U.S. patent application number 11/029547 was filed with the patent office on 2005-07-21 for pyrrolopyridazine compounds and methods of use thereof for the treatment of proliferative disorders.
Invention is credited to Barbosa, Stephanie A., Chen, Zhong, Hunt, John T., Salvati, Mark.
Application Number | 20050159420 11/029547 |
Document ID | / |
Family ID | 28678223 |
Filed Date | 2005-07-21 |
United States Patent
Application |
20050159420 |
Kind Code |
A1 |
Salvati, Mark ; et
al. |
July 21, 2005 |
Pyrrolopyridazine compounds and methods of use thereof for the
treatment of proliferative disorders
Abstract
Disclosed are pyrrolopyridazine compounds, methods of preparing
such compounds, and their use for the treatment of proliferative,
inflammatory, and other disorders.
Inventors: |
Salvati, Mark;
(Lawrenceville, NJ) ; Barbosa, Stephanie A.;
(Lambertville, NJ) ; Chen, Zhong; (Cranbury,
NJ) ; Hunt, John T.; (Princeton, NJ) |
Correspondence
Address: |
STEPHEN B. DAVIS
BRISTOL-MYERS SQUIBB COMPANY
PATENT DEPARTMENT
P O BOX 4000
PRINCETON
NJ
08543-4000
US
|
Family ID: |
28678223 |
Appl. No.: |
11/029547 |
Filed: |
January 5, 2005 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11029547 |
Jan 5, 2005 |
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10396197 |
Mar 25, 2003 |
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6900208 |
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60368249 |
Mar 28, 2002 |
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60402118 |
Aug 8, 2002 |
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Current U.S.
Class: |
514/248 ;
544/235 |
Current CPC
Class: |
A61P 5/28 20180101; A61K
31/5025 20130101; A61K 45/06 20130101; A61K 2300/00 20130101; A61K
31/5025 20130101; A61P 5/00 20180101; A61P 29/00 20180101; Y02A
50/411 20180101; C07D 487/04 20130101; A61P 5/26 20180101; A61P
35/00 20180101 |
Class at
Publication: |
514/248 ;
544/235 |
International
Class: |
A61K 031/503; C07D
487/04 |
Claims
1-10. (canceled)
11. A pharmaceutical composition comprising a compound of formula
(I) 401including enantiomers, diastereomers, pharmaceutically
acceptable salts, prodrugs, and solvates thereof, wherein: R.sub.1
is selected from the group consisting of H, hydroxyl, alkyl,
aralkyl, halogen, OR.sub.1', OC(O)R.sub.1', OC(O)OR.sub.1',
OC(O)NR.sub.1'R.sub.1", OS(O).sub.2R.sub.1'", and
OS(O).sub.2NF.sub.1'R.sub.1"; wherein R.sub.1' and R.sub.1" are
each independently selected from the group consisting of H, alkyl,
aryl, aralkyl, heterocyclo, and cycloalkyl groups; R.sub.1' and
R.sub.1" may also be taken together to form one of a cycloalkyl, an
aryl, and a heterocyclic group; R.sub.1'" is selected from the
group consisting of H, alkyl, aryl, aralkyl, heterocyclo, and
cycloalkyl; R.sub.2 is selected from the group consisting of alkyl,
cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O--,
R.sub.1'C(O)-- R.sub.1"R.sub.1'NC(O)-- R.sub.1'"O(O).sub.2S--
R.sub.1'R.sub.1"N(O).sub.2- S-- and R.sub.1'"(O).sub.nS-- wherein n
is the integer 1 or 2; R.sub.1 and R.sub.2 may be taken together
with the carbon atoms to which they are attached to form
cycloalkene; X is selected from the group consisting of a valence
bond, O, S, and NR.sub.2'; and R.sub.2' is selected from the group
consisting of H, alkyl, aralkyl, C(O)R.sub.1, C(O)OR.sub.1,
SO.sub.2NR.sub.1'R.sub.1", C(O)NR.sub.1'R.sub.1" and SO.sub.2
R.sub.1'"; with the proviso that when X is S, R.sub.2 is selected
from the group consisting of H, alkyl, cycloalkyl, aryl,
heterocycle and aralkyl; R.sub.3 is selected from the group
consisting of H, hydroxyl, alkyl, cycloalkyl, heterocycle, aryl,
aralkyl, acyl, carbalkoxy, carboxamido, halogen, amine, substituted
amine, OR.sub.3', CH.sub.2OR.sub.3', CH.sub.2NR.sub.3'R.sub.3",
CH.sub.2SR.sub.3', OC(O)R.sub.3', OC(O)OR.sub.3",
OC(O)NR.sub.3'R.sub.3", OS(O).sub.2R.sub.3', and
OS(O).sub.2NR.sub.3'R.sub.3"; wherein R.sub.3' and R.sub.3" are
each independently selected from the group consisting of H, alkyl,
aralkyl, heterocycle, cycloalkyl, and aryl; R.sub.3' and R.sub.3"
may also be taken together with the nitrogen atom to which they are
attached to form a heterocyclyl; when R.sub.3 is a carbalkoxy,
acyl, or carboxamido group, these groups are optionally substituted
with one or two substituent groups, said substituent groups are
independently selected from the group consisting of H, alkyl,
aralkyl, heterocycle, cycloalkyl, and aryl; R.sub.2 and R.sub.3 may
also be taken together to form a cycloalkyl, aryl, or heterocyclic
group; R.sub.4 is selected from the group consisting of alkyl,
cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O),
