). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 6.65
(d, J=1.2 Hz, 1H), 2.42 (d, J=1.2 Hz, 3H); MS ESI [M+H]f 184.0,
calcd for [C.sub.7H.sub.5NO.sub.3S+H].sup.+ 184.0.
1-(4-methoxybenzyl)-5-methyl-1H-thieno[2,3-d][1,3]oxazine-2,4-dione
##STR00021##
[0171] To a solution of
5-methyl-1H-thieno[2,3-d][1,3]oxazine-2,4-dione (0.625 g, 3.41
mmol) in anh DMF (9 mL), K.sub.2CO.sub.3 (0.566 g, 4.09 mmol) was
added followed by KI (0.142 g, 0.85 mmol) under stirring at rt.
PMB-Cl (0.56 mL, 4.06 mmol) was added dropwise to the reaction over
10 min and stirred for further 2 h. The reaction mixture was poured
into H.sub.2O (100 mL) to precipitate the product which was
filtered, washed with H.sub.2O and dried to afford the title
compound as a light brown solid (0.935 g, 91%). .sup.1H NMR (400
MHz, CDCl.sub.3) .delta. 7.38 (d, J=8.8 Hz, 2H), 6.90-6.88 (m, 2H),
6.46 (d, J=1.2 Hz, 1H), 5.05 (s, 2H), 3.80 (s, 3H), 2.42 (d, J=1.2
Hz, 3H); MS ESI [M+H].sup.+ 304.2, calcd for
[C.sub.15H.sub.13NO.sub.4S+H].sup.+ 304.1.
4-hydroxy-7-(4-methoxybenzyl)-3-methyl-5-(6-(4-methylpiperazin-1-yl)-1H-be-
nzo[d]imidazol-2-yl) thieno[2,3-b]pyridin-6(7H)-one
##STR00022##
[0173] A solution of LDA (34 mL, 1 M in THF/hexane, 34 mmol) was
added dropwise to a solution of ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate (922
mg, 3.04 mmol) and 1-(4-methoxy
benzyl)-5-methyl-1H-thieno[2,3-d][1,3]oxazine-2,4-dione (925 mg,
3.04 mmol) in anh THF (28 mL) at 40.degree. C. under Ar. The
resulting brown solution was stirred at 40.degree. C. for 2 h and
then quenched with aq, NH.sub.4Cl (25 mL) at rt. The reaction
mixture was diluted with H.sub.2O (25 mL) and extracted with DCM
(2.times.100 mL). The combined organic layers were washed once with
H.sub.2O, dried over Na.sub.2SO.sub.4, and concentrated to give
mixture of product and uncyclized ester. The crude mass was
purified by flash chromatography (gradient: EtOAc/hex 0-40%,
followed by MeOH/DCM 0-25%) to give mixture of product and
uncyclized ester (900 mg).
[0174] Above mixture of product and uncyclized ester (900 mg) was
dissolved in anh THF (9 mL) at rt under Ar. A solution of LDA (5
mL, 1 M in THF/hexane) was added dropwise at 40.degree. C. The
resulting brown solution was stirred at 40.degree. C. for 1 h and
worked up it as per above to give crude product. The crude product
was purified by flash chromatography (gradient: MeOH/DCM 0-20%) to
give the title compound as a cream solid (325 mg, 21%). .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 12.54 (s, 1H), 7.38 (d, J=8.4 Hz,
1H), 7.33 (d, J=8.4 Hz, 2H), 7.01-6.98 (m, 2H), 6.85 (d, J=8.8 Hz,
2H), 6.40 (s, 1H), 5.26 (s, 2H), 3.80-3.61 (m, 6H), 3.60-3.51 (m,
4H), 2.89-2.88 (m, 4H), 2.63 (s, 3H); the signal due to OH group
cannot be readily detected. MS ESI [M+H].sup.+ 516.2, calcd for
[C.sub.28H.sub.29N.sub.5O.sub.3S+H].sup.+ 516.2.
7-(4-methoxybenzyl)-3-methyl-5-(5 and/or
6)-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imid-
azol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate
##STR00023##
[0176] The title compound was prepared according to general method
B by utilizing
4-hydroxy-7-(4-methoxy-benzyl)-3-methyl-5-(6-(4-methylpiper-azi-
n-1-yl)-1H-benzo[d]imidazol-2-yl) thieno[2,3-b]pyridin-6(7H)-one
(320 mg, 0.62 mmol), pyridine (1.0 mL, 12.4 mmol), Tf.sub.2O (0.833
mL, 4.96 mmol) in DCM (12 mL) to give a dark brown oil. The crude
product, obtained as an indeterminate mixture of 2 regioisomers,
was used directly in the next step without further purification. MS
ESI [M+H].sup.+ 780.0, calcd for
[C.sub.30H.sub.27F.sub.6N.sub.5O.sub.7S.sub.3+H].sup.+ 780.1.
Ethyl
2-(6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-benzo[d]imidazol-
-2-yl)acetate
A. 2-nitro-5-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)aniline
##STR00024##
[0177] A mixture of 5-chloro-2-nitroaniline (1.73 g, 10 mmol),
(3r,5s)-rel-1,2,6-trimethylpiperazine (1.41 g, 11 mmol) and
K.sub.2CO.sub.3 (2.72 g, 20 mmol) was irradiated in microwave at
140.degree. C. for 4 h. H.sub.2O (150 mL) was then added with
stirring, suction filtered, rinsed with H.sub.2O and dried to give
the title compound as a brown solid (2.47 g, 94%). .sup.1H NMR (400
MHz, DMSO-d.sub.6) .delta. 7.79 (d, J=10.0 Hz, 1H), 7.23 (s, 2H,
NH.sub.2), 6.41 (dd, J=9.6, 1.6 Hz, 1H), 6.20 (d, J=2.4 Hz, 1H),
3.77 (d, J=12.4 Hz, 2H), 2.59 (t, J=11.8 Hz, 2H), 2.19-2.11 (m,
2H), 2.16 (s, 3H), 1.05 (d, J=6.0 Hz, 6H); MS ESI [M+H].sup.+
265.3, calcd for [C.sub.13H.sub.20N.sub.4O.sub.2+H].sup.+
265.2.
B.
4-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)benzene-1,2-diamine
##STR00025##
[0179] To a suspension of
2-nitro-5-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)aniline (2.47
g, 9.4 mmol) in MeOH (30 mL) was added 10% Pd/C (247 mg, 10% wt.).
The resulting mixture was hydrogenated under H.sub.2 balloon O/N.
After additional 10% Pd/C (124 mg, 5% wt.) was added, it was
hydrogenated under H.sub.2 balloon O/N, filtered, concentrated and
dried to give the title compound as a dark brown solid (2.25 g,
quantitative). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 6.66 (d,
J=8.4 Hz, 1H), 6.47 (d, J=2.4 Hz, 1H), 6.31 (dd, J=8.4, 2.8 Hz,
1H), 3.35-3.25 (m, 2H), 2.47-2.40 (m, 4H), 2.34 (s, 3H), 1.18 (d,
J=5.6 Hz, 6H); MS ESI [M+H].sup.+ 235.3, calcd for
[C.sub.13H.sub.22N.sub.4+H].sup.+ 235.2.
C. ethyl
2-(6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-benzo[d]imida-
zol-2-yl)acetate
##STR00026##
[0181] To a solution of
4-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)benzene-1,2-diamine
(2.25 g, 9.4 mmol) in EtOH (40 mL) was added ethyl
3-ethoxy-3-iminopropionate hydrochloride (2.93 g, 15 mmol). The
resulting mixture was heated at 80.degree. C. for 2 h. After
removal of solvents, it was diluted with DCM/MeOH (100 mL/10 mL),
basified with satd aq NaHCO.sub.3 (30 mL) and separated. The
aqueous layer was extracted with DCM (60 mL.times.2) and the
combined extracts were concentrated and purified by flash
chromatography (gradient: 100% EtOAc, then MeOH/DCM 0-20%) to give
the title compound as a dark orange solid (2.32 g, 73%). .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. 10.13 (br s, 1H, NH), 7.53-6.88
(m, 3H), 4.25 (q, J=7.2 Hz, 2H), 4.03 (s, 2H), 3.43 (d, J=11.2 Hz,
2H), 2.61 (t, J=11.2 Hz, 2H), 2.50-2.41 (m, 2H), 2.35 (s, 3H), 1.32
(t, J=7.2 Hz, 3H), 1.19 (d, J=6.0 Hz, 6H); MS ESI [M+H].sup.+
331.3, calcd for [C.sub.18H.sub.26N.sub.4O.sub.2+H].sup.+
331.2.
7-hydroxy-4-(4-methoxybenzyl)-6-(6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-
-yl)-1H-benzo[d]imidazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one
##STR00027##
[0183] To a mixture of
1-(4-methoxybenzyl)-1H-thieno[3,2-d][1,3]oxazine-2,4-dione (1.16 g,
4 mmol) and ethyl
2-(6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl-
)acetate (990 mg, 3 mmol) in THF (20 mL) was added LDA (1.0 M in
THF/hex, 10 mL, 10 mmol) dropwise at rt. After addition, the
resulting mixture was stirred at 40.degree. C. for 1 h, diluted
with DCM, quenched with satd aq NH.sub.4Cl and extracted with DCM.
The combined extracts were concentrated and purified by flash
chromatography (gradient: 20-100% EtOAc/hex, then MeOH (0.5%
NH.sub.3)/DCM 0-20%) to give a mixture of cyclized and uncyclized
product as a brown foam (1.10 g). The mixture was redissolved in
THF (15 mL) and LDA (1.0 M in THF/hex, 6 mL, 6 mmol) was added
dropwise at rt. The process and workup were both the same as above.
The title compound was obtained as an orange solid (630 mg, 40%).
MS ESI [M+H].sup.+ 530.3, calcd for
[C.sub.29H.sub.31N.sub.5O.sub.3S+H].sup.+ 530.2.
4-(4-methoxybenzyl)-5-oxo-6-(1-((trifluoromethyl)sulfonyl)-(5
and/or
6)-((3r,5s)-rel-3,4,5-trimethyl-piperazin-1-yl)-1H-benzo[d]imidazol-2-yl)-
-4,5-dihydrothieno[3,2-b]pyridin-7-yl trifluoromethane
sulfonate
##STR00028##
[0185] According to general method B, a solution of
7-hydroxy-4-(4-methoxybenzyl)-6-(6-((3r,5s)-rel-3,4,5-trimethyl-piperazin-
-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one (106
mg, 0.2 mmol) in DCM (15 mL) at 0.degree. C. was added pyridine
(0.32 mL, 4 mmol), followed by Tf.sub.2O (0.27 mL, 1.2 mmol). The
resulting mixture was stirred at 0.degree. C. for 1 h, diluted with
DCM (10 mL), quenched with satd aq NaHCO.sub.3 (15 mL), extracted
with DCM (20 mL.times.2) and concentrated to give the crude title
compound (an indeterminate mixture of two regioisomers) as a brown
oil which was used directly in the subsequent steps. MS ESI
[M+H].sup.+ 794.1, calcd for
[C.sub.31H.sub.29F.sub.6N.sub.5O.sub.7S.sub.3+H].sup.+ 794.11.
Synthesis of 2-amino-4-ethoxythiophene-3-carbonitrile
##STR00029##
[0187] The mixture of MeC(OMe).sub.3 (2.26 mL, 12.3 mmol) and
CH.sub.2(CN).sub.2 (0.78 mL, 12.3 mmol) was stirred at 65.degree.
C. for 3 h before cooled down to rt. THF (10 mL) and sulfur (395
mg) was added followed by addition of Et.sub.3N (1.72 mL, 12.3
mmol) dropwise. The resulting reaction mixture was stirred at
60.degree. C. for 15 min and concentrated under reduced pressure.
The residue was partitioned between EtOAc and H.sub.2O, extracted
with EtOAc, dried over Na.sub.2SO.sub.4, filtered, and concentrated
to dryness. The residue was triturated with DCM and filtered to
give the title compound as a brown solid (1.23 g, 60%). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 5.27 (s, 1H), 3.99 (q, J=7.0 Hz, 2H),
1.38 (t, J=7.0 Hz, 3H); MS ESI [M+H].sup.+ 169.0, calcd for
[C.sub.7H.sub.8N.sub.2OS+H].sup.+ 169.0.
tert-Butyl
4-(5-amino-4-cyanothiophen-3-yl)piperazine-1-carboxylate
##STR00030##
[0189] A mixture of MeC(OMe).sub.3 (1.3 mL, 10 mmol) and
CH.sub.2(CN).sub.2 (0.66 g, 10 mmol) was heated in closed vial at
80.degree. C. for 17 h. The reaction was cooled to rt and
tert-butyl piperazine-1-carboxylate (2.79 g, 15.0 mmol) was added.
Heating with stirring was continued at 65.degree. C. for 5 h. The
reaction mixture was then concentrated in vacuo. S.sub.8 (0.34 g)
and anh THF (10 mL) were added. The suspension was heated with
stirring at 40.degree. C. Et.sub.3N (1.3 mL, 9.3 mmol) was added
dropwise over 15 min. The oil bath temperature was increased to
60.degree. C. and stirring was continued for 11 h. The reaction was
then concentrated under reduced pressure and purified by flash
chromatography (SiO.sub.2, hexanes:EtOAc 5-50%) to afford
tert-butyl 4-(5-amino-4-cyanothiophen-3-yl)piperazine-1-carboxylate
as a light orange solid (0.71 g, 23%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 7.12 (s, 2H), 5.46 (s, 1H), 3.45-3.37 (m,
4H), 2.90-2.81 (m, 4H), 1.40 (s, 9H). MS ESI [M+H].sup.+ 309.3,
calcd for [C.sub.14H.sub.20N.sub.4O.sub.2S+H].sup.+ 309.1.
tert-butyl
4-(4-amino-5-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7-dihydrothieno-[2,3-b]pyridin-3-yl)piperazine-1-carboxylate
##STR00031##
[0191] LiHMDS (1.0 M in THF, 2.8 mL, 2.8 mmol) was added dropwise
at rt to a stirred suspension of ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(0.170 g, 0.56 mmol) and tert-butyl
4-(5-amino-4-cyanothiophen-3-yl)piperazine-1-carboxylate (0.175 g,
0.56 mmol) in anh THF (10 mL) under Ar. The reaction was stirred at
rt for additional 5 min and then heated in an oil bath at
40.degree. C. for 1 h. The reaction was cooled to rt, quenched with
satd aq NH.sub.4Cl, concentrated under reduced pressure and
purified by flash chromatography (MeOH/DCM 0-20%) to give the title
compound as a light tan solid (83 mg, 26%). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.45 (d, J=8.8 Hz, 1H), 7.10 (d, J=2.3 Hz, 1H),
7.03 (dd, J=8.8, 2.3 Hz, 1H), 6.18 (s, 1H), 3.62-3.50 (m, 4H),
3.24-3.18 (m, 4H), 3.05-2.98 (m, 4H), 2.75-2.67 (m, 4H), 2.41 (s,
3H), 1.49 (s, 9H); MS ESI [M+H].sup.+ 565.3, calcd for
[C.sub.28H.sub.36N.sub.8O.sub.3S+H].sup.+ 565.4.
Ethyl
2-(6-(4-methylpiperazine-1-carbonyl)-1H-benzo[d]imidazol-2-yl)acetat-
e
A. (3,4-Dinitrophenyl)(4-methylpiperazin-1-yl)methanone
##STR00032##
[0193] To a suspension of 3,4-dinitrobenzoic acid (1.23 g, 5.8
mmol) in anh DCM (20 mL) at rt was added dropwise oxalyl chloride
(1.0 mL, 11.7 mmol) followed by anh DMF (2 drops). The reaction was
stirred overnight and then concentrated at rt. The residue was
dissolved in anh THF (40 mL) at 0.degree. C. under Ar.
1-Methylpiperazine (1.3 mL, 11.7 mmol) was added dropwise (thick
white suspension was stirred with intermittent shaking). After the
addition the cooling was continued for 10 min before the cooling
bath was removed. After stirring the reaction at rt for 3 h,
H.sub.2O was added. THF was removed under reduced pressure and the
aqueous residue was extracted (CH.sub.2Cl.sub.2; 2% MeOH in
CH.sub.2Cl.sub.2. 2.times.). The combined organic extracts were
dried over Na.sub.2SO.sub.4 and concentrated under reduced pressure
to afford (3,4-dinitrophenyl)(4-methylpiperazin-1-yl)methanone as a
light orange solid (1.77 g, quant). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.29 (d, J=8.3 Hz, 1H), 8.27 (d, J=1.5 Hz,
1H), 7.97 (dd, J=8.3, 1.8 Hz, 1H), 3.59-3.68 (m, 2H), 3.24-2.53
(in, 2H), 2.42-2.35 (in, 2H), 2.21-2.32 (in, 2H), 2.20 (s, 3H). MS
ESI [M+H].sup.+ 295.2, calcd for
[C.sub.12H.sub.14N.sub.4O.sub.5+H].sup.+ 295.1.
B. (3,4-Diaminophenyl)(4-methylpiperazin-1-yl)methanone
##STR00033##
[0195] A solution of
(3,4-dinitrophenyl)(4-methylpiperazin-1-yl)methanone (0.53 g, 1.8
mmol) in THF (25 mL) and EtOH (50 mL) was degassed with N.sub.2.
Pd/C (191 mg, 0.18 mmol) was added and the reaction was stirred
under H.sub.2 (1 atm) overnight at rt. The reaction mixture was
then filtered through Celite and concentrated under reduced
pressure to afford
(3,4-diaminophenyl)(4-methylpiperazin-1-yl)methanone as a purple
solid (0.44 g, quant). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta.
6.61-6.55 (m, 1H), 6.47-6.45 (m, 2H), 4.81 (br.s, 2H), 4.58 (br. s,
2H), 3.50-3.39 (m, 4H), 2.34-2.22 (m, 4H), 2.18 (s, 3H). MS ESI
[M+H].sup.+ 235.1, calcd for [C.sub.12H.sub.18N.sub.4O+H].sup.+
235.1.
C. Ethyl
2-(6-(4-methylpiperazine-1-carbonyl)-1H-benzo[d]imidazol-2-yl)ace-
tate
##STR00034##
[0197] (3,4-Diaminophenyl)(4-methylpiperazin-1-yl)methanone (0.44
g, 1.8 mmol) and 3-ethoxy-3-iminopropanoic acid hydrochloride (1.07
g, 5.5 mmol) in anh EtOH (100 mL) under Ar were heated with
stirring overnight at 65.degree. C. The reaction mixture was then
concentrated under reduced pressure. The residue was taken in to
H.sub.2O (15 mL), neutralized with 10% aq Na.sub.2CO.sub.3,
extracted with CH.sub.2Cl.sub.2 (2.times.), washed (brine) and
dried over Na.sub.2SO.sub.4. Purification by flash chromatography
(0-50% MeOH in CH.sub.2Cl.sub.2) afforded the title compound as a
yellow foam (0.31 g, 52%). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.71-7.57 (m, 2H), 7.33 (d, J=8.2 Hz, 1H), 4.23 (q, J=7.2
Hz, 2H), 3.91-3.40 (m, 4H), 2.62-2.38 (m, 4H), 2.34 (s, 3H), 1.28
(t, J=7.1 Hz, 3H); Signals due to CH.sub.2-ester are absent in
CD.sub.3OD. MS ESI [M+H].sup.+ 331.2, calcd for
[C.sub.17H.sub.22N.sub.4O.sub.3+H].sup.+ 331.2.
Ethyl
2-(6-(morpholine-4-carbonyl)-1H-benzo[d]imidazol-2-yl)acetate
A. (3,4-dinitrophenyl)(morpholino)methanone
##STR00035##
[0199] To a suspension of 3,4-dinitrobenzoic acid (1.30 g, 6.1
mmol) in anh DCM (50 mL) at rt was added dropwise (COCl).sub.2 (1.0
mL, 11.7 mmol) followed by anh DMF (2 drops). The reaction was
stirred overnight and then concentrated at rt. The residue was
dissolved in anh THF (24 mL) at 0.degree. C. under Ar. morpholine
(1.0 mL, 11.6 mmol) was added dropwise (thick white suspension was
stirred with intermittent shaking). After the addition the cooling
was continued for 10 min before the cooling bath was removed. After
stirring the reaction at rt for 3 h, H.sub.2O was added. THF was
removed under reduced pressure and the aqueous residue was
extracted (CH.sub.2Cl.sub.2. 2.times.). The combined organic
extracts were dried (Na.sub.2SO.sub.4) and concentrated under
reduced pressure to afford (3,4-dinitrophenyl)(morpholino)methanone
as a light orange solid (1.8 g, quant). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 8.28-8.31 (m, 2H), 8.00 (dd, J=8.28, 1.76 Hz,
1H), 3.39-3.80 (m, 8H).