R.sub.1'C(O), R.sub.1"R.sub.1'NC(O), R.sub.1.sup.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS, wherein n is
the integer 1 or 2; Y is selected from the group consisting of a
valence bond, O, S, and NR.sub.4'; R.sub.4' is selected from the
group consisting of H, alkyl, aralkyl, a heterocycle, C(O)R.sub.1,
C(O)OR.sub.1, S(O.sub.2)NR.sub.1'R.sub.1", C(O)NR.sub.1'R.sub.1",
and S(O.sub.2)R.sub.1; with the proviso that when Y is S, R.sub.4
is selected from the group consisting of alkyl, cycloalkyl, aryl,
heterocycle and aralkyl; when Y is NR.sub.4', R.sub.4' can be taken
together with R.sub.3 with the N atom and carbon atoms to which
they are attached to form a heterocyclic ring system; R.sub.5 is
selected from the group consisting of H, halogen, cyano, alkyl,
cycloalkyl, a heterocycle, aryl, aralkyl, acyl, substituted
alkylene group, R.sub.1'OC(O), R.sub.1'C(O), R.sub.1"R.sub.1'NC(O),
R.sub.1'"O(O).sub.2S, R.sub.1'R.sub.1"N(O).sub.2S and
R.sub.1'"(O).sub.nS; wherein n the integer 1 or 2; Z is selected
from the group consisting of a valence bond, O, S, and NR.sub.5';
R.sub.5' is selected from the group consisting of H, alkyl, aralkyl
and a heterocycle; with the proviso that when Z is a valence bond,
R.sub.5 is selected from the group consisting of H, halogen, a
substituted alkylene group and a cyano group; and, with the further
proviso that when Z is S, R.sub.5 is selected from the group
consisting of H, alkyl, cycloalkyl, aryl, heterocycle and aralkyl;
and, R.sub.6 is selected from the group consisting of H, alkyl,
cycloalkyl, aryl, aralkyl, a heterocycle, acyl, carbalkoxy, and
carboxamido; said carbalkoxy, acyl, and carboxamido groups are
optionally substituted with one or two substituent groups, each of
which is independently selected from the group consisting of H,
alkyl, aralkyl, and a heterocycle, further comprising one or more
additional therapeutic agent.
12. The pharmaceutical composition of claim 11, wherein the
additional therapeutic agent is selected from the group consisting
of angiogenesis inhibitors, antiestrogens, progestogens, aromatase
inhibitors, antihormones, antiprogestogens, antiandrogens, LHRH
agonists and antagonists, testosterone 5.alpha.-dihydroreductase
inhibitors, farnesyl transferase inhibitors, anti-invasion agents,
growth factor inhibitors, antimetabolites, intercalating antitumour
antibiotics, platinum derivatives, alkylating agents, antimitotic
agents, topoisomerase inhibitors, cell cycle inhibitors, and
biological response modifiers.
13. The pharmaceutical composition of claim 11, wherein the
additional therapeutic agent is selected from the group consisting
of linomide, integrin .alpha.v.beta.3 function inhibitors,
angiostatin, razoxin, tamoxifen, toremifen, raloxifene,
droloxifene, iodoxyfene, megestrol acetate, anastrozole, letrazole,
borazole, exemestane, flutamide, nilutamide, bicalutamide,
cyproterone acetate, gosereline acetate, luprolide, finasteride,
metalloproteinase inhibitors, urokinase plasminogen activator
receptor function inhibitors, growth factor antibodies, growth
factor receptor antibodies, tyrosine kinase inhibitors,
serine/threonine kinase inhibitors, methotrexate, 5-fluorouracil,
purine, adenosine analogues, cytosine arabinoside, doxorubicin,
daunomycin, epirubicin, idarubicin, mitomycin-C, dactinomycin,
mithramycin, cisplatin, carboplatin, nitrogen mustard, melphalan,
chlorambucil, busulphan, cyclophosphamide, ifosfamide nitrosoureas,
thiotephan, vincristine, taxol, taxotere, epothilone analogs,
discodermolide analogs, eleutherobin analogs, etoposide,
teniposide, amsacrine, topotecan, flavopyridols.
14. The pharmaceutical composition of claim 11, wherein the
additional therapeutic agent is selected from the group consisting
of taxol, Erbitux.TM., paraplatin and Ifex.
15. A method of treating a proliferative or inflammatory disease,
in a patient in need thereof, comprising administering a
therapeutically effective amount of the compound of claim 1.
16. The method of claim 15, further comprising administering at
least one additional therapeutic agent.
17. The method of claim 16, wherein the additional therapeutic
agent is selected from the group consisting of angiogenesis
inhibitors, antiestrogens, progestogens, aromatase inhibitors,
antihormones, antiprogestogens, antiandrogens, LHRH agonists and
antagonists, testosterone 5.alpha.-dihydroreductase inhibitors,
farnesyl transferase inhibitors, anti-invasion agents, growth
factor inhibitors, antimetabolites, intercalating antitumour
antibiotics, platinum derivatives, alkylating agents, antimitotic
agents, topoisomerase inhibitors, cell cycle inhibitors, and
biological response modifiers.
18. The method of claim 16, wherein the additional therapeutic
agent is selected from the group consisting of linomide, integrin
.alpha.v.beta.3 function inhibitors, angiostatin, razoxin,
tamoxifen, toremifen, raloxifene, droloxifene, iodoxyfene,
megestrol acetate, anastrozole, letrazole, borazole, exemestane,
flutamide, nilutamide, bicalutamide, cyproterone acetate,
gosereline acetate, luprolide, finasteride, metalloproteinase
inhibitors, urokinase plasminogen activator receptor function
inhibitors, growth factor antibodies, growth factor receptor
antibodies, tyrosine kinase inhibitors, serine/threonine kinase
inhibitors, methotrexate, 5-fluorouracil, purine, adenosine
analogues, cytosine arabinoside, doxorubicin, daunomycin,
epirubicin, idarubicin, mitomycin-C, dactinomycin, mithramycin,
cisplatin, carboplatin, nitrogen mustard, melphalan, chlorambucil,
busulphan, cyclophosphamide, ifosfamide nitrosoureas, thiotephan,
vincristine, taxol, taxotere, epothilone analogs, discodermolide
analogs, eleutherobin analogs, etoposide, teniposide, amsacrine,
topotecan, flavopyridols.
19. The method of claim 16, wherein the additional therapeutic
agent is selected from the group consisting of Erbitux.TM., taxol,
paraplatin and Ifex.
20. The method of claim 16, wherein the at least one additional
therapeutic agent is administered simultaneously, sequentially or a
combination thereof, with the compound of formula I.
Description
RELATED APPLICATIONS
[0001] This application is a divisional of Ser. No. 10/396,197
filed on Mar. 25, 2003 which claims priority benefit under Title 35
.sctn. 119(e) of U.S. provisional Application Nos. 60/368,249,
filed Mar. 28, 2002, and 60/402,118, filed Aug. 8, 2002, the
contents of which are herein incorporated by reference.
FIELD OF THE INVENTION
[0002] The present invention relates to pyrrolopyridazine
compounds, methods of preparing such compounds, and their use for
the treatment of proliferative and other disorders.
BACKGROUND OF THE INVENTION
[0003] Protein kinases are a class of enzymes that catalyze the
transfer of a phosphate group from ATP to a tyrosine, serine,
threonine, or histidine residue located on a protein substrate.
Protein kinases clearly play a role in normal cell growth. Many of
the growth factor receptor proteins have intracellular domains that
function as protein kinases and it is through this function that
they effect signaling. The interaction of growth factors with their
receptors is a necessary event in the normal regulation of cell
growth, and the phosphorylation state of substrate proteins often
is related to the modulation of cell growth.
[0004] The human epidermal growth factor receptor (HER) family
consists of four distinct receptor tyrosine kinases referred to as
HER1, HER2, HER3, and HER4. These kinases are also referred to as
erbB1, erbB2, etc. HER1 is also commonly referred to as the
epidermal growth factor receptor (EGFr). With the exception of
HER3, these receptors have intrinsic protein kinase activity that
is specific for tyrosine residues of phosphoacceptor proteins. The
HER kinases are expressed in most epithelial cells as well as tumor
cells of epithelial origin. They are also often expressed in tumor
cells of mesenchymal origin such as sarcomas or rhabdomyosarcomas.
Receptor tyrosine kinases (RTKs) such as HER1 and HER2 are involved
in cell proliferation and are associated with diseases such as
psoriasis and cancer. Disruption of signal transduction by
inhibition of these kinases in target cells is known to have an
antiproliferative and therapeutic effect.
[0005] The enzymatic activity of receptor tyrosine kinases can be
stimulated by either overexpression, or by ligand-mediated
dimerization. The formation of homodimers as well as heterodimers
has been demonstrated for the HER receptor family. An example of
homodimerization is the dimerization of HER1 (EGF receptor) by one
of the EGF family of ligands (which includes EGF, transforming
growth factor alpha, betacellulin, heparin-binding EGF, and
epiregulin). Heterodimerization among the four HER receptor kinases
can be promoted by binding to members of the heregulin (also
referred to neuregulin) family of ligands. Such heterodimerization,
involving HER2 and HER3, or a HER3/HER4 combination, results in a
significant stimulation of the tyrosine kinase activity of the
receptor dimers even though one of the receptors (HER3) is
enzymatically inert. The kinase activity of HER2 has also been
shown to be activated by virtue of overexpression of the receptor
alone in a variety of cell types. Activation of receptor homodimers
and heterodimers results in phosphorylation of tyrosine residues on
the receptors and on other intracellular proteins. This is followed
by the activation of intracellular signaling pathways such as those
involving the microtubule associated protein kinase (MAP kinase )
and the phosphatidylinositol 3-kinase (PI3 kinase). Activation of
these pathways has been shown to lead to cellular proliferation and
the inhibition of apoptosis. Inhibition of HER kinase signaling has
been shown to inhibit cell proliferation and survival.
[0006] Deregulation of EGF receptors plays a role in the aberrant
growth of epithelial cysts in the disease described as polycystic
kidney disease [Du, J., Wilson, P. D., Amer. J. Physiol., 269 (2 Pt
1), 487 (1995); Nauta, J., et al., Pediatric Research, 37(6), 755
(1995); Gattone, V. H. et al., Developmental Biology, 169(2), 504
(1995); Wilson, P. D. et al., Eur. J. Cell Biol., 61(1), 131,
(1993)]. The compounds of this invention, which inhibit the
catalytic function of the EGF receptors, are consequently useful
for the treatment of this disease.