B. Ethyl
2-(6-(morpholine-4-carbonyl)-1H-benzo[d]imidazol-2-yl)acetate
##STR00036##
[0201] A solution of (3,4-dinitrophenyl)(morpholino)methanone (0.83
g, 2.9 mmol) in THF (30 mL) and EtOH (60 mL) was degassed with
N.sub.2. Pd/C (0.31 mg, 0.29 mmol) was added and the reaction was
stirred under H.sub.2 (1 atm) overnight at rt. The reaction mixture
was then filtered through Celite and concentrated under reduced
pressure to afford (3,4-diaminophenyl)(morpholino)methanone as a
purple foam. LCMS (ESI) m z calcd for
[C.sub.11H.sub.15N.sub.3O.sub.2+H].sup.+ 222.1; found 222.2. The
material and ethyl 3-ethoxy-3-iminopropanoate hydrochloride (1.2 g,
6.2 mmol) in anh EtOH (100 mL) under Ar were heated with stirring
overnight at 65.degree. C. The reaction mixture was then
concentrated under reduced pressure. Purification by flash
chromatography (0-20% MeOH in CH.sub.2Cl.sub.2) afforded the title
compound as a pale red foam (0.43 g, 47%). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.52-7.76 (m, 2H), 7.33 (dd, J=8.28, 1.51 Hz,
1H), 4.22 (q, J=7.19 Hz, 2H), 4.00-4.05 (m, 2H), 3.70 (br. s., 8H),
1.28 (t, J=7.15 Hz, 3H); Signals due to CH.sub.2-ester are absent
in CD.sub.3OD; MS ESI [M+H].sup.+ 318.2, calcd for
[C.sub.17H.sub.22N.sub.4O.sub.3+H].sup.+ 318.1.
Ethyl
2-(5-methyl-6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acet-
ate
A. 4-methyl-5-(4-methylpiperazin-1-yl)-2-nitroaniline
##STR00037##
[0203] 5-Chloro-4-methyl-2-nitroaniline (0.32 g, 1.7 mmol) and
1-methylpiperazine (1.5 mL, 13.5 mmol) were heated in a sealed tube
at 80.degree. C. for 30 min followed by at 105.degree. C. for 1 d
and 120.degree. C. for 2 d. The reaction was later cooled, diluted
with H.sub.2O and filtered. The collected solid was rinsed with
H.sub.2O and dried in vacuo to afford the title compound a as a
yellow solid (0.36 g, 84%). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. 7.72 (s, 1H), 7.27 (s, 2H), 6.44 (s, 1H), 2.97-2.86 (m 4H),
2.49-2.39 (m, 4H), 2.22 (s, 3H), 2.11 (s, 3H). LCMS (ESI) m z calcd
for [C.sub.12H.sub.18N.sub.4O.sub.2+H].sup.+ 251.1; found
235.3.
B. Ethyl
2-(5-methyl-6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)a-
cetate
##STR00038##
[0205] 4-methyl-5-(4-methylpiperazin-1-yl)-2-nitroaniline (0.36 g,
1.4 mmol) and Pd/C (10%, 81 mg, 0.08 mmol) in EtOH (50 mL), THF (25
mL) were degassed with N.sub.2 and then stirred under H.sub.2 (1
atm) for 5 d. The reaction mix was filtered through Celite, the pad
was rinsed with EtOH. The filtrate was concentrated under reduced
pressure to afford
4-methyl-5-(4-methylpiperazin-1-yl)benzene-1,2-diamine as a yellow
tan solid (0.35 g, quant). The material (0.35 g) and ethyl
3-ethoxy-3-iminopropanoate hydrochloride (0.81 g, 4.1 mmol) in anh
EtOH (70 mL) under Ar were heated with stirring overnight at
65.degree. C. The reaction mixture was then concentrated under
reduced pressure, taken into H.sub.2O (20 mL) and basified with 2 M
aq Na.sub.2CO.sub.3 to pH 9. The mixture was extracted with DCM
(2.times.); the organic extracts were dried (Na.sub.2SO.sub.4) and
concentrated under reduced pressure. Purification by flash
chromatography (0-30% MeOH in CH.sub.2Cl.sub.2) afforded the title
compound as a yellow foam (0.36 g, 82%). .sup.1H NMR (500 MHz,
CD.sub.3OD) .delta. 7.35 (s, 1H), 7.25 (s, 1H), 4.22 (q, J=7.09 Hz,
2H), 2.95-3.03 (m, 4H), 2.88-2.58 (m, 4H), 2.43 (s, 3H), 2.41 (s,
3H), 1.28 (t, J=7.09 Hz, 3H); Signals due to CH.sub.2-ester are
absent in CD.sub.3OD; LCMS (ESI) m z calcd for
[C.sub.17H.sub.24N.sub.4O.sub.2+H].sup.+ 317.2; found 317.3.
Ethyl 2-(5-fluoro-6-morpholino-1H-benzo[d]imidazol-2-yl)acetate
A. 4-fluoro-5-morpholino-2-nitroaniline
##STR00039##
[0207] A mixture of 5-chloro-4-fluoro-2-nitroaniline (1.0 g, 5.24
mmol), morpholine (1.37 mL, 15.7 mmol) and DMSO (5 mL) was heated
in oil bath 140.degree. C. for 3 h. Then H.sub.2O (50 mL) was added
with stirring at 80.degree. C. to precipitate the product and
allowed the suspension to rt, suction filtered, washed with
H.sub.2O and dried to give the title compound as a yellow solid
(1.25 g, 94%). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.17 (d,
J=14.0 Hz, 1H), 6.37 (d, J=8.0 Hz, 1H), 3.83 (t, J=4.4 Hz, 4H),
3.22 (t, J=4.8 Hz, 4H); MS ESI [M+H].sup.+ 242.1, calcd for
[C.sub.10H.sub.12FN.sub.3O.sub.3+H].sup.+ 242.1.
B. 4-fluoro-5-morpholinobenzene-1,2-diamine
##STR00040##
[0209] To a 100 mL round-bottom flask was charged with
4-fluoro-5-morpholino-2-nitroaniline (1.23 g) and MeOH (37 mL) at
rt under Ar blanket. Raney Nickel (0.123 g) was added under
stirring with caution at rt. The reaction mass was slowly heated to
60-65.degree. and hydrazine hydrate (0.86 mL) was added to the
reaction mass dropwise in about 5 min. The reaction was H.sub.2N F
stirred at 65-70.degree. C. for 2 hrs. After reaction completion,
cooled it to rt and filtered the catalyst through a Celite pad
under Ar and washed the Celite pad with MeOH (5 mL*2). The combined
filtrate was concentrated and purified by flash chromatography
(gradient: MeOH/DCM 0-25%) to give the title compound as a light
brown solid (0.615 g, 57%). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 6.51-6.47 (m, 2H), 3.81 (t, J=4.8 Hz, 4H), 2.93 (t, J=4.8
Hz, 4H); MS ESI [M+H].sup.+ 212.0, calcd for
[C.sub.10H.sub.14FN.sub.3O+H].sup.+ 212.1.
C. Ethyl
2-(5-fluoro-6-morpholino-1H-benzo[d]imidazol-2-yl)acetate
##STR00041##
[0211] To a solution of 4-fluoro-5-morpholinobenzene-1,2-diamine
(0.615 g, 2.91 mmol) in EtOH (30 mL) at 65.degree. C. was added
ethyl 3-ethoxy-3-iminopropionate hydrochloride (1.14 g, 5.82 mmol)
in two equal lots at an interval of 5 min each. Then stirred the
reaction mass at 65.degree. C. for 2 hrs. After reaction completion
concentrate the reaction mass under reduced pressure to leaving
behind thick brown oil. To the resulting oil H.sub.2O (25 mL) added
and adjusted the .sub.PH.about.10 using 2 M aq Na.sub.2CO.sub.3.
The resultant mixture was extracted with DCM (30 mL*2) and the
combined extracts were concentrated and purified by flash
chromatography (gradient: Hex/EtOAc 0-40%, then MeOH/DCM 0-20%) to
give the title compound as a brown solid (0.786 g, 88%). .sup.1H
NMR (400 MHz, CD.sub.3OD) .delta. 7.26 (d, J=12.4 Hz, 1H), 7.19 (d,
J=7.6 Hz, 1H), 4.25-4.20 (m, 2H), 3.88 (t, J=4.4 Hz, 4H), 3.08 (t,
J=4.8 Hz, 4H), 1.28 (t, J=7.2 Hz, 3H); MS ESI [M+H].sup.+ 308.1.0,
calcd for [C.sub.15H.sub.18FN.sub.3O.sub.3+H].sup.+ 308.1.
Ethyl
2-(6-(4-methyl-1,4-diazepan-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
A. 5-(4-methyl-1,4-diazepan-1-yl)-2-nitroaniline
##STR00042##
[0213] A mixture of 5-chloro-2-nitroaniline (8.63 g, 50 mmol),
1-methyl-1,4-diazepane (6.85 g, 60 mmol) and K.sub.2CO.sub.3 (8.28
g, 60 mmol) was heat at 90.degree. C. for 20 h. After diluting with
H.sub.2O (500 mL), it was extracted with EtOAc (60 mL.times.3),
concentrated and dried to give crude
5-(4-methyl-1,4-diazepan-1-yl)-2-nitroaniline as a dark red oil
(12.50 g). NMR indicated a mixture of product and
5-chloro-2-nitroaniline (2:1). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.72 (d, J=10.0 Hz, 1H), 6.26 (dd, J=9.8, 2.6 Hz, 1H), 6.02
(d, J=2.4 Hz, 1H), 3.66-3.63 (m, 2H), 3.58 (t, J=6.4 Hz, 2H),
2.77-2.74 (m, 2H), 2.62-2.59 (m, 2H), 2.39 (s, 3H), 2.07-2.00 (m,
2H); MS ESI [M+H].sup.+ 251.3, calcd for
[C.sub.12H.sub.18N.sub.4O.sub.2+H].sup.+ 251.15.
B. 4-(4-methyl-1,4-diazepan-1-yl)benzene-1,2-diamine
##STR00043##
[0215] To a mixture of crude
5-(4-methyl-1,4-diazepan-1-yl)-2-nitroaniline (12.50 g) and
Raney-Nickel (1.25 g) in MeOH (150 mL) at 65.degree. C. was added
N.sub.2H.sub.4--H.sub.2O (12.0 mL) over 10 min. After addition, the
resulting mixture was stirred at 70.degree. C. for 30 min. Upon
cooling to rt, it was filtered through Celite and rinsed with MeOH.
The filtrate was concentrated and dried to give crude
4-(4-methyl-1,4-diazepan-1-yl)benzene-1,2-diamine as a dark red
brown oil (10.57 g). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 6.63
(d, J=8.0 Hz, 1H), 6.53 (dd, J=8.4, 2.4 Hz, 1H), 6.26 (d, J=2.4 Hz,
1H), 3.60-3.40 (m, 4H), 2.75-2.71 (m, 2H), 2.62-2.58 (m, 2H), 2.37
(s, 3H), 2.04-1.97 (m, 2H).
C. Ethyl
2-(6-(4-methyl-1,4-diazepan-1-yl)-1H-benzo[d]imidazol-2-yl)acetat-
e
##STR00044##
[0217] A mixture of crude
4-(4-methyl-1,4-diazepan-1-yl)benzene-1,2-diamine (10.57 g) and
ethyl 3-ethoxy-3-iminopropionate hydrochloride (19.50 g, 100 mmol)
in EtOH (200 mL) was heated at 90.degree. C. for 2 h. After removal
of solvents, it was diluted with H.sub.2O (50 mL), basified with 2
M aq Na.sub.2CO.sub.3 (40 mL) and extracted with DCM (60
mL.times.3). The combined extracts were concentrated and purified
by flash chromatography (gradient: 0-100% EtOAc/hexane, then
MeOH/DCM 0-25%) to give the title compound as a dark brown oil
(7.31 g, 46% over 3 steps). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 8.38 (d, J=8.8 Hz, 1H), 6.82-6.77 (m, 2H), 4.22 (q, J=6.8
Hz, 2H), 3.66-3.61 (m, 2H), 3.54 (t, J=6.4 Hz, 2H), 2.85-2.80 (m,
2H), 2.68-2.64 (m, 2H), 2.41 (s, 3H), 2.12-2.05 (m, 2H), 1.29 (t,
J=7.0 Hz, 3H); MS ESI [M+H].sup.+ 317.3, calcd for
[C.sub.17H.sub.24N.sub.4O.sub.2+H].sup.+ 317.20.
Representative Examples
A1:
4-amino-5-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[-
2,3-b]pyridin-6(7H)-one
##STR00045##
[0219] To a solution of ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(2.42 g, 8.05 mmol) and 2-aminothiophene-3-carbonitrile (1.0 g,
8.05 mmol) in anh THF (40 mL) at 40.degree. C. added LDA (40 mL, 1
M in THF/hexane, 40 mmol) dropwise over 15 min under Ar. The
resulting brown solution was stirred at 40.degree. C. for 2 h and
then quenched with aq NH.sub.4Cl (50 mL) at rt. The mixture was
diluted with H.sub.2O (125 mL) and extracted with ethyl acetate
(2.times.200 mL). The combined organic layers were washed once with
H.sub.2O, dried over Na.sub.2SO.sub.4, and concentrated to give
crude product. The crude product was triturated with DCM (20 mL)
followed by MeOH (25 mL) to give the title compound as a light
brown solid (1.95 g, 64%).
[0220] The free base (1.95 g) was suspended in MeOH (50 mL) and
added 1 M HCl-Et.sub.2O (13 mL) at rt. The suspension was stirred
for 15 min at rt and concentrated under vacuum and azeotroped with
MeOH (2.times.25 mL) to give the HCl salt as a dark brown solid
(2.28 g, 62%); .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.69 (d,
J=9.2 Hz, 1H), 7.52 (d, J=5.6 Hz, 1H), 7.36 (dd, J=8.8, 2.4 Hz,
1H), 7.30 (d, J=2.4 Hz, 1H), 7.19 (d, J=5.6 Hz, 1H), 3.97-3.93 (m,
2H), 3.70-3.67 (m, 2H), 3.39-3.35 (m, 2H), 3.34-3.18 (m, 2H), 3.01
(s, 3H); MS ESI [M+H].sup.+ 381.2, calcd for
[C.sub.19H.sub.20N.sub.6OS+H].sup.+ 381.1.
TABLE-US-00002 Yield; Example/IUPAC name Structure description;
salt A2: 4-amino-3-methyl-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl) thieno[2,3-b]pyridin-6(7H)- one ##STR00046##
64 mg (38%); Grey solid 2HCl Reagents (general method A1): ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2- yl)acetate
(110 mg, 0.36 mmol), 2-amino-4-methylthiophene-3-carbonitrile (50
mg, 0.36 mmol), LDA (1.62 mL, 1M in THF/hexane, 1.62 mmol), anh THF
(5.0 mL) .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.72 (d, J = 9.2
Hz, 1H), 7.41 (dd, J = 8.8 Hz, 2.4 Hz, 1H), 7.33 (d, J = 2.0 Hz,
1H), 6.76 (d, J = 0.8 Hz, 1H), 3.98-3.94 (m, 2H), 3.70-3.67 (m,
2H), 3.38-3.35 (m, 2H), 3.27-3.21 (m, 2H), 3.00 (s, 3H), 2.30 (s,
3H); MS calcd; MS ESI [M + H].sup.+ 395.3, calcd for
[C.sub.20H.sub.22N.sub.6OS + H].sup.+ 395.1 A3:
4-amino-2-methyl-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00047##
18 mg (10%); Light brown solid 2HCl Reagents (general method A1):
ethyl 2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-
yl)acetate (110 mg, 0.36 mmol), 2
amino-5-methylthiophene-3-carbonitrile (50 mg, 0.36 mmol), LDA
(1.80 mL, 1M in THF/hexane, 1.80 mmol), anh THF (5.0 mL) .sup.1H
NMR (400 MHz, CD.sub.3OD) .delta. 7.68 (d, J = 8.8 Hz, 1H),
7.38-7.31 (m, 2H), 7.18 (s, 1H), 3.96- 3.93 (m, 2H), 3.70-3.67 (m,
2H), 3.40-3.34 (m, 2H), 3.25-3.19 (m, 2H), 3.01 (s, 3H), 2.52 (s,
3H); MS calcd; MS ESI [M + H].sup.+ 395.3, calcd for
[C.sub.20H.sub.22N.sub.6OS + H].sup.+ 395.1 A4:
4-amino-5-(6-morpholino- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00048## 0.51 g (51%); Light
tan solid; Free base Reagents (general method A1): LiHMDS (1.0M in
THF, 14.3 mL, 14.3 mmol) was added dropwise over 20 min to a
stirred solution of 2-aminothiophene-3-carbonitrile (0.340 g, 2.73
mmol), ethyl 2-(6-morpholino-1H-benzo[d]imidazol-2-yl)acetate
(0.829 g, 2.86 mmol) in anh THF (20 mL) at rt under Ar. The
reaction was heated at 40.degree. C. for 1 h, then cooled to rt,
quenched with satd aq NH.sub.4Cl, concentrated under reduced
pressure and purified by flash chromatography
(MeOH--CH.sub.2Cl.sub.2 0-7%). A small sample was repurified by
prep HPLC to afford the TFA salt (a light yellow solid). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 12.18 (s, 1H), 7.59 (d, J = 5.7
Hz, 1H), 7.56 (br s, 1H), 7.36-7.24 (br., s, 1H), 7.20 (d, J = 5.7
Hz, 1H), 7.17-7.08 (m, 1H), 3.86-3.79 (m, 4H), 3.30-3.18 (m, 4H);
three exchangeable protons may be attributed to two very broad
peaks 13.47-12.46 (brs, 1H) and 9.30-7.67 (brs, 2H), MS ESI [M +
H].sup.+ 368.2, calcd for [C.sub.18H.sub.17N.sub.5O.sub.2S +
H].sup.+ 368.1. A5: 4-amino-3-methoxy-5-(6-
(4-methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00049## 55 mg (11%); brown
solid; TFA Reagents (General method A1): ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2- yl)acetate
(302 mg, 1 mmol), 2-amino-4-methoxythiophene-3-carbonitrile (154
mg, 1 mmol), LDA (1.0M in THF/hex, 5 mL, 5 mmol), THF (10 mL).
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.70 (d, J = 8.8 Hz, 1H),
7.39 (dd, J = 9.2, 2.4 Hz, 1H), 7.31 (d, J = 1.6 Hz, 1H), 6.09 (s,
1H), 3.98- 3.88 (m, 2H), 3.85 (s, 3H), 3.72-3.64 (m, 2H), 3.38-3.28
(m, 2H), 3.25-3.14 (m, 2H), 2.87 (s, 3H); MS ESI [M + H].sup.+
411.3, calcd for [C.sub.20H.sub.22N.sub.6O.sub.2S + H].sup.+ 411.2.
A6: 4-amino-3-ethoxy-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00050##
200 mg (61%); dark brown solid; 2HCl salt Reagents (general method
A1): 2-amino-4-ethoxythiophene-3-carbonitrile (111 mg, 0.66 mmol),
ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate (200
mg, 0.66 mmol), LiHMDS (1M in THF, 2.65 mL, 2.65 mmol). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.72 (d, J = 8.7 Hz, 1H), 7.42 (d, J
= 8.4 Hz, 1H), 7.32 (s, 1 H), 6.08 (s, 1H), 4.08 (q, J = 6.8 Hz,
2H), 3.96 (d, J = 11.5 Hz, 2H), 3.68 (d, J = 11.8 Hz, 2H),
3.41-3.26 (m, 2H), 3.27-3.16 (m, 2H), 3.01 (s, 3H), 1.42 (t, J =
6.8 Hz, 3H); MS ESI [M + H].sup.+ 425.3, calcd for
[C.sub.21H.sub.24N.sub.6O.sub.2S + H].sup.+ 425.2. A7:
5-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-yl)-
4-((tetrahydro-2H-pyran-4- yl)amino)thieno[2,3-b]pyridin- 6(7H)-one
##STR00051## 35 mg (43%); brown solid; TFA Step 1: Reagents
(General method C): a mixture of 7-(4-methoxybenzyl)-5-(5 and/or
6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.11 g, 0.14 mmol), tetrahydro-
2H-pyran-4-amine (0.035 g, 0.35 mmol), DCM (15 mL). MS ESI [M +
H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.21.