[0007] The mitogen-activated protein kinase (MAPK) pathway is a
major pathway in the cellular signal transduction cascade from
growth factors to the cell nucleus. The pathway involves kinases at
two levels: MAP kinase kinases (MAPKK), and their substrates MAP
(mitogen activated protein) kinases (MAPK). There are different
isoforms in the MAP kinase family. [For review, see Seger, R.;
Krebs, E. G. FASEB, 9, 726, (1995)]. The compounds of this
invention can inhibit the action of one or both of these kinases:
MEK, a MAP kinase kinase, and its substrate ERK, a MAP kinase.
ERK(extracellular regulated kinases), a p42 MAPK, is found to be
essential for cell proliferation and differentiation. Over
expression and/or over activation of MEK or ERK has been found to
be associated with various human cancers [For example, Sivaraman,
V. S.et al., C. C. J. Clin. Invest., 99, 1478 (1997)]. It has been
demonstrated that inhibition of MEK prevents activation of ERK and
subsequent activation of ERK substrates in cells, resulting in
inhibition of cell growth stimulation and reversal of the phenotype
of ras-transformed cells [Dudley, D. T. et al., Proc. Nat. Acad.
Sci., 92, 7686 (1995)].
[0008] Members of the raf family of kinases phosphorylate serine
residues on MEK. There are three serine/threonine kinase members of
the raf family known as a-raf, b-raf, and c-raf. While mutations in
the raf genes are rare in human cancers, c-raf is activated by the
ras oncogene which is mutated in a wide number of human tumors.
Therefore, inhibition of the kinase activity of c-raf may provide a
way to prevent ras mediated tumor growth [Campbell, S. L.,
Oncogene, 17, 1395 (1998)].
[0009] The Src family of cytoplasmic protein tyrosine kinases
consists of at least eight members (Src, Fyn, Lyn, Yes, Lck, Fgr,
Hck and Blk) that participate in a variety of signaling pathways
[Schwartzberg, P. L., Oncogene, 17, 1463 (1998)]. The prototypical
member of this tyrosine kinase family is p60.sup.src (Src). Src is
involved in proliferation and migration responses in many cell
types. In limited studies, Src activity has been shown to be
elevated in breast, colon (-90%), pancreatic (>90%) and liver
(>90%) tumors. Greatly increased Src activity is also associated
with metastasis (>90%) and poor prognosis. Antisense Src message
impedes growth of colon tumor cells in nude mice [Staley et al.,
Cell Growth & Differentation, 8, 269 (1997)], suggesting that
Src inhibitors should slow tumor growth. In addition to its role in
cell proliferation, Src also acts in stress response pathways,
including the hypoxia response. Previous studies have shown that
colonic tumor cells genetically engineered to express antisense Src
message form tumors demonstrating reduced vascularization in nude
mouse models [Ellis, et al., J. Biol. Chem., 273, 1052 (1998)],
suggesting that Src inhibitors would be anti-angiogenic as well as
anti-proliferative.
[0010] Apart from its role in cancer, Src also appears to play a
role in osteoporosis. Mice genetically engineered to be deficient
in src production were found to exhibit osteopetrosis, the failure
to resorb bone [Soriano, P., Cell, 64, 693 (1991); Boyce, B. F., J.
Clin. Invest., 90, 1622 (1992)]. This defect was characterized by a
lack of osteoclast activity. Since osteoclasts normally express
high levels of Src, inhibition of Src kinase activity may be useful
in the treatment of osteoporosis [Missbach, M., Bone, 24, 437
(1999)].
[0011] In addition to EGFr, there are several other RTKs including
FGFr, the receptor for fibroblast growth factor (FGF); flk-1, also
known as KDR, and flt-1, the receptors for vascular endothelial
growth factor (VEGF); and PDGFr, the receptor for platelet derived
growth factor (PDGF). The formation of new blood vessels, a process
known as angiogenesis, is essential for tumor growth. Two natural
angiogenesis inhibitors, angiostatin and endostatin, dramatically
inhibited the growth of a variety of solid tumors. [O'Reilly, M.
S., Cell, 79, 315 (1994); O'Reilly, M. S., Nature Medicine, 2, 689
(1996); O'Reilly, M. S., Cell, 88, 277 (1997)]. Since FGF and VEGF
are known to stimulate angiogenesis, inhibition of the kinase
activity of their receptors should block the angiogenic effects of
these growth factors. In addition, the receptor tyrosine kinases
tie-1 and tie-2 also play a key role in angiogenesis [Sato, T. N.,
Nature, 376, 70 (1995)]. Compounds that inhibit the kinase activity
of FGFr, flk-1, flt-1, tie-1 or tie-2 may inhibit tumor growth by
their effect on angiogenesis.
[0012] PDGF is a potent growth factor and chemoattractant for
smooth muscle cells (SMCs), and the renarrowing of coronary
arteries following angioplasty is due in part to the enhanced
proliferation of SMCs in response to increased levels of PDGF.
Therefore, compounds that inhibit the kinase activity of PDGFr may
be useful in the treatment of restenosis. In addition, since PDGF
and PDGFr are overexpressed in several types of human gliomas,
small molecules capable of suppressing PDGFr activity have
potential utility as anticancer therapeutics [Nister, M., J. Biol.
Chem., 266, 16755 (1991); Strawn, L. M., J. Biol. Chem. 269, 21215
(1994)].
[0013] In addition, a large number of cytokines participate in the
inflammatory response, including IL-1, IL-6, IL-8 and TNF-.alpha..
Overproduction of cytokines such as IL-1 and TNF-.alpha. are
implicated in a wide variety of diseases, including inflammatory
bowel disease, rheumatoid arthritis, psoriasis, multiple sclerosis,
endotoxin shock, osteoporosis, Alzheimer's disease, and congestive
heart failure, among others [Henry et al., Drugs Fut., 24:1345-1354
(1999); Salituro et al., Curr. Med. Chem., 6:807-823 (1999)].
Evidence in human patients indicates that protein antagonists of
cytokines are effective in treating chronic inflammatory diseases,
such as monoclonal antibody to TNF-.alpha. (Enbrel) [Rankin et al.,
Br. J. Rheumatol., 34:334-342 (1995)], and soluble TNF-.alpha.
receptor-Fc fusion protein (Etanercept) [Moreland et al., Ann.
Intern. Med., 130:478-486 (1999)].
[0014] The biosynthesis of TNF-.alpha. occurs in many cell types in
response to an external stimulus, such as a mitogen, an infectious
organism, or trauma. Important mediators of TNF-.alpha. production
are the mitogen-activated protein (MAP) kinases, and in particular,
p38 kinase. These kinases are activated in response to various
stress stimuli, including but not limited to proinflammatory
cytokines, endotoxin, ultraviolet light, and osmotic shock.
Activation of p38 requires dual phosphorylation by upstream MAP
kinase kinases (MKK3 and MKK6) on threonine and tyrosine within a
Thr-Gly-Tyr motif characteristic of p38 isozymes.
[0015] There are four known isoforms of p38, i.e., p38-.alpha.,
p38.beta., p38.gamma., and p38.delta.. The .alpha. and .beta.
isoforms are expressed in inflammatory cells and are key mediators
of TNF-.alpha. production. Inhibiting the p38.alpha. and .beta.
enzymes in cells results in reduced levels of TNF-.alpha.
expression. Also, administering p38.alpha. and .beta. inhibitors in
animal models of inflammatory disease has proven that such
inhibitors are effective in treating those diseases. Accordingly,
the p38 enzymes serve an important role in inflammatory processes
mediated by IL-1 and TNF-.alpha.. Compounds that reportedly inhibit
p38 kinase and cytokines such as IL-1 and TNF-.alpha. for use in
treating inflammatory diseases are disclosed in the following
published international patent applications: WO 00/12497
(quinazoline derivatives as p38 kinase inhibitors); WO 00/56738
(pyridine and pyrimidine derivatives for the same purpose); WO
00/12497 (discusses the relationship between p38 kinase
inhibitors); and WO 00/12074 (piperazine and piperidine compounds
useful as p38 inhibitors).
[0016] In summary, the tight regulation of signal transduction
normally exerted by the array of kinase enzymes is often lost in
malignant cells. Compounds which modulate these kinases are thus
highly desirable for the treatment of disorders associated with
aberrant cellular proliferation. Moreover, compounds which modulate
the cytokines associated with the inflammatory response are highly
desirable for the treatment of inflammatory disorders.
SUMMARY OF THE INVENTION
[0017] Advantageously, the present invention provides compositions
and methods for the treatment of proliferative disorders, including
cancer, and inflammatory diseases. The methods comprise
administering a therapeutically effective amount of a kinase
inhibitor of formula I, below, or a salt, solvate, prodrug or
stereoisomer thereof, and, optionally, at least one additional
therapeutic agent. The treatment is preferably administered to a
mammalian species, more preferably to a human, in need thereof.