Step 2: Reagents (general method D): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4-
((tetrahydro-2H-pyran-4-yl)amino)thieno[2,3-b]pyridin-6(7H)-one
(0.10 g, 0.14 mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.65 (d, J = 9.0 Hz, 1H), 7.58 (d, J
= 5.8 Hz, 1H), 7.29 (s, 2H), 7.19 (d, J = 6.0 Hz, 1H), 4.02-3.83
(m, 4H), 3.74-3.60 (m, 2H), 3.58-3.46 (m, 1H), 3.43-3.33 (m, 2H),
3.29-3.09 (m, 4H), 3.01 (s, 3H), 2.03-1.90 (m, 2H), 1.85-1.69 (m,
2H); MS ESI [M + H].sup.+ 465.3, calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2. A8:
4-(((1r,3r)-3- hydroxycyclobutyl)amino)-5-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00052## 18 mg (16%); yellow
solid; 2 HCl Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.17 g, 0.22 mmol), (1r,3r)-3-
aminocyclobutanol HCl salt (0.068 g, 0.55 mmol), DCM (10 mL). MS
ESI [M + H].sup.+ 703.2, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 703.19.
Step 2: Reagents (General method D): a mixture of
4-(((1r,3r)-3-hydroxycyclobutyl)amino)-7- (4-methoxybenzyl)-5-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]-imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (0.16 g, 0.22
mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.69 (d, J = 9.3 Hz, 1H), 7.61 (d, J = 6.0 Hz,
1H), 7.37 (dd, J = 9.0, 2.5 Hz, 1H), 7.34-7.29 (m, 1H), 7.20 (d, J
= 6.0 Hz, 1H), 4.45-4.33 (m, 1H), 4.03-3.90 (m, 3H), 3.76-3.61 (m,
2H), 3.42-3.36 (m, 2H), 3.30-3.18 (m, 2H), 3.01 (s, 3H), 2.54-2.42
(m, 2H), 2.14-2.03 (m, 2 H); MS ESI [M + H].sup.+ 451.3, calcd for
[C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.2. A9:
4-(((1R*,3R*)-3- hydroxycyclopentyl)amino)-5-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00053## 15 mg (11%); yellow
solid; 2 HCl Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.17 g, 0.22 mmol), (1R*,3R*)-
3-aminocyclopentanol HCl salt (0.076 g, 0.55 mmol), DCM (10 mL). MS
ESI [M + H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.21.
Step 2: Reagents (general method D): a mixture of 4-(((1R*,3R*)-3-
hydroxycyclopentyl)amino)-7-(4-methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-
((trifluoromethyl)sulfonyl)-1H-benzo[d]-imidazol-2-yl)thieno[2,3-b]pyridin-
-6(7H)-one (0.16 g, 0.22 mmol), TFA (7 mL), and conc. HCl (1 mL).
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.68 (d, J = 8.8 Hz, 1H),
7.67 (d, J = 6.0 Hz, 1H), 7.39 (dd, J = 8.8, 2.3 Hz, 1H), 7.32 (d,
J = 2.0 Hz, 1H), 7.20 (d, J = 5.8 Hz, 1H), 4.32-4.24 (m, 1H),
4.01-3.91 (m, 2H), 3.86-3.75 (m, 1H), 3.73-3.64 (m, 2H), 3.41-3.36
(m, 2H), 3.30-3.19 (m, 2H), 3.01 (s, 3H), 2.06-1.94 (m, 2H),
1.91-1.84 (m, 2H), 1.74-1.62 (m, 1H), 1.47-1.40 (m, 1H); MS ESI [M
+ H].sup.+ 465.2, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S +
H].sup.+ 465.2. A10: (R)-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-4- ((tetrahydrofuran-3-
yl)amino)thieno[2,3-b]pyridin- 6(7H)-one ##STR00054## 87 mg (53%);
yellow solid; TFA Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.23 g, 0.30 mmol), (R)-
tetrahydrofuran-3-amine (0.11 g, 0.90 mmol), DCM (12 mL). MS ESI [M
+ H].sup.+ 703.2, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 703.19.
Step 2: Reagents (general method D): a mixture of
(R)-7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4- ((tetrahydrofuran-3-yl)amino)thieno[2,3-b]pyridin-6(7H)-one
(0.21 g, 0.30 mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.61 (d, J = 8.8 Hz, 1H), 7.59 (d, J
= 6.0 Hz, 1H), 7.28-7.22 (m, 2H), 7.17 (d, J = 5.8 Hz, 1H),
4.30-4.22 (m, 1H), 4.05-3.97 (m, 1H), 3.92-3.75 (m, 5H), 3.65 (br.
s., 2H), 3.38-3.26 (m, 2H), 3.24-3.08 (m, 2H), 3.00 (s, 3H),
2.28-2.16 (m, 1H), 2.14-2.05 (m, 1H); MS ESI [M + H].sup.+ 451.2,
calcd for [C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.2. A11:
(S)-5-(6-(4- methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-4-
((tetrahydrofuran-3- yl)amino)thieno[2,3-b]pyridin- 6(7H)-one
##STR00055## 61 mg (36%); brown solid; TFA Step 1: Reagents
(general method C): a mixture of 7-(4-methoxybenzyl)-5-(5 and/or
6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7- dihydro-thieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.23 g, 0.30 mmol), (S)-
tetrahydrofuran-3-amine (0.11 g, 0.90 mmol), DCM (12 mL). MS ESI [M
+ H].sup.+ 703.2, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 703.19.
Step 2: Reagents (general method D): a mixture of
(S)-7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4- ((tetrahydrofuran-3-yl)amino)thieno[2,3-b]pyridin-6(7H)-one
(0.21 g, 0.30 mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.63 (d, J = 8.8 Hz, 1H), 7.61 (d, J
= 5.8 Hz, 1H), 7.29-7.23 (m, 2H), 7.19 (d, J = 5.8 Hz, 1H),
4.33-4.22 (m, 1H), 4.06-3.97 (m, 1H), 3.95-3.76 (m, 5H), 3.66 (br.
s., 2H), 3.41-3.32 (m, 2H), 3.24-3.08 (m, 2H), 3.01 (s, 3H),
2.29-2.16 (m, 1H), 2.14-2.02 (m, 1H); MS ESI [M + H].sup.+ 451.2,
calcd for [C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.18. A12:
4-(((1s,3s)-3- hydroxycyclobutyl)amino)-5-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00056## 101 mg (60%);
yellow
solid; TFA Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7- dihydro-thieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.23 g, 0.30 mmol), (1s,3s)-3-
aminocyclobutanol HCl salt (0.11 g, 0.90 mmol), DMF (8 mL). MS ESI
[M + H].sup.+ 703.1, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 703.19.
Step 2: Reagents (general method D): a mixture of
4-(((1s,3s)-3-hydroxycyclobutyl)amino)-7- (4-methoxybenzyl)-5-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]-imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (0.21 g, 0.30
mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.62 (d, J = 9.5 Hz, 1H), 7.57 (d, J = 6.0 Hz,
1H), 7.30-7.21 (m, 2H), 7.16 (d, J = 6.0 Hz, 1H), 3.97-3.79 (m,
3H), 3.73-3.50 (m, 3H), 3.40-3.26 (m, 2H), 3.17 (m, 2H), 3.00 (s,
3H), 2.69-2.56 (m, 2H), 2.16-2.00 (m, 2H); MS ESI [M + H].sup.+
451.2, calcd for [C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.2.
A13: 4-(((1R*,3S*)-3- hydroxycyclopentyl)amino)-5-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00057## 96 mg (57%); yellow
solid; TFA Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7- dihydro-thieno[2,3-b]pyridin-4-yl (0.23 g, 0.30
mmol), (1S*,3R*)-3-aminocyclopentanol HCl salt (0.12 g, 0.90 mmol),
DMF (8 mL). MS ESI [M + H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.21.
Step 2: Reagents (general method D): a mixture of 4-(((1R*,3S*)-3-
hydroxycyclopentyl)amino)-7-(4-methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-
((trifluoromethyl)sulfonyl)-1H-benzo[d]-imidazol-2-yl)thieno[2,3-b]pyridin-
-6(7H)-one (0.21 g, 0.30 mmol), TFA (7 mL), and conc. HCl (1 mL).
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.62 (d, J = 9.0 Hz, 1H),
7.53 (d, J = 5.8 Hz, 1H), 7.28 (dd, J = 2.1, 8.9 Hz, 1H), 7.24 (d,
J = 1.8 Hz, 1H), 7.19 (d, J = 5.8 Hz, 1H), 4.47-4.39 (m, 1H),
4.23-4.12 (m, 1H), 3.96-3.81 (m, 2H), 3.73- 3.60 (m, 2H), 3.39-3.26
(m, 2H), 3.25-3.11 (m, 2H), 3.00 (s, 3H), 2.09-1.82 (m, 6H); MS ESI
[M + H].sup.+ 465.2, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S +
H].sup.+ 465.2. A14: 4-(((3R,4R)-3- fluoropiperidin-4-yl)amino)-5-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00058## 126 mg (58%);
orange solid; 2 TFA Step 1: Reagents (general method C): a mixture
of 7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.24 g, 0.31 mmol), (3R,4R)-tert- butyl
4-amino-3-fluoropiperidine-1-carboxylate (0.20 g, 0.93 mmol), DMF
(5 mL). MS ESI [M - CF.sub.3O.sub.2S + 2H].sup.+ 702.2, calcd for
[C.sub.37H.sub.44FN.sub.7O.sub.4S + H].sup.+ 702.3. Step 2:
Reagents (general method D): a mixture of (3R,4R)-tert-butyl
3-fluoro-4-((7-(4- methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]-imidazol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl)amino)p-
iperidine-1- carboxylate (0.26 g, 0.31 mmol), TFA (7 mL), and conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.57 (d, J =
8.8 Hz, 1H), 7.51 (d, J = 6.0 Hz, 1H), 7.23 (d, J = 1.8 Hz, 1H),
7.19 (d, J = 6.0 Hz, 1H), 7.15 (dd, J = 1.8, 8.8 Hz, 1H), 5.13-4.87
(m, 2H), 4.33-4.21 (m, 1H), 3.94-3.50 (m, 6H), 3.44-3.34 (m, 2H),
3.28-3.07 (m, 3H), 3.00 (s, 3H), 2.49-2.37 (m, 1H), 2.19-2.05 (m,
1H); MS ESI [M + H].sup.+ 482.2, calcd for
[C.sub.24H.sub.28FN.sub.7OS + H].sup.+ 482.2. A15:
4-((3,3-difluoropiperidin- 4-yl)amino)-5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00059## 134 mg (59%);
yellow solid; 2 TFA Step 1: Reagents (general method C): A mixture
of 7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.24 g, 0.31 mmol), tert-butyl 4-
amino-3,3-difluoropiperidine-1-carboxylate (0.22 g, 0.93 mmol), DMF
(5 mL). MS ESI ([M - CF.sub.3O.sub.2S + H].sup.+ 719.2, calcd for
[C.sub.37H.sub.43F.sub.2N.sub.7O.sub.4S].sup.+ 719.31. Step 2:
Reagents (general method D): a mixture of tert-butyl
3,3-difluoro-4-((7-(4- methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl)amino)pi-
peridine-1- carboxylate (0.26 g, 0.31 mmol), TFA (7 mL), and conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.60-7.50 (m,
2H), 7.26-7.17 (m, 2H), 7.13 (dd, J = 1.8, 9.0 Hz, 1H), 4.65-4.48
(m, 1H), 3.96-3.45 (m, 8H), 3.29-3.04 (m, 4H), 2.98 (s, 3H),
2.59-2.41 (m, 1H), 2.40-2.22 (m, 1H); MS ESI [M + H].sup.+ 500.2,
calcd for [C.sub.24H.sub.27F.sub.2N.sub.7OS + H].sup.+ 500.2. A16:
(R)-4-((1-hydroxy-3- methylbutan-2-yl)amino)-5-(6-
(4-methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00060## 40 mg (29%), yellow
solid; free base Step 1: Reagents (general method C):
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-oxo-6,7-di-
hydrothieno[2,3- b]pyridin-4-yl trifluoromethanesulfonate (crude,
0.30 mmol), (R)-2-amino-3-methylbutan-1-ol (0.12 g, 1.2 mmol), DMF
(7 mL). ESI [M + H].sup.+ 719.2, calcd for
[C.sub.33H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ Step 2:
Reagents (general method D):
(R)-4-((1-hydroxy-3-methylbutan-2-yl)amino)-7-(4-
methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]-imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (crude, 0.30
mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.65 (d, J = 6.0 Hz, 1H), 7.47-7.42 (m, 1H), 7.19-7.13 (m,
1H), 7.10 (d, J = 5.7 Hz, 1H), 7.02-6.94 (m, 1H), 4.27-4.19 (m,
1H), 3.92-3.82 (m, 2H), 3.21 (br s, 4H), 2.69 (br s, 4H), 2.38 (s,
3H), 2.32-2.22 (m, 1H), 1.23 (d, J = 7.0 Hz, 3H), 1.02 (d, J = 7.0
Hz, 3H); ESI [M + H].sup.+ 467.3, calcd for
[C.sub.24H.sub.30N.sub.6O.sub.2S + H].sup.+ 467.2 A17: 4-((5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-6-oxo-
6,7-dihydrothieno[2,3- b]pyridin-4- yl)amino)piperidine-1-
carbaldehyde ##STR00061## 40 mg (27%), Yellow solid; Free base Step
1: Reagents (general method C): 7-(4-methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-oxo-6,7-di-
hydrothieno[2,3- b]pyridin-4-yl trifluoromethanesulfonate (crude,
0.30 mmol), 4-aminopiperidine-1- carbaldehyde (0.15 g, 1.2 mmol),
DMF (7 mL). ESI [M + H].sup.+ 744.1, calcd for
[C.sub.34H.sub.36F.sub.3N.sub.7O.sub.5S.sub.2 + H].sup.+ 744.2 Step
2: Reagents (general method D): 4-((7-(4-methoxybenzyl)-5-(5 and/or
6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7-
dihydrothieno[2,3-b]pyridin-4-yl)amino)piperidine-1-carbaldehyde
(crude, 0.30 mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 8.06 (s, 1H), 7.45 (s, 2H), 7.18-7.06 (m, 2H),
7.00-6.91 (m, 1H), 4.52-4.40 (m, 1H), 4.09-3.95 (m, 1H), 3.84-3.73
(m, 1H), 3.52-3.38 (m, 2H), 3.20 (br s, 4H), 2.66 (br s, 4H), 2.36
(s, 3H), 2.14 (br s, 2H), 1.91-1.69 (m, 2H); ESI [M + H].sup.+
492.2, calcd for [C.sub.25H.sub.29N.sub.7O.sub.2S + H].sup.+ 492.2
A18: 4-((1-methylpiperidin-4- yl)amino)-5-(6-morpholino-
1H-benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin-6(7H)- one
##STR00062## 15 mg (8%), yellow solid; Free base Step 1: Reagents:
(general method C): 7-(4-methoxybenzyl)-5-(5 and/or 6-morpholino-1-
((trifluoro-methyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-oxo-6,7-dihydroth-
ieno[2,3- b]pyridin-4-yl trifluoro-methanesulfonate (crude, 0.41
mmol), tert-butyl 4-aminopiperidine-1- carboxylate (0.19 g, 1.6
mmol), DMF (7 mL). ESI [M + H].sup.+ 717.1, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2 Step
2: Reagents (general method D):
7-(4-methoxybenzyl)-4-((1-methylpiperidin-4- yl)amino)-5-(5 and/or
6-morpholino-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)-one (crude, 0.30 mmol), TFA (5 mL),
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.52-7.43 (m,
2H), 7.19-7.10 (m, 2H), 7.03-6.95 (m, 1H), 4.38-4.23 (m, 1H),
3.92-3.82 (m, 4H), 3.19-3.10 (m, 4H), 2.98-2.87 (m, 2H), 2.59-2.44
(m, 2H), 2.38 (s, 3H), 2.26-2.14 (m, 2H), 2.00-1.87 (m, 2H); ESI [M
+ H].sup.+ 465.2, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S +
H].sup.+ 465.2 A19: 4-(cyclopentylamino)-3- methyl-5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin-6(7H)- one ##STR00063## 59 mg (36%); Yellow
solid 2HCl Step-01: Reagents (general method C):
7-(4-methoxybenzyl)-3-methyl-5-(5 and/or 6-(4-
methylpipera-zin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benoz[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]py-ridin-4-yl
trifluoromethanesulfonate (242 mg, 0.31 mmol), cyclopentylamine (80
mg, 0.93 mmol), DMF (4 mL); MS calcd; MS ESI [M + H].sup.+ 715.2,
calcd for [C.sub.34H.sub.37F.sub.3N.sub.6O.sub.4S.sub.2 + H].sup.+
715.2 Step-02: Reagents (general method D):
4-(cyclopentylamino)-7-(4-methoxybenzyl)-3-methyl- 5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imida-
zol-2- yl)thieno[2,3-b]pyridin-6(7H)-one (245 mg), TFA (6 mL),
conc, HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.72
(d, J = 9.2 Hz, 1H), 7.44-7.41 (m, 1H), 7.32 (d, J = 1.6 Hz, 1H),
6.86 (s, 1H), 3.99-3.96 (m, 2H), 3.71-3.68 (m, 2H), 3.40-3.34 (m,
2H), 3.28-3.23 (m, 2H), 3.18-3.15 (m, 1H), 3.01 (s, 3H), 2.65 (s,
3H), 1.68-1.61 (m, 6H), 1.46-1.44 (m, 2H); MS ESI [M + H].sup.+
463.2, calcd for [C.sub.25H.sub.30N.sub.6OS + H].sup.+ 463.2 A20:
3-methyl-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-4- ((tetrahydro-2H-pyran-4-
yl)amino)thieno[2,3-b]pyridin- 6(7H)-one ##STR00064## 60 mg (35%);
Yellow solid 2HCl Step-01: Reagents (general method C):
7-(4-methoxybenzyl)-3-methyl-5-(5 and/or 6-(4-
methy-lpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno-[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (242 mg, 0.31 mmol),
tetrahydro-2H-pyran-4-amine (95 mg, 0.93 mmol), DMF (4 mL). MS
calcd; MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2
Step-02: Reagents (general method D):
7-(4-methoxybenzyl)-3-methyl-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-4-
((tetrahydro-2H-pyran-4-yl)amino)thieno[2,3-b]pyridin-6(7H)-one
(250 mg), TFA (4 mL), conc HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.75 (d, J = 9.2 Hz, 1H), 7.44 (dd, J =
9.2,
2.0 Hz, 1H), 7.35 (d, J = 2.0 Hz, 1H), 6.88 (s, 1H), 4.01-3.98 (m,
2H), 3.81-3.78 (m, 2H), 3.71- 3.68 (m, 2H), 3.40-3.35 (m, 2H),
3.31-3.23 (m, 2H), 3.01 (s, 3H), 2.86-2.80 (m, 2H), 2.68 (s, 3H),
2.62-2.56 (m, 1H), 1.83-1.80 (m, 2H), 1.65-1.55 (m, 2H); MS ESI [M
+ H].sup.+ 479.1, calcd for [C.sub.25H.sub.30N.sub.6O.sub.2S +
H].sup.+ 479.2 A21: 4-amino-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-3- (piperazin-1-yl)thieno[2,3-
b]pyridin-6(7H)-one ##STR00065## 74 mg (82%); tan solid; 2TFA
tert-Butyl
4-(4-amino-5-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2--
yl)-6-oxo-6,7-
dihydro-theino[2,3-b]pyridin-3-yl)piperazine-1-carboxylate (77.8
mg, 0.13 mmol) in DCM (20 mL) was treated with TFA (2 mL) at rt.
The reaction was stirred 2.5 h before being concentrated under
reduced pressure and purified by prep HPLC. .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.71 (d, J = 9.0 Hz, 1H), 7.40 (dd, J = 9.0,
2.3 Hz, 1H), 7.32 (d, J = 2.0 Hz, 1H), 6.40 (s, 1H), 4.02-3.85 (m,
2H), 3.74-3.61 (br m, 2H), 3.46-3.38 (m, 4H), 3.37-3.28 (m, 6H),
3.26- 3.11 (m, 2H), 3.00 (s, 3H); MS ESI [M + H].sup.+ 465.4, calcd
for [C.sub.23H.sub.28N.sub.8OS + H].sup.+ 465.2. A22: 6-(6-(4-
methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2- yl)-7-((pyridin-4-
ylmethyl)amino)thieno[3,2-b] pyridin-5(4H)-one ##STR00066## 16 mg
(9%); yellow solid; TFA salt Step 1: Reagents (general method C): a
mixture of 4-(4-methoxybenzyl)-6-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-5-oxo-4,5- dihydro-thieno[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (crude, 0.3 mmol), pyridin-4-
ylmethanamine (0.09 mL, 0.89 mmol). MS ESI [M + H].sup.+ 724.2,
calcd for [C.sub.34H.sub.32F.sub.3N.sub.7O.sub.4S.sub.2 + H].sup.+
724.2. Step 2: Reagents (general method D):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-7-((pyridin--
4- ylmethyl)amino)thieno[3,2-b]pyridin-5(4H)-one, TFA (4 mL), conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 8.77 (d, J =
6.8 Hz, 2H), 8.05 (d, J = 6.8 Hz, 2H), 7.84 (d, J = 5.6 Hz, 1H),
7.58 (d, J = 8.8 Hz, 1H), 7.25 (d, J = 2.0 Hz, 1H), 7.14 (dd, J =
8.8, 2.0 Hz, 1H), 7.06 (d, J = 5.6 Hz, 1H), 5.41 (s, 2H), 3.91-3.78
(m, 2H), 3.75-3.59 (m, 2H), 3.41-3.33 (m, 2H), 3.21-3.05 (m, 2H),
3.00 (m, 3H). MS ESI [M + H].sup.+ 472.3, calcd for
[C.sub.25H.sub.25N.sub.7OS + H].sup.+ 472.2. A23: 4-amino-3-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[3,4-b]pyridin- 2(1H)-one ##STR00067## 43 mg (18%); Pale
yellow solid 2HCl Reagents (general method A1): ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2- yl)acetate
(160 mg, 0.52 mmol), 4-aminothiophene-3-carbonitrile (65 mg, 0.52
mmol), LDA (2.6 mL, 1M in THF/hexane, 2.35 mmol), anh THF (6.0 mL)
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 8.39 (d, J = 3.2 Hz, 1H),
7.69 (d, J = 8.8 Hz, 1H), 7.39-7.37 (m, 1H), 7.30 (d, J = 2.0 Hz,
1H), 6.96 (d, J = 3.6 Hz, 1H), 3.97-3.94 (m, 2H), 3.71-3.67 (m,
2H), 3.39-3.35 (m, 2H), 3.24-3.18 (m, 2H), 3.01 (s, 3H); MS ESI [M
+ H].sup.+ 381.1, calcd for [C.sub.19H.sub.20N.sub.6OS + H].sup.+
381.1 A24: 4-(cyclopentylamino)- 3-(6-(4-methylpiperazin-1-
yl)-1H-benzo[d]imidazol-2- yl)thieno[3,4-b]pyridin- 2(1H)-one
##STR00068## 11 mg (5%); Yellow solid 2HCl Step-01: Reagents
(general method C): 1-(4-methoxybenzyl)-3-(5- and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-2-oxo-1,2- dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate (223 mg, 0.29 mmol), cyclopentylamine (73
mL, 0.72 mmol), DCM (10 mL). MS ESI [M + H].sup.+ 701.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.4S.sub.2 + H].sup.+ 701.2
Step-02: Reagents (general method D):
4-(cyclopentylamino)-1-(4-methoxybenzyl)-3-(5- and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d-
]imidazol-2- yl)thieno[3,4-b]pyridin-2(1H)-one (180 mg, 0.25 mmol),
TFA (7 mL), conc HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 8.53 (d, J = 3.2 Hz, 1H), 7.71 (d, J = 9.2 Hz, 1H), 7.42
(dd, J = 9.2 Hz, 2.4 Hz, 1H), 7.32 (d, J = 2.0 Hz, 1H), 6.97 (d, J
= 3.2 Hz, 1H), 3.99-3.96 (m, 2H), 3.70-3.67 (m, 2H), 3.39-3.35 (m,
2H), 3.28-3.22 (m, 2H), 3.01 (s, 3H), 1.71-1.69 (m, 6H), 1.38-1.37
(m, 2H), 1H merged with H.sub.2O; MS ESI [M + H].sup.+ 449.2, calcd
for [C.sub.24H.sub.28N.sub.6OS + H].sup.+ 449.2. A25: 3-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-4-
((tetrahydro-2H-pyran-4- yl)amino)thieno[3,4- b]pyridin-2(1H)-one
##STR00069## 12 mg (4%); Dark brown solid 2HCl Step-01: Reagents
(general method C): 1-(4-methoxybenzyl)-3-(5- and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-2-oxo-1,2- dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate (380 mg, 0.49 mmol), tetrahydro-
2H-pyran-4-amine (125 mg, 1.23 mmol), DCM (10 mL). MS ESI [M +
H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2
Step-02: Reagents (general method D): 1-(4-methoxybenzyl)-3-(5-
and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4-
((tetrahydro-2H-pyran-4-yl)amino)thieno[3,4-b]pyridin-2(1H)-one
(110 mg, 0.15 mmol), TFA (4 mL), conc HCl (0.5 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 8.50 (d, J = 3.2 Hz, 1H), 7.75 (d, J
= 8.2 Hz, 1H), 7.45 (dd, J = 9.2 Hz, 2.4 Hz, 1H), 7.34 (d, J = 2.0
Hz, 1H), 6.98 (d, J = 3.2 Hz, 1H), 4.02-3.99 (m, 2H), 3.84-3.81 (m,
2 H), 3.71-3.68 (m, 2H), 3.50-3.37 (m, 2H), 3.27-3.21 (m, 2H), 3.02
(s, 3H), 2.80-2.77 (m, 2H), 2.75-2.64 (m, 1H), 1.80-1.66 (m, 4H);
MS ESI [M + H].sup.+ 465.2, calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2 A26: 3-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-4-
((2-morpholinoethyl)amino) thieno[3,4-b]pyridin-2(1H)- one
##STR00070## 38 mg (26%); Yellow solid Free base Step-01: Reagents
(general method C): 1-(4-methoxybenzyl)-3-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-2-oxo-1,2- dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate (229 mg, 0.30 mmol), 2-
morpholinoethanamine (97 mg, 0.75 mmol), DCM (6 mL). MS ESI [M +
H].sup.+ 746.2, calcd for
[C.sub.34H.sub.38F.sub.3N.sub.7O.sub.5S.sub.2 + H].sup.+ 746.2
Step-02: Reagents (general method D): 1-(4-methoxybenzyl)-3-(5
and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4-((2- morpholinoethyl)amino)thieno[3,4-b]pyridin-2(1H)-one
(210 mg, 0.28 mmol), TFA (4 mL), conc HCl (0.5 mL). .sup.1H NMR
(Free base, 400 MHz, CD.sub.3OD) .delta. 8.37 (d, J = 2.8 Hz, 1H),
7.42 (d, J = 8.8 Hz, 1H), 7.14 (s, 1H), 6.98 (d, J = 7.2 Hz, 1H),
6.87 (d, J = 3.2 Hz, 1H), 4.05 (t, J = 6.0 Hz, 2H), 3.75 (br. s,
4H), 3.21 (br. s, 4H), 2.85 (t, J = 6.0 Hz, 2H), 2.68-2.64 (m, 8
H), 2.38 (s, 3H); MS ESI [M + H].sup.+ 494.2, calcd for
[C.sub.25H.sub.31N.sub.7O.sub.2S + H].sup.+ 494.2 A27:
4-(cyclobutylamino)- 3-(6-(4-methylpiperazin-1-
yl)-1H-benzo[d]imidazol-2- yl)thieno[3,4-b]pyridin- 2(1H)-one
##STR00071## 30 mg (20%); Yellow solid 2HCl Step-01: Reagents
(General Method C): 1-(4-methoxybenzyl)-3-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-2-oxo-1,2- dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate (229 mg, 0.30 mmol), cyclobutylamine (53
mg, 0.75 mmol), DCM (6 mL) MS ESI [M + H].sup.+ 687.1, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.4S.sub.2 + H].sup.+ 687.2
Step-02: Reagents (general method D):
4-(cyclobutylamino)-1-(4-methoxybenzyl)-3-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidaz-
ol-2- yl)thieno[3,4-b]pyridin-2(1H)-one (210 mg, 0.28 mmol), TFA (4
mL), conc, HCl (0.5 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta.
8.51 (d, J = 3.6 Hz, 1H), 7.70 (d, J = 9.2 Hz, 1H), 7.41 (dd, J =
9.2 Hz, 2.0 Hz, 1H), 7.31 (d, J = 2.0 Hz, 1H), 6.96 (d, J = 3.2 Hz,
1H), 3.99-3.96 (m, 2H), 3.71-3.68 (m, 2H), 3.63-3.59 (m, 1H),
3.39-3.35 (m, 2H), 3.28-3.22 (m, 2H), 3.01 (s, 3H), 2.23-2.21 (m,
2H), 2.06-2.00 (m, 2H), 1.74-1.66 (m, 1H), 1.49-1.38 (m, 1H); MS
ESI [M + H].sup.+ 435.2, calcd for [C.sub.23H.sub.26N.sub.6OS +
H].sup.+ 435.1 A28: 3-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-4- ((pyridin-2- ylmethyl)amino)thieno[3,4-
b]pyridin-2(1H)-one ##STR00072## 15 mg (13%); Pale yellow solid
2HCl Step-01: Reagents (general method C): 1-(4-methoxybenzyl)-3-(5
and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-2-oxo-1,2- dihydrothieno[3,4-b]pyridin-4-yl
trifluoromethanesulfonate (168 mg, 0.22 mmol), 2- Picolylamine (60
mg, 0.55 mmol), DCM (4 mL) MS ESI [M + H].sup.+ 724.1, calcd for
[C.sub.34H.sub.32F.sub.3N.sub.7O.sub.4S.sub.2 + H].sup.+ 724.2
Step-02: Reagents (general method D): 1-(4-methoxybenzyl)-3-(5
and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-4-((pyridin-2- ylmethyl)amino)thieno[3,4-b]pyridin-2(1H)-one
(65 mg, 0.09 mmol), TFA (3 mL), conc HCl (0.5 mL). .sup.1H NMR (400
MHz, CD.sub.3OD) .delta. 8.62 (d, J = 5.6 Hz, 1H), 8.49-8.43 (m,
2H), 7.97- 7.93 (m, 1H), 7.85 (d, J = 8.0 Hz, 1H), 7.62 (d, J = 9.2
Hz, 1H), 7.40 (dd, J = 9.2 Hz, 2.4 Hz, 1H), 7.23 (d, J = 2.0 Hz,
1H), 7.04 (d, J = 3.2 Hz, 1H), 4.84 (s, 2H), 4.00-3.96 (m, 2H),
3.70- 3.67 (m, 2H), 3.40-3.37 (m, 2H), 3.28-3.22 (m, 2H), 3.02 (s,
3H); MS ESI [M + H].sup.+ 472.2, calcd for
[C.sub.25H.sub.25N.sub.7OS + H].sup.+ 472.2 A29: 7-hydroxy-6-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00073## 6.6 mg (10%); Pale
yellow solid; TFA Step 1: Reagents (general method A1):
1-(4-methoxybenzyl)-1H-thieno[3,2-d][1,3]oxazine- 2,4-dione (0.75
g, 2.6 mmol), ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2- yl)acetate
(0.79 g, 2.6 mmol), KHMDS (13 mL, 13 mmol), THF (30 mL). .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. 13.64 (br. s, 1H), 12.64 (br. s.,
1H), 7.52 (br. s., 1H), 7.40-7.29 (m, 1H), 7.21 (d, J = 7.5 Hz,
2H), 7.04-6.88 (m, 3H), 6.84 (d, J = 8.0 Hz, 2H), 5.35 (br. s.,
2H), 3.76 (s, 3H), 3.20 (br. s., 4H), 2.62 (br. s., 4H), 2.39 (br.
s., 3 H); MS ESI [M + H].sup.+ 502.4, calcd for
[C.sub.27H.sub.27N.sub.5O.sub.3S + H].sup.+ 502.18. Step 2:
Reagents (general method D): 7-hydroxy-4-(4-methoxybenzyl)-6-(6-(4-
methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[3,2-b]pyridin-5(4H)--
one (0.090 g, 0.18 mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 13.52- 13.14 (m, 2 H), 11.25
(s, 1H), 9.79 (br. s., 1H), 7.81 (d, J = 5.3 Hz, 1H), 7.64 (d, J =
8.8 Hz, 1H), 7.37 (d, J = 2.3 Hz, 1H), 7.10 (dd, J = 9.0, 2.5 Hz,
1H), 6.92 (d, J = 5.0 Hz, 1H), 3.85-3.66 (m, 2H), 3.64-3.46 (m,
2H), 3.29-3.13 (m, 2H), 3.09-2.92 (m, 2H), 2.87 (s, 3H); MS ESI [M
+ H].sup.+ 382.3, calcd for [C.sub.19H.sub.19N.sub.5O.sub.2S +
H].sup.+ 382.45. A30: 7-amino-6-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00074##
1.5 g (79%) yellow solid free base A solution of
3-aminothiophene-2-carbonitrile (951 mg, 7.67 mmol) and
ethyl 2-(6-(4-
methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate (2.316 g,
7.67 mmol) in anhydrous THF (55 mL) was heated up to 40.degree. C.
in oil bath and LiHMDS (30.7 mL, 1.0M in THF, 30.7 mmol) was added
dropwise over 30 minutes. The resulting reaction mixture was
stirred at 40.degree. C. for 2 h then cooled down to rt and
quenched with satd aq NH.sub.4Cl in ice bath. The aqueous layer was
extracted with EtOAc. The combined organic layers were dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure. The
residue was triturated with DCM and filtered. The filter cake was
triturated again with MeOH then filtered to give the title compound
as a bright yellow solid (1.495 g, 79%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 12.34 (s, 1H), 11.81 (s, 1H), 10.73-10.47 (m,
1H), 7.96-7.91 (m, 1H), 7.90-7.78 (m, 1H), 7.52-7.43 (m, 1H),
7.09-7.25 (m, 1H), 7.00 (d, J = 5.3 Hz, 1H), 6.93-6.86 (m, 1H),
3.18-3.14 (m, 4H), 2.65-2.54 (m, 4H), 2.30 (s, 3H); MS ESI [M +
H].sup.+ 381.5, calcd for [C.sub.29H.sub.20N.sub.6OS + H].sup.+
381.1. A31: 7-amino-6-(6- morpholino-1H- benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00075## 16 mg (13%); white
solid; HCl LiHMDS (1.0M in THF, 1.7 mL, 1.7 mmol) was added
dropwise over 10 min to a stirred solution of
3-aminothiophene-2-carbonitrile (0.425 g, 0.34 mmol), ethyl
2-(6-morpholino-1H- benzo[d]imidazol-2-yl)acetate (0.103 g, 0.36
mmol) in anh THF (10 mL) at rt under Ar. The reaction was heated at
40.degree. C. for 1 h and directly purified by flash chromatography
(MeOH in CH.sub.2Cl.sub.2 0-10%) followed by prep HPLC. The
material was further recrystallized form EtOAc/hexanes and
triturated with MeOH. The resulting grey solid (25 mg) was
suspended in MeOH and treated with HCl (1.0M in Et.sub.2O, 0.14 mL)
at rt. The reaction was concentrated under reduced pressure Free
base: .sup.1H NMR (400 MHz, DMF-d.sub.7) .delta. 11.94 (s, 1H),
8.18 (m, 1H), 7.78 (d, J = 8.3 Hz, 1H), 7.52 (s, 1H), 7.35 (d, J =
5.2 Hz, 1H), 7.25 (d, J = 7.8 Hz, 1H), 4.06-4.00 (m, 4H), 3.40 (br
s, 4H). *three exchangeable protons are likely obscured by a peak
due to H.sub.2O and DMF-d.sub.7; MS ESI [M + H].sup.+ 368.2, calcd
for [C.sub.18H.sub.17N.sub.5O.sub.2S + H].sup.+ 368.1. A32:
7-amino-2-methyl-6-(6- (4-methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00076##
124 mg (44%); brown solid; TFA Reagents (general method A1):
3-amino-5-methylthiophene-2-carbonitrile (0.080 g, 0.58 mmol),
ethyl
2-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(0.18 g, 0.58 mmol), LDA (2.6 mL, 2.6 mmol), THF (5 mL). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 11.75 (br. s., 1H), 9.84 (br.
s., 1H), 7.53 (br. s., 1H), 7.24 (br. s., 1H), 6.98 (br. s., 1H),
6.77 (br. s., 1H), 3.82-3.39 (m, 7H), 3.22 (br. s., 2H), 3.08-2.76
(m, 5H), 2.56 (br. s., 3H); MS ESI [M + H].sup.+ 395.3, calcd for
[C.sub.20H.sub.22N.sub.6OS + H].sup.+ 395.5. A33:
7-amino-6-(6-(3r,5s)- rel-3,4,5-trimethylpiperazin-
1-yl)-1H-benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one
##STR00077## 43 mg (18%); yellow solid; 2HCl Reagents (general
method A1): ethyl
2-(6-((3r,5r)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)acetate (165 mg, 0.5 mmol),
3-aminothiophene-2-carbonitrile (124 mg, 1 mmol), LDA (1.0M in
THF/hex, 2.5 mL, 2.5 mmol), THF (8 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.99 (d, J = 1.6 Hz, 1H), 7.68 (d, J = 9.2 Hz,
1H), 7.38 (dd, J = 9.0, 2.2 Hz, 1H), 7.31 (d, J = 2.0 Hz, 1H), 7.11
(d, J = 5.6 Hz, 1H), 4.03-3.95 (m, 2H), 3.65-3.55 (m, 2H),
3.10-3.04 (m, 2H), 3.03 (s, 3H), 1.56 (d, J = 6.4 Hz, 6H); MS ESI
[M + H].sup.+ 409.3, calcd for [C.sub.21H.sub.24N.sub.6OS +
H].sup.+ 409.17. A34: 7-(cyclopentylamino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00078## 77 mg (26%); brown
solid; TFA Step 1: Reagents (general method C): a mixture or
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-5-oxo-4,5- dihydro-thieno[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (0.41 g, 0.54 mmol), cyclopentylamine
(0.13 mL, 1.3 mmol), MeCN (10 mL). MS ESI [M + H].sup.+ 701.3,
calcd for [C.sub.33H.sub.35F.sub.3N.sub.6O.sub.4S.sub.2 + H].sup.+
701.2. Step 2: Reagents (general method D): a mixture of
7-(cyclopentylamino)-4-(4- methoxybenzyl)-6-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one (0.38 g, 0.54
mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 12.99 (br. s, 1H), 12.11 (br. s, 1H), 11.87
(s, 1H), 9.65 (br. s, 1H), 8.04 (d, J = 5.5 Hz, 1H), 7.61-7.40 (m,
1H), 7.32-7.13 (m, 1H), 7.04 (d, J = 5.5 Hz, 1H), 6.93 (dd, J =
8.5, 2.5 Hz, 1H), 4.72-4.60 (m, 1H), 3.80-3.69 (m, 2H), 3.59-3.52
(m, 2H), 3.29-3.15 (m, 2H), 3.02-2.90 (m, 2H), 2.88 (d, J = 3.5 Hz,
3H), 2.21-2.06 (m, 2H), 1.93-1.67 (m, 6H); MS ESI [M + H].sup.+
449.3, calcd for [C.sub.24H.sub.28N.sub.6OS + H].sup.+ 449.2. A35:
6-(6-(4- methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7-
((tetrahydro-2H-pyran-4- yl)amino)thieno[3,2- b]pyridin-5(4H)-one
##STR00079## 69 mg (52%); yellow solid; TFA Step 1: Reagents
(general method C): a mixture of 4-(4-methoxybenzyl)-6-(5 and/or
6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-5-oxo-4,5- dihydro-thieno[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (0.18 g, 0.23 mmol), tetrahydro-
2H-pyran-4-amine (0.058 mL, 0.58 mmol), MeCN (5 mL). MS ESI [M +
H].sup.+ 717.3, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D): a mixture of
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-
methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2--
yl)-7-
((tetrahydro-2H-pyran-4-yl)amino)thieno[3,2-b]pyridin-5(4H)-one
(0.17 g, 0.23 mmol), TFA (7 mL), and conc. HCl (1 mL). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 13.02 (br. s, 1H), 12.18 (br. s,
1H), 11.92 (s, 1H), 9.63 (br. s, 1H), 8.04 (d, J = 5.5 Hz, 1H),
7.54 (d, J = 8.0 Hz, 1H), 7.30-7.14 (m, 1 H), 7.04 (d, J = 5.5 Hz,
1H), 6.96 (dd, J = 8.3, 2.3 Hz, 1H), 4.49-4.34 (m, 1H), 4.02-3.93
(m, 2H), 3.81-3.74 (m, 2H), 3.57 (d, J = 9.0 Hz, 4H), 3.31-3.15 (m,
2H), 3.02-2.91 (m, 2H), 2.88 (d, J = 3.8 Hz, 3H), 2.13 (d, J = 3.3
Hz, 2H), 1.81-1.64 (m, 2H); MS ESI [M + H].sup.+ 465.3, calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2. A36:
7-(((1R*,3S*)-3- hydroxycyclopentyl)amino)-
6-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00080## 33 mg (28%); yellow
solid; TFA Step 1: Reagents (general method C): a mixture of
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-5-oxo-4,5- dihydrothieno[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (0.15 g, 0.20 mmol), (1S*,3R*)-3-
aminocyclopentanol (0.070 g, 0.50 mmol), DCM (10 mL). MS ESI [M +
H].sup.+ 717.3, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D): a mixture of 7-(((1R*,3S*)-3-
hydroxycyclopentyl)amino)-4-(4-methoxybenzyl)-6-(5 and/or
6-(4-methylpiperazin-1-yl)-1-
((trifluoromethyl)sulfonyl)-1H-benzo[d]-imidazol-2-yl)thieno[3,2-b]pyridin-
-5(4H)-one (0.14 g, 0.20 mmol), TFA (7 mL), and conc. HCl (1 mL).