[0018] More specifically, the instant invention provides a compound
of formula I: 1
[0019] including enantiomers, diastereomers, pharmaceutically
acceptable salts, prodrugs, and solvates thereof, wherein:
[0020] R.sub.1 is selected from the group consisting of H,
hydroxyl, alkyl, aralkyl, halogen, OR.sub.1', OC(O)R.sub.1',
OC(O)OR.sub.1', OC(O)NR.sub.1'R.sub.1", OS(O).sub.2R.sub.1'", and
OS(O).sub.2NR.sub.1'R.s- ub.1"; wherein R.sub.1' and R.sub.1" are
each independently selected from the group consisting of H, alkyl,
aryl, aralkyl, heterocyclo, and cycloalkyl groups; R.sub.1' and
R.sub.1" may also be taken together to form one of a cycloalkyl, an
aryl, and a heterocyclic group; R.sub.1'" is selected from the
group consisting of H, alkyl, aryl, aralkyl, heterocyclo, and
cycloalkyl;
[0021] R.sub.2 is selected from the group consisting of H, alkyl,
cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O),
R.sub.1'C(O), R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS; wherein n is
the integer 1 or 2;
[0022] R.sub.1 and R.sub.2 may be taken together to form a
cycloalkyl, aryl, or heterocyclic group;
[0023] X is selected from the group consisting of a valence bond,
O, S, and NR.sub.2'; and R.sub.2' is selected from the group
consisting of H, alkyl, aralkyl, C(O)R.sub.1, C(O)OR.sub.1,
SO.sub.2NR.sub.1'R.sub.1", C(O)NR.sub.1'R.sub.1" and
SO.sub.2R.sub.1'"; with the proviso that when X is S, R.sub.2 is
selected from the group consisting of H, alkyl, cycloalkyl, aryl,
heterocycle and aralkyl;
[0024] R.sub.3 is selected from the group consisting of H,
hydroxyl, alkyl, cycloalkyl, heterocycle, aryl, aralkyl, acyl,
carbalkoxy, carboxamido, halogen, amine, substituted amine,
OR.sub.3', CH.sub.2OR.sub.3', CH.sub.2NR.sub.3'R.sub.3",
CH.sub.2SR.sub.3', OC(O)R.sub.3', OC(O)OR.sub.3",
OC(O)NR.sub.3'R.sub.3", OS(O).sub.2R.sub.3', and
OS(O).sub.2NR.sub.3'R.sub.3"; wherein R.sub.3' and R.sub.3" are
each independently selected from the group consisting of H, alkyl,
aralkyl, heterocycle, cycloalkyl, and aryl; R.sub.3' and R.sub.3"
may also be taken together to form a cycloalkyl, aryl, or
heterocyclic group; when R.sub.3 is a carbalkoxy, acyl, or
carboxarnido group, these groups are optionally substituted with
one or two substituent groups, said substituent groups are
independently selected from the group consisting of H, alkyl,
aralkyl, heterocycle, cycloalkyl, and aryl; said substituent groups
may also be taken together to form a cycloalkyl, aryl, or
heterocyclic group;
[0025] R.sub.2 and R.sub.3 may also be taken together to form a
cycloalkyl, aryl, or heterocyclic group;
[0026] R.sub.4 is selected from the group consisting of H, alkyl,
cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O),
R.sub.1"C(O), R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS, wherein n is
the integer 1 or 2;
[0027] Y is selected from the group consisting of a valence bond,
O, S, and NR.sub.4'; R.sub.4' is selected from the group consisting
of H, alkyl, aralkyl, a heterocycle, C(O)R.sub.1, C(O)OR.sub.1,
S(O.sub.2)NR.sub.1'R.sub.1", C(O)NR.sub.1'R.sub.1", and
S(O.sub.2)R.sub.1; with the proviso that when Y is S, R.sub.4 is
selected from the group consisting of alkyl, cycloalkyl, aryl,
heterocycle and aralkyl; when Y is NR.sub.4', R.sub.4' can be taken
together with R.sub.3 to form a heterocyclic ring system;
[0028] R.sub.5 is selected from the group consisting of H, halogen,
cyano, alkyl, cycloalkyl, a heterocycle, aryl, aralkyl, acyl,
substituted alkylene group, R.sub.1'OC(O), R.sub.1"C(O),
R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS; wherein n is 1
or 2;
[0029] Z is selected from the group consisting of a valence bond,
O, S, and NR.sub.5'; R.sub.5' is selected from the group consisting
of H, alkyl, aralkyl and a heterocycle; with the proviso that when
Z is a valence bond, R.sub.5 is selected from the group consisting
of H, halogen, a substituted alkylene group and a cyano group; and,
with the further proviso that when Z is S, R.sub.5 is selected from
the group consisting of H, alkyl, cycloalkyl, aryl, heterocycle and
aralkyl; and,
[0030] R.sub.6 is selected from the group consisting of H, alkyl,
cycloalkyl, aryl, aralkyl, a heterocycle, acyl, carbalkoxy, and
carboxamido; said carbalkoxy, acyl, and carboxamido groups are
optionally substituted with one or two substituent groups, each of
which is independently selected from the group consisting of H,
alkyl, aralkyl, and a heterocycle.
[0031] Further provided are pharmaceutical compositions comprising
a compound of formula I, above, or a salt, solvate, or stereoisomer
thereof, and at least one pharmaceutically acceptable carrier.
Optionally, the pharmaceutical composition may also comprise at
least one additional therapeutic agent.
[0032] Also provided are methods of treating proliferative or
inflammatory diseases, in patients in need thereof, by
administering a compound of formula I, above, or a salt, solvate,
or stereoisomer thereof, and, optionally, at least one additional
therapeutic agent.
DETAILED DESCRIPTION OF THE INVENTION
[0033] Definitions
[0034] The following definitions apply to the terms as used
throughout this specification, unless otherwise limited in specific
instances.
[0035] Unless otherwise indicated, the term "lower alkyl", "alkyl"
or "alk" as employed herein alone or in combined form, e.g.,
aralkyl or haloalkyl, includes both straight and branched chain
hydrocarbons, preferably containing 1 to 12 carbons in the case of
alkyl or alk, in the normal chain, and preferably 1 to 4 carbons in
the case of lower alkyl. Examples of alkyl groups include methyl,
ethyl, propyl, isopropyl, butyl, t-butyl, or isobutyl, pentyl,
hexyl, isohexyl, heptyl, 4,4-dimethylpentyl, octyl,
2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, and the
like. Each alkyl group may be optionally substituted with 1 to 4
substituents which may include alkyl, alkenyl, alkynyl, aryl,
cycloalkyl, halogen, heterocyclo, hydroxy, cyano, nitro, amino,
monoalkylamino, dialkylamino, hydroxylamine, sulfonate, sulfamide,
cyano-guanidine, oxo, carbalkoxy, carboxamido, SO.sub.n where n is
0, 1, or 2, and/or acyl groups.