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 8.04 (d, J = 5.5 Hz, 1H),
7.61 (d, J = 9.0 Hz, 1H), 7.29-7.20 (m, 2H), 7.11 (d, J = 5.5 Hz,
1H), 4.82-4.73 (m, 1H), 4.57-4.47 (m, 1H), 3.96-3.79 (m, 2H),
3.74-3.58 (m, 2H), 3.42-3.33 (m, 2H), 3.22- 3.08 (m, 2H), 3.01 (s,
3H), 2.33-2.19 (m, 2H), 2.16-2.02 (m, 3H), 2.02-1.88 (m, 1H); MS
ESI [M + H].sup.+ 465.2, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S
+ H].sup.+ 465.2. A37: 7-(((1r,4r)-4- hydroxycyclohexyl)amino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00081## 10 mg (9%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate(crude,
0.24 mmol), trans-4-aminocyclohexanol (0.11 g, 0.96 mmol), MeCN (10
mL). MS ESI [M + H].sup.+ 713.2 calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: (general method D):
7-(((1r,4r)-4-hydroxycyclohexyl)amino)-4-(4-methoxybenzyl)-6- (5
and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benz-
o[d]imidazol-2- yl)thieno[3,2-b]py-ridin-5(4H)-one (crude, 0.24
mmol), TFA (7 mL), HCl (1 mL); .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.88 (d, J = 5.5 Hz, 1H), 7.51-7.41 (m, 1H), 7.24-7.14 (m,
1H), 7.08 (d, J = 5.5 Hz, 1H), 7.03-6.95 (m, 1H), 4.32-4.21 (m,
1H), 3.82-3.71 (m, 1H), 3.28-3.20 (m, 4H), 2.86-2.74 (m, 4H), 2.47
(s, 3H), 2.24-2.36 (m, 2H), 2.17-2.06 (m, 2H), 1.78-1.63 (m, 2H),
1.62-1.49 (m, 2H); MS ESI [M + H].sup.+ 479.2, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.3. A38: 6-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7-
(((tetrahydro-2H-pyran-4- yl)methyl)amino)thieno[3,2-
b]pyridin-5(4H)-one ##STR00082## 24 mg (32%) yellow solid Free base
Step 1: Reagents (general method C): 4-(4-methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.16 mmol), 4-aminomethyltetrahydropyran (0.074 g, 0.64 mmol), MeCN
(10 mL). MS ESI [M + H].sup.+ 731.3, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: Reagents (general method D): 4-(4-methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-7-(((tetrahy-
dro-2H-pyran-4- yl)methyl)amino)-thieno[3,2-b]pyridin-5(4H)-one
(crude, 0.16 mmol), TFA (7 mL), HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.93-7.83 (m, 1H), 7.49-7.40 (m, 1H), 7.21-7.13
(m, 1H), 7.12-7.06 (m, 1H), 7.05-6.96 (m, 1H), 4.10-3.97 (m, 2H),
3.89-3.79 (m, 2H), 3.56-3.44 (m, 2H), 3.26-3.16 (m, 4H), 2.77-2.65
(m, 4H), 2.40 (s, 3H), 2.19-2.07 (m, 1H), 1.99-1.88 (m, 2H),
1.65-1.50 (m, 2H); MS ESI [M + H].sup.+ 479.3, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2. A39: 6-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7- (piperidin-4-
ylamino)thieno[3,2-b]pyridin- 5(4H)-one ##STR00083## 17 mg (19%)
Brown solid Free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.20 mmol), tert-butyl 4-aminopiperidine-1- carboxylate (0.16 g,
0.80 mmol), MeCN (10 mL). MS ESI [M + H].sup.+ 816.2, calcd for
[C.sub.38H.sub.44F.sub.3N.sub.7O.sub.6S.sub.2 + H].sup.+ 816.2 Step
2: Reagents (general method D): tert-butyl
4-((4-(4-methoxybenzyl)-6-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-5-oxo-4,5-
dihydrothieno[3,2-b]pyridin-7-yl)amino)piperidine-1-carboxylate
(crude,
0.20 mmol), TFA (6 mL), HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.90-7.83 (m, 1H), 7.52-7.42 (m, 1H), 7.25-7.11
(m, 1H), 7.10-7.05 (m, 1H), 7.04-6.96 (m, 1H), 4.50-4.42 (m, 1 H),
3.27-3.16 (m, 6H), 2.91-2.79 (m, 2H), 2.73-2.61 (m, 4H), 2.37 (s,
3H), 2.26-2.17 (m, 2H), 1.89-1.73 (m, 2H); MS ESI [M + H].sup.+
464.2, calcd for [C.sub.24H.sub.29N.sub.7OS + H].sup.+ 464.2. A40:
7-(((1S,4S)-4- hydroxycyclohexyl)amino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00084## 35 mg (30%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.24 mmol), cis-4-aminocyclohexanol (0.11 g, 0.96 mmol), MeCN (10
mL). MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2 Step
2: (general method D):
7-(((1S,4S)-4-hydroxycyclohexyl)amino)-4-(4-methoxybenzyl)-6- (5
and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benz-
o[d]imidazol-2- yl)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.24
mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.88-7.82 (m, 1H), 7.56-7.41 (m, 1H), 7.29-7.14 (m, 1H),
7.08 (d, J = 5.2 Hz, 1H), 7.04-6.96 (m, 1H), 4.50-4.40 (m, 1H),
3.90-3.80 (m, 1H), 3.26-3.17 (m, 4H), 2.73- 2.63 (m, 4H), 2.38 (s,
3H), 2.13-1.81 (m, 8H); MS ESI [M + H].sup.+ 479.2, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2 A41:
7-(((1S,2S)-2- hydroxycyclohexyl)amino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00085## 16 mg (23%), yellow
solid; free base Step 1: (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-1-yl)-1-
((trifluoro-methyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-dihydroth-
ieno[3,2- b]pyridin-7-yl trifluoro-methanesulfonate (crude, 0.20
mmol), (1S,2S)-2-aminocyclohexanol (0.091 g, 0.80 mmol), DMF (7
mL). MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H] 731.2 Step 2:
Reagents (general method D):
7-(((1S,2S)-2-hydroxycyclohexyl)amino)-4-(4- methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno-[3,2-b]pyridin-5(4H)-one (crude, 0.14
mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.84 (d, J = 5.5 Hz, 1H), 7.55-7.40 (m, 1H), 7.18-7.12 (m,
1H), 7.06 (d, J = 5.5 Hz, 1H), 7.03-6.94 (m, 1H), 4.24-4.14 (m,
1H), 3.88-3.77 (m, 1H), 3.27-3.18 (m, 4H), 2.81-2.70 (m, 4H), 2.44
(s, 3H), 2.33-2.23 (m, 1H), 2.20-2.10 (m, 1H), 1.91-1.78 (m, 2H),
1.65-1.42 (m, 4H); MS ESI [M + H].sup.+ 479.3, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.3 A42:
7-(((1S,2S)-2- hydroxycyclopentyl)amino)-
6-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00086## 20 mg (13%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.32 mmol), (1S,2S)-2-aminocyclopentanol (0.13 g, 1.3 mmol), DMF (7
mL). MS ESI [M + H].sup.+ 717.2 calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2 Step
2: Reagents (general method D):
7-(((1S,2S)-2-hydroxycyclopentyl)amino)-4-(4- methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno-[3,2-b]pyridin-5(4H)-one (crude, 0.32
mmol), TFA (5 mL), HCl (5 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.90-7.84 (m, 1H), 7.48-7.40 (m, 1H), 7.20-7.11 (m, 1H),
7.07 (d, J = 5.2 Hz, 1H), 7.30-6.93 (m, 1H), 4.61-4.53 (m, 1H),
4.42-4.34 (m, 1H), 3.27- 3.15 (m, 4H), 2.76-2.64 (m, 4H), 2.45-2.32
(m, 4H), 2.30-2.17 (m, 1H), 2.06-1.94 (m, 2H), 1.93-1.82 (m, 1H),
1.82-1.70 (m, 1H); MS ESI [M + H].sup.+ 465.2 calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2 A43:
6-(6-morpholino-1H- benzo[d]imidazol-2-yl)-7- (piperidin-4-
ylamino)thieno[3,2-b]pyridin- 5(4H)-one ##STR00087## 11 mg (12%),
brown solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-morpholino-1-
((trifluoro-methyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-dihydroth-
ieno[3,2- b]pyridin-7-yl trifluoro-methanesulfonate (crude, 0.20
mmol), tert-butyl 4-aminopiperidine-1- carboxylate (0.16 g, 0.8
mmol), DMF (7 mL). MS ESI [M + H].sup.+ 803.2, calcd for
[C.sub.37H.sub.41F.sub.3N.sub.6O.sub.7S.sub.2 + H].sup.+ 803.2.
Step 2: Reagents (general method D): tert-butyl
4-((4-(4-methoxybenzyl)-6-(5 and/or 6-
morpholino-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4-
,5-
dihydrothieno[3,2-b]pyridin-7-yl)-amino)piperidine-1-carboxylate
(crude, 0.20 mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.90-7.84 (m, 1H), 7.52-7.42 (m, 1H), 7.23-7.13
(m, 1H), 7.08 (d, J = 5.2 Hz, 1H), 7.04-6.95 (m, 1H), 4.54-4.43 (m,
1H), 3.93-3.83 (m, 4H), 3.29-3.24 (m, 2H), 3.20-3.10 (m, 4H),
2.98-2.86 (m, 2H), 2.29-2.18 (m, 2H),, 1.90- 1.78 (m, 2H); MS ESI
[M + H].sup.+ 451.3, calcd for [C.sub.23H.sub.26N.sub.6O.sub.2S +
H].sup.+ 451.2. A44: 7-(((1S,2R)-2- hydroxycyclohexyl)amino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00088## 20 mg (21%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.20 mmol), (1R,2S)-2-aminocyclohexanol (0.091 g, 0.80 mmol), DMF
(7 mL). MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: Reagents (general method D):
7-(((1S,2R)-2-hydroxycyclohexyl)amino)-4-(4- methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno-[3,2-b]pyridin-5(4H)-one (crude, 0.20
mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.84 (d, J = 5.8 Hz, 1H), 7.56-7.40 (m, 1H), 7.29-7.13 (m,
1H), 7.07 (d, J = 5.5 Hz, 1H), 7.03-6.94 (m, 1H), 4.56-4.46 (m,
1H), 4.15-4.04 (m, 1H), 3.27-3.13 (m, 4H), 2.79-2.64 (m, 4H), 2.41
(s, 3H), 2.14-1.95 (m, 2H), 1.91-1.74 (m, 4H), 1.60-1.44 (m, 2H);
MS ESI [M + H].sup.+ 479.2, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2. A45:
7-((1-methylpiperidin-4- yl)amino)-6-(6-morpholino-
1H-benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one
##STR00089## 33 mg (30%), brown solid; free base Step 1: Reagents
(general method C): 4-(4-methoxybenzyl)-6-(5 and/or 6-morpholino-1-
((trifluoro-methyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-dihydroth-
ieno[3,2- b]pyridin-7-yl trifluoro-methanesulfonate (crude, 0.20
mmol), tert-butyl 4-aminopiperidine-1- carboxylate (0.16 g, 0.8
mmol), DMF (7 mL). MS ESI [M + H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D):
4-(4-methoxybenzyl)-7-((1-methylpiperidin-4- yl)amino)-6-(5 and/or
6-morpholino-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.24 mmol), TFA (5 mL),
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.91-7.84 (m,
1H), 7.53-7.43 (m, 1H), 7.22-7.13 (m, 1H), 7.08 (d, J = 5.5 Hz,
1H), 7.05-6.94 (m, 1H), 4.48-4.33 (m, 1H), 3.93-3.83 (m, 4H),
3.20-3.10 (m, 4H), 3.02-2.87 (m, 2H), 2.61-2.43 (m, 2H), 2.40 (s,
3H), 2.31-2.16 (m, 2H), 2.01-1.87 (m, 2H); MS ESI [M + H].sup.+
465.2, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2.
A46: 7-(((1R,2S)-2- hydroxycyclohexyl)amino)-6-
(6-(5-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00090## 12 mg (4%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.20 mmol), (1S,2R)-2-aminocyclohexanol (0.091 g, 0.80 mmol), DMF
(7 mL). MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: Reagents (general method D):
7-(((1R,2S)-2-hydroxycyclohexyl)amino)-4-(4- methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno-[3,2-b]pyridin-5(4H)-one (crude, 0.20
mmol), TFA (5 mL), HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.82 (d, J = 5.2 Hz, 1H), 7.55-7.38 (m, 1H), 7.29- 7.10 (m,
1H), 7.05 (d, J = 5.5 Hz, 1H), 7.01-6.93 (m, 1H), 4.55-4.43 (m,
1H), 4.15-4.05 (m, 1H), 3.22 (br s, 4H), 2.70 (br s, 4H), 2.39 (s,
3H), 2.14-1.96 (m, 3H), 1.92-1.73 (m, 5H); MS ESI [M + H].sup.+
479.2, calcd for [C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2
A47: 7-(cyclopentylamino)-6- (6-(3r,5s)-rel-3,4,5-
trimethylpiper-azin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00091## 21 mg (18%); brown
solid; TFA Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-5-oxo-6-(1- ((trifluoromethyl)sulfonyl)-5
and/or 6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-4,5-dihydrothieno[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (crude, 0.2 mmol), cyclopentylamine (0.1
mL), DMF (6 mL). MS ESI [M + H].sup.+ 729.2, calcd for
[C.sub.35H.sub.39F.sub.3N.sub.6O.sub.4S.sub.2 + H].sup.+ 729.2.
Step 2: Reagents (general method D):
7-(cyclopentylamino)-4-(4-methoxybenzyl)-6-(1-
((trifluoro-methyl)sulfonyl)-(5 and/or
6-((3s,5r)-rel-3,4,5-trimethylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)-thieno[3,2-b]pyridin-5(4H)-one (crude, 0.2
mmol), TFA (6 mL), and conc. HCl (0.5 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 7.94 (d, J = 5.6 Hz, 1H), 7.56 (d, J = 8.8 Hz,
1H), 7.26-7.33 (m, 1H), 7.18-7.14 (m, 1H), 7.09 (d, J = 5.6 Hz,
1H), 4.67-4.60 (m, 1H), 3.92-3.85 (m, 2H), 3.60-3.50 (m, 2H),
3.05-2.88 (m, 5H), 2.22-2.13 (m, 2H), 1.94-1.70 (m, 6H), 5.10 (d, J
= 6.0 Hz, 6H); MS ESI [M + H].sup.+ 477.3, calcd for
[C.sub.26H.sub.32N.sub.6OS + H].sup.+ 477.2. A48:
7-((tetrahydro-2H- pyran-4-yl)amino)-6-(6- (3r,5s)-rel-3,4,5-
trimethylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00092## 19 mg (16%); yellow
solid; TFA Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-5-oxo-5 and/or 6-(1-
((trifluoromethyl)-sulfonyl)-6-((3r,5s)-rel-3,4,5-trimethylpiperazin-1-yl)-
-1H- benzo[d]imidazol-2-yl)-4,5-dihydrothieno-[3,2-b]pyridin-7-yl
trifluoromethanesulfonate (crude, 0.2 mmol),
tetrahydro-2H-pyran-4-amine (0.1 mL), DMF (6 mL). MS ESI [M +
H].sup.+ 745.1, calcd for
[C.sub.35H.sub.39F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 745.2.
Step 2: Reagents (general method D):
4-(4-methoxybenzyl)-7-((tetrahydro-2H-pyran-4- yl)amino)-(5 and/or
6-(1-((trifluoromethyl)sulfonyl)-6-((3s,4r)-rel-3,4,5-trimethylpiperazin--
1- yl)-1H-benzo[d]imi-dazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one
(crude, 0.2 mmol) TFA (6 mL), and conc. HCl (0.5 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.95 (d, J = 5.6 Hz, 1H), 7.60 (d, J
= 8.8 Hz, 1H), 7.28 (d, J = 2.0 Hz, 1H), 7.21 (dd, J = 8.8, 1.6 Hz,
1H), 7.10 (d, J = 5.6 Hz, 1H), 4.40-4.30 (m, 1H), 4.08-4.02 (m,
2H), 3.95-3.87 (m, 2H), 3.63-3.53 (m, 4H), 3.03 (s, 3H), 3.02-2.93
(m, 2H), 2.18-2.11 (m, 2H), 1.86-1.76 (m, 2H), 1.52 (d, J = 6.4 Hz,
6H); MS ESI [M + H].sup.+ 493.3, calcd for
[C.sub.26H.sub.32N.sub.6O.sub.2S + H].sup.+ 493.2. A49: 6-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7-
morpholinothieno[3,2- b]pyridin-5(4H)-one ##STR00093## 134 mg
(79%); yellow solid; TFA salt Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.3 mmol), morpholine (0.08 mL, 0.897 mmol). MS ESI [M + H].sup.+
703.2, calcd for [C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 +
H].sup.+ 703.2. Step 2: Reagents (general method D):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-(((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-7-morpholin-
othieno[3,2- b]pyridin-5(4H)-one (crude, 0.3 mmol), TFA (5 mL),
conc. HCl (1 mL). .sup.1H NMR (400 MHz, DMSO-d.sub.6) .delta. 12.46
(s, 1H), 9.98 (br. s, 1H), 8.17 (d, J = 5.6 Hz, 1H), 7.71 (d, J =
9.2 Hz, 1H), 7.33 (dd, J = 9.2, 1.2 Hz, 1H), 7.24 (d, J = 2.0 Hz,
1H), 7.08 (d, J = 5.6 Hz, 1H), 3.92-3.84 (m, 2H), 3.70-3.62 (m,
4H), 3.62-3.57 (m, 2H), 3.35-3.28 (m, 4H), 3.28-3.13 (m, 2H),
3.11-2.99 (m, 2H), 2.89 (s, 3H); MS ESI [M + H].sup.+ 451.3, calcd
for [C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.2. A50:
7-(4-hydroxypiperidin- 1-yl)-6-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00094##
126 mg (73%); yellow solid; TFA salt Step 1: Reagents (general
method C): 4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.30 mmol), piperidin-4-ol (91 mg, 0.897 mmol), TFA (4 mL), conc.
HCl (1 mL). MS ESI [M + H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D):
7-(4-hydroxypiperidin-1-yl)-4-(4-methoxybenzyl)-6-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d-
]imidazol-2- yl)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.3 mmol),
TFA (5 mL), conc. HCl (1 mL). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. 14.26 (br. s, 1H), 12.33 (s, 1H), 10.04 (br. s, 1H), 8.14
(s, 1H), 7.70 (d, J = 7.28 Hz, 1H), 7.36-7.28 (m, 1H), 7.23 (br.
s., 1H), 7.06 (s, 1H), 3.98-3.83 (m, 2H), 3.75-3.66 (m, 2H),
3.66-3.39 (m, 3H), 3.25-3.10 (m, 4H), 3.10-2.97 (m, 2H), 2.73 (s,
3H), 1.86-1.72 (m, 2H), 1.54-1.39 (m, 2H); MS ESI [M + H].sup.+
465.3, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2.
A51: 7-(((1R,2S)-2- hydroxycyclohexyl)amino)-6-
(6-(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[3,2-b]pyridin- 5(4H)-one ##STR00095## 15 mg (15%), yellow
solid; free base Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5 and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.20 mmol), (1S,2R)-2-aminocyclohexanol (0.091 g, 0.80 mmol), DMF
(7 mL). MS ESI [M + H].sup.+ 731.2; calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: Reagents (General Method D):
7-(((1R,2S)-2-hydroxycyclohexyl)amino)-4-(4- methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.20
mmol), TFA (5 mL), HCl (1 mL) .sup.1H NMR (400 MHz, MeOD-d.sub.4)
.delta. 7.82 (d, J = 5.2 Hz, 1H), 7.55-7.38 (m, 1H), 7.29- 7.10 (m,
1H), 7.05 (d, J = 5.5 Hz, 1H), 7.01-6.93 (m, 1H), 4.55-4.43 (m,
1H), 4.15-4.05 (m, 1H), 3.22 (br s, 4H), 2.70 (br s, 4H), 2.39 (s,
3H), 2.14-1.96 (m, 3H), 1.92-1.73 (m, 5H); MS ESI [M + H].sup.+
479.2; calcd for [C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2.
A52: 6-(6-(4- methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7-
((pyridin-3- ylmethyl)amino)thieno[3,2- b]pyridin-5(4H)-one
##STR00096## 57 mg (27%); yellow solid; TFA salt Step 1: Reagents
(general method C): 4-(4-methoxybenzyl)-6-(5 and/or
6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.3 mmol), pyridin-3-ylmethanamine (0.09 mL, 0.90 mmol). MS ESI [M
+ H].sup.+ 724.2, calcd for
[C.sub.34H.sub.32F.sub.3N.sub.7O.sub.4S.sub.2 + H].sup.+ 724.2.
Step 2: Reagents (general method D): 4-(4-methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-7-((pyridin--
3- ylmethyl)amino)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.3 mmol),
TFA (4 mL), conc. HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 8.92-8.81 (m, 1H), 8.78-8.67 (m, 1H), 8.56-8.45 (m, 1H),
8.00-7.85 (m, 2H), 7.62-7.50 (m, 1H), 7.30-7.21 (m, 1H), 7.20-7.13
(m, 1H), 7.12-7.03 (m, 1H), 5.31 (s, 2H), 3.91-3.77 (m, 2H),
3.73-3.57 (m, 2H), 3.41-3.22 (m, 2H), 3.20-3.07 (m, 2H), 3.01 (s,
3H); MS ESI [M + H].sup.+ 472.3, calcd for
[C.sub.25H.sub.25N.sub.7OS + H].sup.+ 472.2. A53: 6-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-7-
((pyridin-2-ylmethyl)amino)- thieno[3,2-b]pyridin-5(4H)- one
##STR00097## 100 mg (48%); yellow solid; TFA salt Step 1: Reagents
(general method C): 4-(4-methoxybenzyl)-6-(5 and/or
6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-di-
hydrothieno[3,2- b]pyridin-7-yl trifluoromethanesulfonate (crude,
0.3 mmol), pyridin-2-ylmethanamine (0.09 mL, 0.897 mmol). MS ESI [M
+ H].sup.+ 724.2, calcd for
[C.sub.34H.sub.32F.sub.3N.sub.7O.sub.4S.sub.2 + H].sup.+ 724.2.
Step 2: Reagents (general method D): 4-(4-methoxybenzyl)-6-(5
and/or 6-(4-methylpiperazin-
1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-7-((pyridin--
2- ylmethyl)amino)thieno[3,2-b]pyridin-5(4H)-one (crude, 0.3 mmol),
TFA (4 mL), conc. HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 8.67 (d, J = 5.5 Hz, 1H), 8.05 (t, J = 7.4 Hz, 1H), 7.94
(d, J = 5.5 Hz, 1H), 7.68 (d, J = 5.5 Hz, 1H), 7.63 (d, J = 8.8 Hz,
1H), 7.58-7.51 (m, 1H), 7.25-7.21 (m, 2H), 7.08 (d, J = 5.5 Hz,
1H), 5.25 (s, 2H), 3.95-3.84 (m, 2H), 3.72-3.62 (m, 2H), 3.41-3.33
(m, 2H), 3.21-3.10 (m, 2H), 3.01 (m, 3H). MS ESI [M + H].sup.+
472.3, calcd for [C.sub.25H.sub.25N.sub.7OS + H].sup.+ 472.2. A54:
4-(((3S,4S)-3- fluoropiperidin-4- yl)amino)-5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00098## 113 mg (53%);
yellow solid; 2 TFA Step 1: Reagents (general method C): a mixture
of 7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.23 g, 0.30 mmol), (3S,4S)-tert- butyl
4-amino-3-fluoropiperidine-1-carboxylate (0.20 g, 0.90 mmol), DMF
(5 mL). MS ESI [M - CF.sub.3O.sub.2S + 2H].sup.+ 702.2, calcd for
[C.sub.37H.sub.44FN.sub.7O.sub.4S + H].sup.+ 702.32. Step 2:
Reagents (general method D): a mixture of (3S,4S)-tert-butyl
3-fluoro-4-((7-(4- methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imi-dazol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl)amino)p-
iperidine-1- carboxylate (0.23 g, 0.30 mmol), TFA (7 mL), and conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.55 (d, J =
8.8 Hz, 1H), 7.48 (d, J = 6.0 Hz, 1H), 7.24-7.16 (m, 2H), 7.10 (d,
J = 8.4 Hz, 1H), 5.12-5.07 (m, 1H), 5.03-4.99 (m, 2H), 4.41 (br. s,
1H), 3.90-3.56 (m, 6H), 3.52- 3.43 (m, 1H), 3.32-3.27 (m, 1H),
3.23-3.09 (m, 2H), 3.01 (s, 3H), 2.52-2.41 (m, 1H), 2.21-2.09 (m,
1H); MS ESI [M + H].sup.+ 482.2, calcd for
[C.sub.24H.sub.28FN.sub.7OS + H].sup.+ 482.2. A55: 4-(((3R*,4S*)-3-
fluoropiperidin-4- yl)amino)-5-(6-(4- methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00099##
114 mg (54%); brown solid; 2 TFA Step 1: Reagents (general method
C): a mixture of 7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.23 g, 0.30 mmol), (3R*,4S*)-
tert-butyl 4-amino-3-fluoropiperidine-1-carboxylate (0.20 g, 0.90
mmol), DMF (5 mL). MS ESI [M - CF.sub.3O.sub.2S + 2H].sup.+ 702.2,
calcd for [C.sub.37H.sub.44FN.sub.7O.sub.4S + H].sup.+ 702.32. Step
2: Reagents (general method D): a mixture of (3R*,4S*)-tert-butyl
3-fluoro-4-((7-(4- methoxybenzyl)-5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imi-dazol-2-yl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-4-yl)amino)p-
iperidine-1- carboxylate (0.23 g, 0.30 mmol), TFA (7 mL), and conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.57 (d, J =
8.8 Hz, 1H), 7.52 (d, J = 6.0 Hz, 1H), 7.24 (d, J = 2.1 Hz, 1H),
7.21 (d, J = 5.9 Hz, 1H), 7.15 (dd, J = 2.1, 8.9 Hz, 1H), 5.19 (d,
J = 46.7 Hz, 1H), 4.43-4.27 (m, 1H), 3.78 (d, J = 12.6 Hz, 3H),
3.72-3.57 (m, 2H), 3.49 (d, J = 13.9 Hz, 2 H), 3.41 (d, J = 14.2
Hz, 1H), 3.23 (d, J = 3.5 Hz, 4H), 3.00 (s, 3H), 2.42-2.23 (m, 2H);
MS ESI [M + H].sup.+ 482.2, calcd for [C.sub.24H.sub.28FN.sub.7OS +
H].sup.+ 482.2. A56: 4-amino-5-(5-fluoro- 6-morpholino-1H-benzo[d]-
imidazol-2-yl)-thieno- [2,3b]-pyridin-6(7H)-one ##STR00100## 40 mg
(16%); Light brown solid HCl Reagents (general method-A1): ethyl
2-(5-fluoro-6-morpholino-1H-benzo[d]imidazol-2- yl)acetate (186 mg,
0.60 mmol), 2-amino-2-cyanothiophene (75 mg, 0.60 mmol), LDA (3.0
mL, 1M in THF/hex, 3.0 mmol) anh THF (6.0 mL); .sup.1H NMR (400
MHz, CD.sub.3OD) .delta. 7.58 (d, J = 11.6 Hz, 1H), 7.55-7.51 (m,
2H), 7.21 (d, J = 6.0 Hz, 1H), 3.97-3.94 (m, 4H), 3.30-3.27 (m,
4H); MS ESI [M + H].sup.+ 386.2, calcd for
[C.sub.18H.sub.16FN.sub.5O.sub.2S + H].sup.+ 386.1. A57:
4-(((1S,2R)-2- hydroxy- cyclohexyl)amino)-5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-
yl)thieno-[2,3-b]pyridin- 6(7H)-one ##STR00101## 36 mg (25%),
Yellow solid; Free base Step 1: (General Method C)
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-methylpiperazin-1-yl)-1-
((trifluoro-methyl)-sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-oxo-6,7-dihydrot-
hieno[2,3- b]pyridin-4-yl trifluoro-methanesulfonate (crude, 0.30
mmol), (1R,2S)-2-aminocyclohexanol (0.14 g, 1.2 mmol), DMF (7 mL).
MS ESI [M + H].sup.+ 731.2, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ calcd
731.2 Step 2: (General Method D)
4-(((1S,2R)-2-hydroxycyclohexyl)amino)-7-(4-methoxybenzyl)-
5-(5 and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imida-
zol-2- yl)thieno[2,3-b]pyridin-6(7H)-one (crude, 0.30 mmol), TFA (5
mL), HCl (1 mL); .sup.1H NMR (400 MHz, MeOD-d.sub.4) .delta. ppm
7.53 (d, J = 5.99 Hz, 1 H), 7.48 (br. s, 1 H), 7.20 (br. s., 1 H),
7.12 (d, J = 5.99 Hz, 1 H), 6.97-7.04 (m, 1 H), 4.38-4.47 (m, 1 H),
4.04-4.11 (m, 1 H), 3.19- 3.28 (m, 4 H), 2.65-2.75 (m, 4 H), 2.40
(s, 3 H), 2.08-2.17 (m, 1 H), 2.00-2.08 (m, 1 H), 1.75-1.90 (m, 4
H), 1.46-1.61 (m, 2 H); MS ESI [M + H].sup.+ 479.2, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2. A58:
4-(((1S,2R)-2- hydroxy- cyclopentyl)amino)-5-(6-(4-
methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-
thieno[2,3-b]pyridin-6(7H)- one ##STR00102## 23 mg (17%), yellow
solid; Free base Step 1: (general method C)
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-methylpiperazin-1-yl)-1-
((trifluoromethyl)-sulfonyl)-1H-benzo[d]imidazol-2-yl)-6-oxo-6,7-dihydroth-
ieno[2,3- b]pyridin-4-yl trifluoromethane-sulfonate (crude, 0.30
mmol), (1R,2S)-2-aminocyclopentanol (0.10 g, 1.2 mmol), DMF (7 mL).
MS ESI [C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+
calcd 717.2, observed 717.2 Step 2: (General Method D)
4-(((1S,2R)-2-hydroxycyclopentyl)amino)-7-(4-methoxybenzyl)- 5-(5
and/or
6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imida-
zol-2- yl)thieno[2,3-b]pyridin-6(7H)-one (crude, 0.30 mmol), TFA (5
mL), HCl (1 mL); .sup.1H NMR (400 MHz, MeOD-d.sub.4) .delta. ppm
7.55 (d, J = 5.99 Hz 1 H), 7.42-7.51 (m, 1 H), 7.12-7.25 (m, 1 H),
7.09 (d, J = 5.87 Hz, 1 H), 6.94-7.01 (m, 1 H), 4.43-4.51 (m, 1 H),
4.32-4.39 (m, 1 H), 3.16-3.26 (m, 4 H), 2.62-2.74 (m, 4 H), 2.38
(s, 3 H), 2.17-2.26 (m, 1 H), 1.89-2.15 (m, 4 H), 1.69-1.81 (m, 1
H); MS ESI [M + H].sup.+ 465.2, calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2 A59:
4-amino-5-(6-(4- methyl-1,4-diazepan-1-yl)- 1H-benzo-[d]imidazol-2-
yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00103## 28 mg (7%); brown
solid; TFA Reagents (General method A2): ethyl
2-(6-(4-methyl-1,4-diazepan-1-yl)-1H- benzo[d]imidazol-2-yl)acetate
(255 mg, 0.88 mmol), 2-aminothiophene-3-carbonitrile (100 mg, 0.8
mmol), LDA (1.0M in THF/hex, 4 mL, 4 mmol), THF (10 mL), 45.degree.
C., 2 h. 82 mg of mixture of uncyclized and cyclized was obtained
which was recyclized with KOBu.sup.t (1.0M in THF, 1.0 mL, 1 mmol)
in THF (10 mL), rt, 30 min then 35.degree. C., 1 h; .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 6.23 (d, J = 9.2 Hz, 1H), 7.52 (d, J
= 5.6 Hz, 1H), 7.19 (d, J = 5.6 Hz, 1H), 7.14 (dd, J = 9.0, 2.2 Hz,
1H), 7.03 (d, J = 2.0 Hz, 1H), 4.00-3.30 (m, 8H), 3.00 (s, 3H),
2.40-2.30 (m, 2H); MS ESI [M + H].sup.+ 395.5, calcd for
[C.sub.20H.sub.22N.sub.6OS + H].sup.+ 395.2
[0221] The following compounds were prepared according to the
general method A3.
A60:
4-amino-S-(5-(4-methylpiperazine-1-carbonyl)-1H-benzo[d]imidazol-2-yl-
)thieno[2,3-b]-pyridin-6(7H)-one
##STR00104##
[0223] LDA (1.0 M in THF/hexanes, 2.3 mL, 2.3 mmol) was added
dropwise over 15 min at rt to a stirred suspension of ethyl
2-(6-(4-methylpiperazine-1-carbonyl)-1H-benzo[d]imidazol-2-yl)acetate
(0.150 g, 0.45 mmol) and 2-aminothiophene-3-carbonitrile (0.056 g,
0.45 mmol) in anh. THF (20 mL) under Ar. The addition was done
initially at rt and after 5 minutes at 35.degree. C. The heating
was continued at 35.degree. C. for 1 h before the reaction mixture
was cooled to rt, quenched with aq NH.sub.4Cl and concentrated
under reduced pressure. Purification by RP HPLC afforded
N-(3-cyanothiophen-2-yl)-2-(5-(4-methylpiperazine-1-carbonyl)-1H-benzo[d]-
imidazol-2-yl)acetamide*TFA as a light brown solid (82 mg, 35%).
.sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.96 (s, 1H), 7.89 (d,
J=8.5 Hz, 1H), 7.68 (dd, J=8.4, 1.4 Hz, 1H), 7.09-7.14 (m, 2H),
3.25-3.81 (m, 8H), 2.97 (s, 3H).
Step 2. The product of the previous reaction was filtered through
PoraPak (2 g, using MeOH then 2 M NH.sub.3 in MeOH) and dried. An
anh THF (12 mL) solution of the material (0.055 g, 0.13 mmol) under
Ar was treated with LiHMDS (1.0 M in THF, 0.7 mL, 0.7 mmol) over 3
min at rt, stirred for 10 min and heated at 45.degree. C. for 95
min. The reaction was then cooled to rt, quenched with aq
NH.sub.4Cl, concentrated under reduced pressure and purified by
prep HPLC. Filtration through PoraPak (2 g) and trituration with
CH.sub.2Cl.sub.2 afforded the title compound as a light yellow
solid 3.6 mg (3%). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta.
7.58-7.77 (m, 2H), 7.51 (d, J=5.80 Hz, 1H), 7.30 (dd, J=8.30, 1.30
Hz, 1H), 7.14 (d, J=5.80 Hz, 1H), 3.53-3.92 (m, 4H), 2.48-2.70 (m,
4H), 2.43 (s, 3H). MS ESI [M+H].sup.+ 409.2, calcd for
[C.sub.20H.sub.20N.sub.6O.sub.2S+H].sup.+ 409.2.
TABLE-US-00003 Yield; description; Example/IUPAC name Structure
salt A61: 4-amino-5-(6-methyl-5- (4-methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00105##
19 mg (7%); Ornage-tan solid; TFA Reagents (method A3): Step 1:
ethyl 2-(5-methyl-6-(4-methylpiperazin-1-yl)-1H-
benzo[d]imidazol-2-yl)acetate (0.17 g, 0.53 mmol),
2-aminothiophene-3-carbonitrile (72 mg, 0.53 mmol), LDA (1.0 M in
THF/hexanes, 1.7 mL, 1.7 mmol) in anh THF (12 mL). Step 2: LiHMDS
(1.0 M in THF, 1.7 mL, 1.7 mmol) in anh THF (20 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.48 (d, J = 5.77 Hz, 1 H), 7.44 (s,
1 H), 7.38 (s, 1 H), 7.12 (d, J = 5.77 Hz, 1 H), 3.58-3.59 (m, 2
H), 3.43-3.23 ( m., 4 H), 3.07-3.22 (m, 2 H), 3.01 (s, 3 H), 2.45
(s, 3 H). MS ESI [M + H].sup.+ 395.1, calcd for
[C.sub.20H.sub.22N.sub.6OS + H]+ 395.2. A62: 4-amino-5-(5-(morpho-
line-4-carbonyl)-1H-benzo- [d]imidazol-2-yl)thieno[2,3-
b]pyridin-6(7H)-one ##STR00106## 6.5 mg (2%); white solid; free
base Reagents (method A3): Step 1: ethyl
2-(6-(morpholine-4-carbonyl)-1H-benzo[d]imidazol-2- yl)acetate
(0.22 g, 0.70 mmol) and 2-aminothiophene-3-carbonitrile (84 mg,
0.70 mmol), LiHMDS (1.0 M in THF, 3.5 mL, 3.5 mmol) in anh. THF (24
mL). Step 2: LiHMDS (1.0 M in THF, 1.2 mL, 1.2 mmol) in anh THF (20
mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.63-7.76 (m, 2 H),
7.48-7.52 (m, 1 H), 7.31 (dd, J = 8.30, 1.50 Hz, 1 H), 7.14 (d, J =
5.77 Hz, 1 H), 3.58-3.88 (m, 8 H). MS ESI [M + H].sup.+ 396.2,
calcd for [C.sub.19H.sub.17N.sub.5O.sub.3S + H].sup.+ 396.1
A63:
7-(cyclopropylamino)-6-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazo-
l-2-yl)thieno[3,2-b]-pyridin-5(4H)-one 2,2,2-trifluroaceatate
##STR00107##
[0225] A suspension of
7-hydroxy-6-(6-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[3-
,2-b]pyridin-5(4H)-one (58 mg, 0.152 mmol) in anhydrous DCM (1 mL)
was added Tf.sub.2O (0.55 mL, 0.916 mmol) dropwise at rt. The
resulting reaction mixture was stirred at rt overnight before
addition of cyclopropanamine (100 mg, 1.83 mmol) at 0.degree. C.
dropwise. The resulting reaction mixture was stirred at 40.degree.
C. overnight and diluted with DCM followed by washing with satd
NaHCO.sub.3. The organic layer was dried over Na.sub.2SO.sub.4,
filtered, and concentrated to dryness. The residue was dissolved in
MeOH and run through PoraPak followed by removal of solvent under
reduced pressure. The crude product was purified by prep HPLC to
give the title compound as a yellow solid (5 mg, 6% yield). .sup.1H
NMR (400 MHz, CD.sub.3OD) .delta. 7.98 (d, J=5.5 Hz, 1H), 7.61 (d,
J=9.0 Hz, 1H), 7.26 (d, J=2.0 Hz, 1H), 7.21 (dd, J=8.4, 2.0 Hz,
1H), 7.10 (d, J=5.5 Hz, 1H), 3.92-3.84 (m, 2H), 3.71-3.62 (m, 2H),
3.41-3.36 (m, 2H), 3.20-3.10 (m, 2H), 3.09-3.03 (m, 1H), 3.01 (s,
3H), 1.04-0.96 (m, 2H), 0.93-0.89 (m, 2H); MS ESI [M+H].sup.+
421.2, calcd for [C.sub.22H.sub.24N.sub.6OS+H].sup.+ 421.2.