[0036] Unless otherwise indicated, the term "cycloalkyl" as
employed herein alone or in combined form includes saturated cyclic
hydrocarbon groups or partially unsaturated (containing 1 or 2
double bonds) cyclic hydrocarbon groups, containing at least one
ring and a total of 3 to 7 carbons, preferably 3 to 6 carbons,
forming the ring. Examples of cycloalkyl groups include
cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl,
cyclopentenyl, cyclohexenyl, and the like. Cycloalkyl groups may
optionally be substituted in the same manner as described above for
alkyl groups.
[0037] The term "aryl" as employed herein alone or in combined
form, e.g., aryloxy, refers to monocyclic and bicyclic aromatic
groups containing 6 to 10 carbons in the ring portion, such as
phenyl, indenyl, indanyl, or naphthyl including 1-naphthyl and
2-naphthyl and the like. Aryl groups may be optionally substituted
through available carbon atoms with 1, 2, or 3 groups selected from
hydrogen, halo, alkyl, cycloalkyl, aralkyl, heterocyclo, haloalkyl,
alkoxy, aryloxy, haloalkoxy, alkenyl, trifluoromethyl,
trifluoromethoxy, alkynyl, hydroxy, amino, nitro, cyano,
carbalkoxy, acyl, hydroxylamine, sulfonate, sulfamide,
cyano-guanidine, SO.sub.n where n is 0, 1, or 2, carboxamido
groups, or monosubstituted amino, or disubstituted amino, wherein
the amino substituents are independently alkyl, aralkyl, aryl,
acyl, or carbalkoxy groups.
[0038] The term "aralkyl" as used herein refers to an aryl group,
as defined above, bonded directly through an alkyl moiety, such as
a benzyl group, for example. An aralkyl group may be optionally
substituted with any group described herein as an aryl or alkyl
substitutent.
[0039] Unless otherwise indicated, the term "lower alkenyl" or
"alkenyl" as used herein by itself or in combined form refers to
straight or branched chain radicals of 2 to 12 carbons, preferably
2 to 5 carbons, in the normal chain, which include one to six
double bonds in the normal chain, such as vinyl, 2-propenyl,
3-butenyl, 2-butenyl, 4-pentenyl, 3-pentenyl, 2-hexenyl, 3-hexenyl,
2-heptenyl, 3-heptenyl, 4-heptenyl, 3-octenyl, 3-nonenyl,
4-decenyl, 3-undecenyl, 4-dodecenyl, and the like, which may be
optionally substituted in the same manner as that described for
alkyl groups.
[0040] Unless otherwise indicated, the term "lower alkynyl" or
"alkynyl" as used herein by itself or in combined form refers to
straight or branched chain radicals of 2 to 12 carbons, preferably
2 to 8 carbons, in the normal chain; which include one triple bond
in the normal chain, such as 2-propynyl, 3-butynyl, 2-butynyl,
4-pentynyl, 3-pentynyl, 2-hexynyl, 3-hexynyl, 2-heptynyl,
3-heptynyl, 4-heptynyl, 3-octynyl, 3-nonynyl, 4-decynyl,
3-undecynyl, 4-dodecynyl and the like, which may optionally be
substituted in the same manner as that described for alkyl
groups.
[0041] The normal carbon chain of any alkyl, alkenyl, alkynyl, or
aralkyl group may optionally be interrupted by one or more
heteroatoms.
[0042] As used herein, the term "acyl" refers to a group of the
formula C(O)R, wherein R represents a hydrogen atom, an alkyl
group, an aryl group, a heterocycle, or an aralkyl group.
[0043] As used herein, the term "carbalkoxy" refers to a group of
the formula C(O)OR, wherein R represents a hydrogen atom, an alkyl
group, an aryl group, a heterocycle, or an aralkyl group.
[0044] As used herein, the term "carboxamido" refers to a group of
the formula C(O)NR.sub.2, wherein the R groups, which may be the
same or different, represent a hydrogen atom, an alkyl group, an
aryl group, a heterocycle, or an aralkyl group. Alternatively, the
two R groups, when taken together with the nitrogen atom, may form
a heterocyclo group.
[0045] The terms "heterocyclo", "heterocyclic" and "heterocycle" as
used herein refer to an optionally substituted, aromatic or
non-aromatic cyclic group, which, for example, is a 4 to 7 membered
monocyclic, 7 to 11 membered bicyclic, or 10 to 15 membered
tricyclic ring system, which has at least one heteroatom in at
least one carbon atom-containing ring. Each ring of the
heterocyclic group containing a heteroatom may have 1, 2, 3, or 4
heteroatoms selected from nitrogen atoms, oxygen atoms and sulfur
atoms, where the nitrogen and sulfur heteroatoms may also
optionally be oxidized and the nitrogen heteroatoms may also
optionally be quaternized. The heterocyclic group may be attached
at any heteroatom or carbon atom.