A64:
4-amino-5-(6-(piperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]p-
yridin-6(7H)-one tert-butyl
4-(3-amino-4-nitrophenyl)piperazine-1-carboxylate
##STR00108##
[0227] A mixture of 5-chloro-2-nitroaniline (2.5 g, 14.48 mmol),
tert-butyl piperazine-1-carboxylate (3.24 g, 17.38 mmol) and
K.sub.2CO.sub.3 (4.0 g, 28.96 mmol) in DMSO (100 mL) was stirred at
100.degree. C. for 3 days. H.sub.2O (150 mL) was then added with
stirring, suction filtered, rinsed with H.sub.2O and dried to give
the title compound as a brown solid (2.6 g, 57%). .sup.1H NMR (400
MHz, CDCl.sub.3) .delta. 8.04 (d, J=9.79 Hz, 1H), 6.27 (dd, J=9.66,
2.64 Hz, 1H), 6.21-6.11 (m, 2H), 5.95 (d, J=2.51 Hz, 1H), 3.61-3.54
(m, 4H), 3.40-3.34 (m, 4H), 1.50 (s, 9H); MS ESI [M+H].sup.+ 323.2,
calcd for [C.sub.15H.sub.22N.sub.4O.sub.4+H].sup.+ 323.2.
Tert-butyl 4-(3,4-diaminophenyl)piperazine-1-carboxylate
##STR00109##
[0229] To a suspension of tert-butyl
4-(3-amino-4-nitrophenyl)piperazine-1-(2.6 g, 8.04 mmol) in MeOH
(150 mL) was added 10% Pd/C (130 mg, 5% wt.). The resulting mixture
was hydrogenated under H.sub.2 balloon O/N. The resulting reaction
mixture was filtered, concentrated and dried to give the title
compound as a dark brown solid (2.29 g, 97%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 6.66 (d, J=8.28 Hz, 1H), 6.39 (d, J=2.51 Hz,
1H), 6.34 (dd, J=8.28, 2.51 Hz, 1H), 3.60-3.53 (m, 4H), 3.46-3.23
(m, 4H), 3.02-2.95 (m, 4H), 1.49 (s, 9H); MS ESI [M+H].sup.+ 293.1,
calcd for [C.sub.15H.sub.24N.sub.4O.sub.2+H].sup.+ 293.2.
Tert-butyl
4-(2-(2-ethoxy-2-oxoethyl)-1H-benzo[d]imidazol-6-yl)piperazine--
1-carboxylate
[0230] To a solution of tert-butyl
4-(3,4-diaminophenyl)piperazine-1-carboxylate (100 mg, 0.34 mmol)
in EtOH (3 mL) was added ethyl 3-ethoxy-3-iminopropionate
hydrochloride (190 mg, 0.68 mmol). The resulting mixture was heated
at 60.degree. C. for 3 h. After removal of solvents, it was diluted
with DCM (10 mL), adjust pH.apprxeq.8 with satd NaHCO.sub.3 and
separated. The aqueous was extracted with DCM (10 mL.times.2) and
the combined extracts were dried over NaSO.sub.4, then concentrated
and purified by flash chromatography (gradient: 100% EtOAc, then
MeOH/DCM 0-20%) to give the title compound as a dark orange solid
(116 mg, 87%). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.49-7.40
(m, 1H), 7.15-7.10 (m, 2H), 4.22 (q, J=7.11 Hz, 2H), 3.95 (s, 1H),
3.61 (br. s., 4H), 3.11 (br. s., 4H), 1.50 (s, 9H), 1.28 (t, J=7.15
Hz, 3H); MS ESI [M+H].sup.+ 389.2, calcd for
[C.sub.20H.sub.28N.sub.4O.sub.4+H].sup.+ 389.2.
Tert-butyl
4-(2-(4-amino-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-5-yl)-1H-be-
nzo[d]imidazol-6-yl)piperazine-1-carboxylate
##STR00110##
[0232] According to general method A, to a solution of
2-amino-4-ethoxythiophene-3-carbonitrile (64 mg, 0.52 mmol),
tert-butyl
4-(2-(2-ethoxy-2-oxoethyl)-1H-benzo[d]imidazol-6-yl)piperazine-1-carboxyl-
ate (200 mg, 0.52 mmol), LiHMDS (1 M in THF, 2.0 mL, 2.06 mmol).)
were used to generate the title compound as a light brown solid (88
mg, 35%). .sup.1H NMR (500 MHz, DMSO-d.sub.6) .delta. 12.72-12.61
(m, 1H), 12.13-12.02 (m, 1H), 10.72-10.55 (m, 1H), 8.01-7.93 (m,
1H), 7.57 (d, J=5.62 Hz, 1H), 7.52-7.43 (m, 1H), 7.24-7.10 (m, 2H),
6.93-6.87 (m, 1H), 3.52-3.44 (m, 4H), 3.07-3.00 (m, 4H), 1.45-1.40
(m, 9H); MS ESI [M+H].sup.+ 467.2, calcd for
[C.sub.23H.sub.26N.sub.6O.sub.3S+H].sup.+ 467.2.
4-amino-5-(6-(piperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridi-
n-6(7H)-one
##STR00111##
[0234] A mixture of tert-butyl
4-(2-(4-amino-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-5-yl)-1H-benzo[d]imid-
azol-6-yl)piperazine-1-carboxylate (83 mg, 0.178 mmol) in TFA (1
mL) was stirred at rt for 2 h before concentrated. The residue was
dissolved in MeOH (20 mL) and run through PoraPak then concentrated
to give the title compound as a yellow solid (45 mg, 69%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6) .delta. 12.65-12.58 (m, 1H),
10.77-10.61 (m, 1H), 8.03-7.94 (m, 1H), 7.59 (d, J=5.77 Hz, 1H),
7.54-7.42 (m, 1H), 7.19-7.10 (m, 2H), 6.92-6.86 (m, 1H), 3.09-3.01
(m, 4H), 2.94-2.88 (m, 4H); the signal due to NH.sub.2 cannot be
readily detected. MS ESI [M+H].sup.+ 367.2, calcd for
[C.sub.18H.sub.18N.sub.6OS+H].sup.+ 367.1.
A65:
4-amino-5-(6-(4-(oxetan-3-yl)piperazin-1-yl)-1H-benzo[d]imidazol-2-yl-
)thieno[2,3-b]pyridin-6(7H)-one
##STR00112##
[0236] A mixture of
4-amino-5-(6-(piperazin-1-yl)-H-benzo[d]imidazol-2-yl)thieno[23-b]pyridin-
-6(7H)-one (45 mg, 0.123 mmol), oxetan-3-one (8.8 mg, 0.123 mmol),
and NaBH(OAc).sub.3 (120 mg, 0.552 mmol) in DCE (2 mL) was stirred
at rt overnight then filtered. The filtrate was concentrated and
purified by prep. HPLC to give the title compound as TFA salt as a
yellow solid (50 mg, 76%). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.66 (d, J=9.03 Hz, 1H), 7.51 (d, J=5.77 Hz, 1H), 7.29 (t,
J=8.91 Hz, 2H), 7.18 (d, J=6.02 Hz, 1H), 4.98-4.87 (m, 4H),
4.54-4.45 (m, 1H), 3.63-3.40 (i, 8H); MS ESI [M+H].sup.+ 423.2,
calcd for [C.sub.21H.sub.2N.sub.6O.sub.2S+H].sup.+ 423.2.
TABLE-US-00004 Yield; description; Example/IUPAC name Structure
salt A66: 4-(((1S,2S)-2- hydroxycyclopentyl)amino)-
5-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00113## 35 mg (23%); brown
solid; 2 HCl Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.22 g, 0.29 mmol), (1S,2S)-2-
aminocyclopenta-1-ol (0.12 g, 1.2 mmol), DMF (5 mL). MS ESI [M -
CF.sub.3O.sub.2S + 2H].sup.+ 585.4, calcd for
[C.sub.32H.sub.36N.sub.6O.sub.3S + H].sup.+ 585.3. Step 2: Reagents
(general method D): a mixture of 2-(4-(((1S,2S)-2-
hydroxycyclopentyl)amino)-7-(4-methoxybenzyl)-6-oxo-6,7-dihydrothieno[2,3--
b]pyridin-5- yl)-5- and
6-(4-methylpiperazin-1-yl)-1H-benzotrifluoromethanesulfonate
(crude, 0.28 mmol), TFA (5 mL), and conc. HCl (1 mL). 1H NMR (400
MHz, CD.sub.3OD) .delta. = 7.75-7.60 (m, 2 H), 7.41 (br. s, 1 H),
7.30 (s, 1 H), 7.19 (d, J = 5.0 Hz, 1 H), 4.15-4.04 (m, 1 H),
4.02-3.89 (m, 2 H), 3.73-3.61 (m, 2 H), 3.28-3.14 (m, 5 H), 3.00
(br. s., 3 H), 1.99- 1.84 (m, 2 H), 1.76-1.46 (m, 3 H), 1.43-1.31
(m, 1 H); MS ESI [M + H].sup.+ 465.3, calcd for
[C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2. A67:
4-(((1R,2R)-2- hydroxycyclopentyl)amino)-
5-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00114## 45 mg (29%); brown
solid; 2 HCl Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.22 g, 0.29 mmol), (1R,2R)-2-
aminocyclopenta-1-ol (0.12 g, 1.2 mmol), DMF (5 mL). MS ESI [M -
CF.sub.3O.sub.2S + 2H].sup.+ 585.4, calcd for
[C.sub.32H.sub.36N.sub.6O.sub.3S + H].sup.+ 585.3. Step 2: Reagents
(general method D): a mixture of 2-(4-(((1R,2R)-2-
hydroxycyclopentyl)amino)-7-(4-methoxybenzyl)-6-oxo-6,7-dihydrothieno[2,3--
b]pyridin-5- yl)-5- and 6-(4-methylpiperazin-1-yl)-1H-benzo
trifluoromethanesulfonate (crude, 0.28 mmol), TFA (5 mL), and conc.
HCl (1 mL). 1H NMR (400 MHz, CD.sub.3OD) .delta. = 7.76-7.62 (m, 2
H), 7.45-7.36 (m, 1 H), 7.35-7.26 (m, 1 H), 7.22-7.15 (m, 1 H),
4.14- 4.05 (m, 1 H), 4.02-3.89 (m, 2 H), 3.74-3.62 (m, 2 H),
3.27-3.17 (m, 5 H), 3.00 (br. s., 3 H), 2.00-1.83 (m, 2 H),
1.74-1.47 (m, 3 H), 1.45-1.32 (m, 1 H); MS ESI [M + H].sup.+ 465.3,
calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2. A68:
7-amino-6-(5-fluoro- 6-(4-methylpiperazin-1-yl)-
1H-benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 5(4H)-one
##STR00115## 91 mg (32%); brown solid; 2 HCl Reagents (general
method A1): 3-amino-2-cyanothiophene (75 mg, 0.6 mmol), ethyl
2-(6-(4- methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(193 mg, 0.6 mmol), LDA (3 ml, 3 mmol), THF (8 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 8.01 (d, J = 5.5 Hz, 1H), 7.59 (d, J
= 11.0 Hz, 1H), 7.49 (d, J = 7.3 Hz, 1H), 7.11 (d, J = 5.3 Hz, 1H),
3.71-3.68 (m, 4H), 3.46-3.41 (m, 2H), 3.23-3.31 (m, 2H), 3.03 (s,
3H); MS ESI [M + H].sup.+ 399.2, calcd for
[C.sub.19H.sub.19FN.sub.6OS + H].sup.+ 399.4. A69:
4-amino-5-(7-fluoro- 6-(4-methylpiperazin-1-yl)-
1H-benzo[d]imidazol-2- yl)thieno[3,2-b]pyridin- 6(7H)-one
##STR00116## 49 mg (13%); brown solid; 2 HCl Reagents (general
method A1): 2-amino-3-cyanothiophene (100 mg, 0.8 mmol), ethyl
2-(6-(4- methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl)acetate
(258 mg, 0.8 mmol), LDA (4 ml, 4 mmol), THF (10 mL). .sup.1H NMR
(400 MHz, CD.sub.3OD) .delta. 7.57-7.51 (m, 2H), 7.39-7.35 (m, 1H),
7.20 (d, J = 5.8 Hz, 1H), 3.69-3.36 (m, 4H), 3.47-3.37 (m, 4H),
3.35 (s, 2H), 3.03 (s, 3H); MS ESI [M + H].sup.+ 399.2, calcd for
[C.sub.19H.sub.19FN.sub.6OS + H].sup.+ 399.4. A70: 4-(((1S,2R)-2-
hydroxycyclohexyl)amino)- 3-(6-(4-methylpiperazin-1-
yl)-1H-benzo[d]imidazol-2 yl)thieno[3,4-b]pyridin- 2(1H)-one
##STR00117## 40 mg (22%); yellow solid; 2 HCl Step 1: Reagents
(general method C): a mixture of 7-(4-methoxybenzyl)-5-(5 and/or
6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.26 g, 0.34 mmol), (1R,2S)-2-
aminocyclohexan-1-ol (0.1 g, 0.85 mmol), DMF (5 mL). MS ESI [M +
H].sup.+ 731.3, calcd for
[C.sub.34H.sub.37F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 731.2.
Step 2: Reagents (general method D): a mixture of
4-(((1S,2R)-2-hydroxycyclohexyl)amino)- 1-(4-methoxybenzyl)-3-(5-
and (6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno[3,4-b]pyridin-2(1H)-one (220 mg
crude), TFA (5 mL), and conc. HCl (2 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 8.43 (d, J = 3.3 Hz, 1H), 7.73 (d, J = 9.0 Hz,
1H), 7.44 (dd, J = 9.0, 2.2 Hz, 1H), 7.34 (d, J = 2.3 Hz, 1H), 7.01
(d, J = 3.0 Hz, 1H), 4.03-3.94 (m, 3H), 3.74-3.65 (m, 2H),
3.42-3.35 (m, 3H), 3.31-3.19 (m, 3H), 3.03 (s, 3H), 1.87-57 (m,
5H), 1.35-1.17 (m, 2H); MS ESI [M + H].sup.+ 479.4, calcd for
[C.sub.25H.sub.30N.sub.6O.sub.2S + H].sup.+ 479.2. A71:
(R)-3-(6-(4- methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-4-
((tetrahydrofuran-3- yl)amino)thieno[3,4- b]pyridin-2(1H)-one
##STR00118## 6 mg (3%); yellow solid; 2 HCl Step 1: Reagents
(general method C): a mixture of 7-(4-methoxybenzyl)-5-(5 and/or
6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.19 g, 0.26 mmol), (R)-3-
aminotetrahydrofuran (0.06 g, 0.64 mmol), DMF (5 mL). MS ESI [M +
H].sup.+ 703.3, calcd for
[C.sub.32H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 703.2.
Step 2: Reagents (general method D): a mixture of (R)-3-(5- and
(6-(4-methylpiperazin-1-yl)-
1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-4-((tetrahydrofura-
n-3- yl)amino)thieno[3,4-b]pyridin-2(1H)-one (crude, 165 mg), TFA
(4 mL), and conc. HCl (1.5 mL). .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 8.54 (d, J = 3.3 Hz, 1H), 7.74 (d, J = 9.5 Hz, 1H), 7.45
(dd, J = 9.0, 2.3 Hz, 1H), 7.34 (d, J = 2.01 Hz, 1H), 7.00 (d, J =
3.3 Hz, 1H), 4.02-3.91 (m, 3H), 3.85 (dd, J = 9.5, 3.3 Hz, 1H),
3.78-3.54 (m, 5H), 3.43-3.35 (m, 2H), 3.28-3.19 (m, 2H), 3.03 (s,
3H), 2.11-1.94 (m, 2H); MS ESI [M + H].sup.+ 451.3, calcd for
[C.sub.23H.sub.26N.sub.6O.sub.2S + H].sup.+ 451.2. A72:
4-(((1S,2R)-2- hydroxycyclohexyl)amino)- 5-(6-morpholino-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00119##
19 mg (7%); brown solid; 2 HCl Step 1: Reagents (general method C):
4-(4-methoxybenzyl)-6-(5- and 6-morpholino-1-
((trifluoro-methyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-5-oxo-4,5-dihydroth-
ieno[3,2- b]pyridin-7-yl trifluoro-methanesulfonate (crude, 0.51
mmol), (1R,2S)-2-aminocyclohexanol (0.23 g, 2.1 mmol), DMF (6 mL).
MS ESI [M - CF.sub.3O.sub.2S + 2H].sup.+ 586.5, calcd for
[C.sub.32H.sub.35N.sub.5O.sub.4S + H].sup.+ 586.2. Step 2: Reagents
(general method D): a mixture of 2-(4-(((1S,2R)-2-
hydroxycyclohexyl)amino)-7-(4-methoxybenzyl)-6-oxo-6,7-dihydrothieno[2,3-b-
]pyridin-5- yl)-5- and 6-morpholino-1H-benzo[d]imidazol-1-yl
trifluoromethanesulfonate (0.38, 0.51 mmol), TFA (5 mL), and conc.
HCl (1 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. = 7.77-7.69
(m, 1 H), 7.64-7.57 (m, 1 H), 7.54-7.45 (m, 2 H), 7.24-7.15 (m, 1
H), 4.09-3.96 (m, 5 H), 3.60- 3.44 (m, 5 H), 1.98-1.52 (m, 6 H),
1.48-1.22 (m, 2 H); MS ESI [M + H].sup.+ 466.4, calcd for
[C.sub.24H.sub.27N.sub.5O.sub.3S + H].sup.+ 466.2. A73:
5-(6-((2S,6R)-2,6- dimethylmorpholino)-1H-
benzo[d]imidazol-2-yl)-4- (((1S,2R)-2- hydroxycyclohexyl)amino)
thieno[2,3-b]pyridin-6(7H)- one ##STR00120## 8 mg (6%); brown
solid; TFA Step 1: Reagents (general method C): 5-(5 and
(6-((2S,6R)-2,6-dimethylmorpholino)-1-
(((trifluoromethyl)sulfonyl)oxy)-1H-benzo[d]imidazol-2-yl)-7-(4-methoxyben-
zyl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (crude, 0.23 mmol), (1R,2S)-2-
aminocyclohexanol (0.26 g, 0.23 mmol), DMF (1 mL). MS ESI [M +
H].sup.+ 746.5, calcd for
[C.sub.35H.sub.38F.sub.3N.sub.5O.sub.6S.sub.2 + H].sup.+ 746.2.
Step 2: Reagents (general method D): a mixture of
5-(6-((2S,6R)-2,6-dimethylmorpholino)-1-
((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-yl)-4-(((1S,2R)-2-
hydroxycyclohexyl)amino)-7-(4-methoxybenzyl)thieno[2,3-b]pyridin-6(7H)-one
and 5-
((2S,6R)-2,6-dimethylmorpholino)-2-(4-(((1S,2R)-2-hydroxycyclohexyl)amino)-
-7-(4-
methoxybenzyl)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-5-yl)-1H-benzo[d]imid-
azol-1-yl trifluoromethanesulfonate (0.094 g, 0.13 mmol), TFA (3
mL), and conc. HCl (1 mL). 1H NMR (400 MHz, CD.sub.3OD) .delta.
7.69 (d, J = 9.0 Hz, 1 H), 7.56-7.48 (m, 2 H), 7.40 (d, J = 10.0
Hz, 1 H), 7.20 (d, J = 6.0 Hz, 1 H), 4.10-4.04 (m, 1 H), 4.04-3.89
(m, 3 H), 3.67 (d, J = 11.3 Hz, 2 H), 2.98-2.86 (m, 2 H), 2.02-1.89
(m, 2 H), 1.86-1.68 (m, 3 H), 1.68-1.58 (m, 1 H), 1.52-1.40 (m, 1
H), 1.39-1.33 (m, 1 H), 1.31 (d, J = 6.3 Hz, 6 H); MS ESI [M +
H].sup.+ 494.5, calcd for [C.sub.26H.sub.31N.sub.5O.sub.3S +
H].sup.+ 494.2. A74: 4-((2- methoxyethyl)amino)-5-(6-
(4-methylpiperazin-1-yl)- 1H-benzo[d]imidazol-2-
yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00121## 55 mg (33%); dark
yellow solid; TFA Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (crude, 0.30 mmol), 2- methoxyethanamine
(0.10 mL, 1.2 mmol), DMF (4 mL). MS ESI [M + H].sup.+ 691.4, calcd
for [C.sub.31H.sub.33F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 691.2.
Step 2: Reagents (general method D): a mixture of
2-(7-(4-methoxybenzyl)-4-((2-
methoxyethyl)amino)-6-oxo-6,7-dihydrothieno[2,3-b]pyridin-5-yl)-5-
and 6-(4- methylpiperazin-1-yl)-1H-benzo[d]imidazol-1-yl
trifluoromethanesulfonate (crude, 0.30 mmol), TFA (5 mL), and conc.
HCl (0.5 mL). .sup.1H NMR (400 MHz, CD.sub.3OD) .delta. 7.68 (d, J
= 8.8 Hz, 1 H), 7.62-7.56 (m, 1 H), 7.34 (d, J = 9.0 Hz, 1 H), 7.29
(d, J = 1.8 Hz, 1 H), 7.20 (d, J = 6.0 Hz, 1 H), 3.99-3.86 (m, 2
H), 3.76-3.64 (m, 2 H), 3.58 (d, J = 5.3 Hz, 2 H), 3.42-3.34 (m, 7
H), 3.25-3.12 (m, 2 H), 3.02 (s, 3 H); MS ESI [M + H].sup.+ 439.5,
calcd for [C.sub.22H.sub.26N.sub.6O.sub.2S + H].sup.+ 439.2. A75:
4-(((1R,2R)-2- hydroxycyclopentyl)amino)-
3-(6-(4-methylpiperazin-1- yl)-1H-benzo[d]imidazol-2-
yl)thieno[3,4-b]pyridin- 2(1H)-one ##STR00122## 22 mg (16%); yellow
solid; 2 HCl Step 1: Reagents (general method C): a mixture of
7-(4-methoxybenzyl)-5-(5 and/or 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (0.19 g, 0.26 mmol), (1R,2R)-2-
aminocyclopentanol (0.06 g, 0.64 mmol), DMF (5 mL). MS ESI [M +
H].sup.+ 717.2, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D): a mixture of
4-(((1R,2R)-2-hydroxycyclopentyl)amino)- 1-(4-methoxybenzyl)-3-(5-
and
(6-(4-methylpiperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-
benzo[d]imidazol-2-yl)thieno[3,4-b]pyridin-2(1H)-one (crude, 165
mg), TFA (4 mL), and conc. HCl (1 mL). .sup.1H NMR (400 MHz,
CD.sub.3OD) .delta. 8.59 (d, J = 3.3 Hz, 1H), 7.72 (d, J = 9.0 Hz,
1H), 7.44 (dd, J = 9.0, 2.3 Hz, 1H), 7.33 (d, J = 2.0 Hz, 1H), 7.00
(d, J = 3.3 Hz, 1H), 4.19-4.12 (m, 1H), 4.00-3.97 (m, 2H),
3.74-3.65 (m, 2H), 3.43-3.35 (m, 2H), 3.30-3.21 (m, 2H), 3.19- 3.10
(m, 1H), 3.03 (s, 3H), 1.99-1.87 (m, 2H), 1.76-1.65 (m, 2H),
1.57-1.44 (m, 1H), 1.44-1.28 (m, 1H); MS ESI [M + H].sup.+ 465.4,
calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+ 465.2. A76:
4-amino-5-(6- ((2S,6R)-2,6- dimethylmorpholino)-1H-
benzo[d]imidazol-2- yl)thieno[2,3-b]pyridin- 6(7H)-one ##STR00123##
49 mg (31%); greenish-yellow solid; TFA Reagents (general method
A2): ethyl 2-(6-((2S,6R)-2,6-dimethylmorpholino)-1H-
benzo[d]imidazol-2-yl)acetate (0.10 g, 0.32 mmol),
2-aminothiophene-3-carbonitrile (0.32 g, 0.32 mmol), LDA (1.0 M in
THF/hex, 1.1 mL, 1.1 mmol), THF (1 mL), 45.degree. C., 1.5 h. 50 mg
of mixture of uncyclized and cyclized was obtained which was
recyclized with KOBu.sup.t (1.0 M in THF, 1.3 mL, 1.3 mmol) in THF
(10 mL), 45.degree. C., 2 h; .sup.1H NMR (400 MHz, CD.sub.3OD)
.delta. 7.69 (d, J = 9.5 Hz, 1 H), 7.52 (d, J = 6.0 Hz, 1 H),
7.49-7.45 (m, 1 H), 7.37 (d, J = 9.0 Hz, 1 H), 7.19 (d, J = 5.8 Hz,
1 H), 4.03-3.92 (m, 2 H), 3.71-3.61 (m, 2 H), 2.92-2.77 (m, 2 H),
1.31 (d, J = 6.3 Hz, 6 H); MS ESI [M + H].sup.+ 396.3, calcd for
[C.sub.20H.sub.21N.sub.5O.sub.2S + H].sup.+ 396.1. A77:
(R)-5-(6-(4- methylpiperazin-1-yl)-1H- benzo[d]imidazol-2-yl)-4-
((tetrahydro-2H-pyran-3- yl)amino)thieno[2,3- b]pyridin-6(7H)-one
##STR00124## 66 mg (38%); yellow solid; TFA Step 1: Reagents
(general method C): a mixture of 7-(4-methoxybenzyl)-5-(5 and 6-(4-
methyl-piperazin-1-yl)-1-((trifluoromethyl)sulfonyl)-1H-benzo[d]imidazol-2-
-yl)-6-oxo-6,7- dihydrothieno[2,3-b]pyridin-4-yl
trifluoromethanesulfonate (crude, 0.30 mmol), (R)-
tetrahydro-2H-pyran-3-amine (crude in DCM, 1.5 mmol), DMF (5 mL).
MS ESI [M + H].sup.+ 717.4, calcd for
[C.sub.33H.sub.35F.sub.3N.sub.6O.sub.5S.sub.2 + H].sup.+ 717.2.
Step 2: Reagents (general method D): a mixture of
(R)-2-(7-(4-methoxybenzyl)-6-oxo-4-
((tetrahydro-2H-pyran-3-yl)amino)-6,7-dihydrothieno[2,3-b]pyridin-5-yl)-5-
and 6-(4- methylpiperazin-1-yl)-1H-benzo[d]imidazol-1-yl
trifluoromethanesulfonate (crude, 0.30 mmol), TFA, and conc. HCl.
1H NMR (400 MHz, CD.sub.3OD) .delta. 7.67 (d, J = 8.8 Hz, 1 H),
7.56 (d, J = 5.8 Hz, 1 H), 7.37-7.26 (m, 2 H), 7.21 (d, J = 6.0 Hz,
1 H), 4.00-3.80 (m, 3 H), 3.75- 3.59 (m, 3 H), 3.53-3.41 (m, 2H),
3.40-3.34 (m, 2H), 3.29-3.11 (m, 3 H), 3.01 (s, 3 H), 2.14-1.96 (m,
1 H), 1.83-1.66 (m, 2 H), 1.42-1.25 (m, 1 H); MS ESI [M + H].sup.+
465.3, calcd for [C.sub.24H.sub.28N.sub.6O.sub.2S + H].sup.+
465.2.
Example B: HPK1 Inhibition Assay
[0237] Active HPK1 (MAP4K1) was purchased as an N-terminal GST
fusion of human HPK1 (aa 1-346) from Invitrogen (cat #PV6355). HPK1
activity was measured using an indirect ELISA detection system.
GST-HPK1 (0.6 nM) was incubated in the presence of 12 .mu.M ATP
(Sigma cat #A7699), 5 mM MOPS (pH 7.2), 2.5 mM
.beta.-glycerol-phosphate, 5 mM MgCl.sub.2, 0.4 mM EDTA, 1 mM EGTA,
0.05 mM DTT, in a 96 well microtitre plate pre-coated with 0.5
jig/well bovine myelin basic protein (MBP) (Millipore, cat
#13-110). The reaction was allowed to proceed for 30 min, followed
by 5 washes of the plate with Wash Buffer (phosphate buffered
saline supplemented with 0.2% Tween 20), and incubation for 30 min
with a 1:3000 dilution of anti-phospho-threonine rabbit polyclonal
antibody (Cell Signaling cat #9381). The plate was washed 5 times
with wash buffer, incubated for 30 min in the presence of goat
anti-rabbit horse radish peroxidase conjugate (BioRad cat #1721019,
1:3000 concentration), washed an additional 5 times with wash
buffer, and incubated in the presence of TMB substrate (Sigma cat
#T0440). The colorimetric reaction was allowed to continue for 5
min, followed by addition of stop solution (0.5 N H.sub.2SO.sub.4),
and quantified by detection at 450 nm with a monochromatic plate
reader (Molecular Devices M5).
[0238] Compound inhibition was determined at either a fixed
concentration (10 .mu.M) or at a variable inhibitor concentration
(typically 50 .mu.M to 0.1 .mu.M in a 10 point dose response
titration). Compounds were pre-incubated in the presence of enzyme
for 15 min prior to addition of ATP and the activity remaining
quantified using the above described activity assay. The %
inhibition of a compound was determined using the following
formula; % inhibition=100.times.(1-(experimental value-background
value)/(high activity control-background value)). The IC.sub.50
value was determined using a non-linear 4 point logistic curve fit
(XLfit4, IDBS) with the formula; (A+(B/(1+((x/C){circumflex over (
)}D)))), where A=background value, B=range, C=inflection point,
D=curve fit parameter.
Example C: FLT3 Inhibition Assay
[0239] FLT3 and LCK compound inhibition were determined using FRET
based Z'-LYTE Kinase Assay Kit with Tyrosine 2 peptide as the
substrate (Invitrogen cat #PV3191). The FLT3 kinase assay was
performed according to the manufacturer's suggested specifications
with an ATP concentration of 940 .mu.M and 1 nM FLT3 (Invitrogen
cat #PV3182) and 180 .mu.M ATP and 25 nM LCK (Invitrogen cat
#P3043) for the LCK kinase reaction. The % inhibition values were
determined according to the manufacturer's directions and IC.sub.50
values were obtained using a non-linear 4 point logistic curve fit
(XLfit4, IDBS).
[0240] In Table 1 below, IC.sub.50 value ranges for exemplary
compounds are given. The IC.sub.50 ranges are indicated as "A,"
"B," and "C," for values less than or equal to 0.05 .mu.M; those
greater than 0.05 .mu.M and less than or equal to 0.5 .mu.M; and
those greater than 0.5 .mu.M, respectively.
TABLE-US-00005 TABLE 1 Inhibition Data of HPK1, Lck and Flt3
IC.sub.50 Range Example HPK1 Lck Flt3 A1 A B A A2 A B A A3 C -- --
A4 A B A A5 C C -- A6 B -- -- A7 A B A A8 A B A A9 A A A A10 A B A
A11 A A12 A -- -- A13 A B A A14 A A A A15 A A A A16 A -- -- A17 A A
A A18 A B A A19 B A20 C A21 A C B A22 A B A A23 A A A A24 A A A A25
A -- -- A26 A -- -- A27 A A A A28 A -- -- A29 A B -- A30 A A A A31
A B A A32 C A -- A33 A B A A34 A A A A35 A A A A36 A A A A37 A A A
A38 A A A A39 A A A A40 A A A A41 A A A A42 A A A A43 A B A A44 A A
A A45 A A A A46 A B A A47 A -- -- A48 A A A A49 C -- -- A50 C C --
A51 A -- -- A52 A A A A53 A A A A54 A B A A55 A B A A56 A -- -- A57
A B A A58 A B A A59 -- -- -- A60 B B -- A61 A B A A62 B -- -- A63 A
-- -- A64 A B A A65 A B A A66 A A A A67 A B A A68 A A A A69 -- B A
A70 -- A A A71 A A A A72 A C A A73 B -- -- A74 A -- -- A75 A -- --
A76 A -- -- A77 A -- --
Example D: In Vitro Phosphorylation Assays
[0241] Jurkat E6.1 cells were obtained from American Type Culture
Collection (ATCC, Manassas, Va.), and maintained according to the
supplier's instructions. Cells were washed three times and starved
in RPMI 1640 medium supplemented with 0.5% fetal calf serum for 18
h at 37.degree. C. Serum starved cells were pretreated with the
indicated concentration of inhibitor for 4 hours before stimulation
with 10 .mu.g/ml ax-CD3 antibody (BioLegend, Inc., San Diego,
Calif.) for 10 min at 37.degree. C. The cells were washed once in
phosphate-buffered saline (pH 7.4) containing 10 mm sodium
pyrophosphate, 10 mm sodium fluoride, 10 mm EDTA, and 1 mm sodium
orthovanadate. Protein lysates were prepared using ice-cold
radioimmunoprecipitation assay (RIPA) lysis buffer. A total of 100
.mu.g of cell lysate was loaded onto Bis-Tris gels (Life
Technologies, Carlsbad, Calif.) with full-range molecular weight
marker as a size reference, and resolved by SDS-PAGE
electrophoresis. Proteins were transferred to PVDF membrane
(Millipore, Billerica, Mass.), blocked and probed with antibodies
for phospho-SLP-76 (Ser376) (rabbit polyclonal #13177; Cell
Signaling Technology Inc., Danvers, Mass.), SLP-76 (rabbit
polyclonal #4958; Cell Signaling Technology Inc., Danvers, Mass.),
phospho-ERK (mouse monoclonal sc-7383; Santa Cruz Biotechnology
Inc., Santa Cruz, Calif.) and ERK1/2 (rabbit polyclonal 06-182;
Millipore, Billerica, Mass.). Secondary antibodies were diluted 1
in 15,000 and incubated for 1 h at rt. Protein bands were
visualized and quantified using Odyssey near infrared imager
(LI-COR, Lincoln, Nebr.). Table 2 below lists effects of
representative compounds of the present invention against SLP-76
serine 376 phosphorylation and ERK1/2 T202/Y204 phosphorylation in
.alpha.-CD3 stimulated Jurkat E6.1 cells.
TABLE-US-00006 TABLE 2 Effects of HPK1 inhibitors against SLP-76
serine 376 phosphorylation and ERK1/2 T202/Y204 phosphorylation in
.alpha.-CD3 stimulated Jurkat E6.1 cells. SLP76 S376 ERK1/2
T202/Y204 Phosphorylation Phosphorylation Compound
Onset-Substantive* Onset Example Inhibition (.mu.M) Inhibition
(.mu.M) A1 0.3-1.0 >3.0 A30 0.3-1.0 >3.0 A43 0.1-0.3 >3.0
A18 0.3-1.0 >3.0 A10 1.0-3.0 >3.0 A57 0.3-1.0 >3.0 A23
0.3-1.0 >3.0 A58 0.1-0.3 >3.0 A34 1.0-3.0 3.0 A21 >3.0
>3.0 A37 0.1-0.3 1.0-3.0 *>75% inhibition as estimated by
immunoblot analysis
Example E: Syngeneic CT26 Cell Line Xenograft Model
[0242] The CT26 WT cell line, which is an
N-nitroso-N-methylurethane-(NNMU) induced, mouse-derived,
undifferentiated colon carcinoma cell line, was obtained from
American Type Culture Collection (ATCC CRL-2638, Manassas, Va., DC,
USA). Cells were grown in Roswell Park memorial Institute medium
commonly referred to as RPMI 1640 Medium containing 4.5 g/L
glucose, 0.11 g/L sodium pyruvate, 1.5 g/L sodium bicarbonate,
L-glutamine& 2.385 g/L HEPES plus 10% fetal bovine serum. Six
to eight week old female BALB/c mice were purchased from Jackson
Laboratories and received and acclimated at the MaRS-TMDT Animal
Resources Centre for 1 week prior to the start of the experiment.
The mice were fed ad libitum autoclaved water and Rodent Lab Diet
(Harlan Teklad LM-485) consisting of 19% crude protein, 5% crude
fat, and 5% crude fiber. Mice were housed in microisolator cages
and maintained in an environment with a 12 h light cycle at
20-22.degree. C. and 40-60% humidity. On the day of implantation,
CT26 cells were harvested and re-suspended with serum free RPMI1640
to a concentration of 1.times.10.sup.7/mL and each mouse was
injected subcutaneously with a volume of 0.1 mL containing
1.times.10.sup.6 CT26 cells in the right rear flank. After 6 d,
palpable tumors with an average volume of .about.65 mm.sup.3
(calculated using the formula: tumor
volume=width.sup.2.times.length/2) had formed. At this time,
animals were separated into five groups of eight animals per group
such that each group contained animals bearing tumors of similar
average size and treatment was initiated. For dosing, Example A1
was dissolved in water to a concentration of 7.5 mg/mL or 15 mg/mL
for dosing of the 75 mg/kg and the 150 mg/kg doses, respectively.
As a positive control and to investigate the combinatorial activity
of Example A1, a rat IgG2b anti-PD1 antibody (BioXcell (NH, USA))
was dosed used. The five groups were treated with: i) 10 mL/kg
water QD for 21 d administered by oral gavage (PO) plus 150 .mu.g
rat IgG2b isotype control antibody dosed by intraperitoneal (IP)
injection on day 0, 3, 6 and 10 (the control arm); ii) 150 .mu.g
anti-PD-1 antibody dosed by intraperitoneal (IP) injection on day
0, 3, 6 and 10; iii) 75 mg/kg Example A1 QD for 21 days
administered PO; iv) 150 mg/kg Example A1 QD for 21 days
administered PO v) 150 mg/kg Example A1 QD for 21 days administered
PO plus 150 .mu.g anti-PD-1 antibody dosed by intraperitoneal (IP)
injection on day 0, 3, 6 and 10. Toxicity was evaluated by body
weight measurements and clinical observations. Tumour measurements
and body weights were taken three times per week. Percent tumor
growth inhibition (TGI) was calculated by the formula:
%
TGI=100.times.[1-(TV.sub.f,treated-TV.sub.i,treated)/(TV.sub.f,control-
-TV.sub.i,control)]
Tumour growth inhibition at day 21, is shown in FIG. 2. A
dose-dependent effect was observed in response to treatment with
Example A1, with 75 mg/kg and 150 mg/kg QD inhibiting tumour growth
by 44% and 64%, respectively. Whilst the anti-PD-1 antibody alone
resulted in an average TGI of 34%, when combined with 150 mg/kg QD
Example A1, the TGI increased to 86%. According to University
Health Network (UHN) Animal Use Protocols (AUPs), mice in efficacy
experiments should be sacrificed when the tumour size is above 1500
mm.sup.3 or if the bodyweight of the animal decreases or if the
animals are displaying clinical signs that require termination for
humane reasons. In this study, the compound was well tolerated with
all animals gaining weight over the course of the study and no
animals were terminated due to clinical signs. A tumour size of
<1500 mm.sup.3 at day 21 was used as a cutoff to represent
survival. Using this cutoff, at day 21 no animals survived in the
control arm, 1 of 8 animals (12.5%) survived in the anti-PD-1 arm,
2 of 8 animals (25%) survived in the 75 mg/kg/day Example A1 arm, 3
of 8 animals (37.5%) survived in the 150 mg/kg/day Example A1 arm,
and 7 of 8 animals (87.5%) survived in the 150 mg/kg/day Example A1
and anti-PD-1 arm. These results demonstrate that compounds of the
invention, as exemplified by compound A1, have in vivo antitumor
activity and can be efficaciously combined with other
immunomodulatory approaches.
Example F: EAE Disease Progression Model
[0243] C57/BL6 mice were obtained from Jackson Laboratories. The
Institutional Animal Care and Use Committee of the University
Health Network approved all animal procedures. Mice were
subcutaneously (SC) immunized with MOG35-55 peptide emulsified in
Complete Freund's Adjuvant (CFA) supplemented with Mycobacterium
tuberculosis. On days 0 and 2 after immunization, the mice were
intraperitoneal (IP) injected with pertussis toxin. Clinical signs
of EAE were monitored daily, according to the following criteria:
0, no disease; 1, decreased tail tone; 2, hind limb weakness or
partial paralysis; 3, complete hind limb paralysis; 4, front and
hind limb paralysis; 5, death, or sacrifice due to moribund state.
For treatment with compound during EAE induction, mice were dosed
orally (PO) with 50 mg/kg A30 (n=4) or water (vehicle control; n=5)
every day (QD). Data are the mean score.+-.SEM. The test results is
shown in FIG. 3.
* * * * *