[0046] Examples of suitable monocyclic heterocyclic groups include
pyrrolidinyl, pyrrolyl, pyrazolyl, oxetanyl, pyrazolinyl,
imidazolyl, imidazolinyl, imidazolidinyl, oxazolyl, oxazolidinyl,
isoxazolinyl, isoxazolyl, thiazolyl, thiadiazolyl, thiazolidinyl,
isothiazolyl, isothiazolidinyl, furyl, tetrahydrofuryl, thienyl,
oxadiazolyl, piperidinyl, piperazinyl, 2-oxopiperazinyl,
2-oxopiperidinyl, 2-oxopyrrolidinyl, 2-oxazepinyl, azepinyl,
4-piperidonyl, pyridyl, N-oxo-pyridyl, pyrazinyl, pyrimidinyl,
pyridazinyl, tetrahydrothiopyranyl, tetrahydropyranyl, morpholinyl,
thiamorpholinyl, thiamorpholinyl sulfoxide,
tetrahydrothiopyranylsulfone, thiamorpholinyl sulfone,
1,3-dioxolane and tetrahydro-1,1-dioxothienyl, dioxanyl,
isothiazolidinyl, thietanyl, thiiranyl, triazinyl, triazolyl, and
the like.
[0047] Examples of suitable bicyclic hetrocyclic groups include
indolyl, benzothiazolyl, benzoxazolyl, benzothienyl, quinuclidinyl,
quinolinyl, quinolinyl-N-oxide, tetrahydroisoquinolinyl,
isoquinolinyl, benzimidazolyl, benzopyranyl, indolizinyl,
benzofuryl, chromonyl, coumarinyl, cinnolinyl, quinoxalinyl,
indazolyl, pyrrolopyridyl, furopyridinyl (such as
furo[2,3-c]pyridinyl, furo[3,1-b]pyridinyl] or
furo[2,3-b]pyridinyl), dihydroisoindolyl, dihydroquinazolinyl (such
as 3,4-dihydro-4-oxo-quinazolinyl), benzisothiazolyl,
benzisoxazolyl, benzodiazinyl, benzofurazanyl, benzothiopyranyl,
benzotriazolyl, benzpyrazolyl, dihydrobenzofuryl,
dihydrobenzothienyl, dihydrobenzothiopyranyl,
dihydrobenzothiopyranyl sulfone, dihydrobenzopyranyl, indolinyl,
isochromanyl, isoindolinyl, naphthyridinyl, phthalazinyl,
piperonyl, purinyl, pyridopyridyl, quinazolinyl,
tetrahydroquinolinyl, thienofuryl, thienopyridyl, thienothienyl,
benzoxodiazol, benzothiodiazol, and the like.
[0048] In preferred embodiments, at least one of the heteroatoms in
the heterocycle is a nitrogen atom.
[0049] Examples of suitable substituents for heterocyclic groups
include one or more alkyl groups as described above or one or more
groups described above as alkyl or aryl substituents. Also suitable
are aryl groups and smaller heterocycles, such as epoxides and
aziridines.
[0050] The term "heteroatom" as used herein includes oxygen, sulfur
and nitrogen, where the nitrogen and sulfur heteroatoms may also
optionally be oxidized and the nitrogen heteroatoms may also
optionally be quaternized.
[0051] The term "halogen" or "halo" as used herein alone or as part
of another group refers to fluorine, chlorine, bromine, and
iodine.
[0052] As used herein, the expression "optionally substituted," as
in "optionally substituted lower alkyl", "optionally substituted
aryl" or the like, refers to alkyl, aryl, and other groups which
may be unsubstituted or substituted with the substituents mentioned
above. Further, when a moiety is described herein as optionally
substituted with more than one substituent, it is intended that
each of the multiple substituents be chosen independently from
among the substituents mentioned above.
[0053] As used herein, the term "about" means that amounts, sizes,
formulations, parameters, and other quantities and characteristics
are not and need not be exact, but may be approximate and/or larger
or smaller, as desired, reflecting tolerances, conversion factors,
rounding off, measurement error and the like, and other factors
known to those of skill in the art. In general, an amount, size,
formulation, parameter or other quantity or characteristic is
"about" or "approximate" whether or not expressly stated to be
such.
[0054] Compounds of the Invention
[0055] In accordance with the present invention, compounds having
formula I, below, are provided. 2
[0056] In compounds of formula I, R.sub.1 is selected from H,
hydroxyl, alkyl, aralkyl, halogen, OR.sub.1', OC(O)R.sub.1',
OC(O)OR.sub.1', OC(O)NR.sub.1'R.sub.1", OS(O).sub.2R.sub.1'", and
OS(O).sub.2NR.sub.1'R.s- ub.1". The groups R.sub.1' and R.sub.1"
may each independently be H, alkyl, aryl, aralkyl, heterocyclo, or
cycloalkyl groups, or may be taken together to form a cycloalkyl,
aryl, or heterocyclic group, any of which may be optionally
substituted. The group R.sub.1'" is H, alkyl, aryl, aralkyl,
heterocyclo, or cycloalkyl group.
[0057] The R.sub.2 is selected from the group consisting of H,
alkyl, cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O),
R.sub.1'C(O), R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS where n is an
integer of 1 or 2.
[0058] R.sub.1 and R.sub.2 when taken together may form a
cycloalkyl, aryl, or heterocyclic group, any of which may be
optionally substituted.
[0059] The group X represents a valence bond, O, S, and NR.sub.2',
and R.sub.2' is H, alkyl, or aralkyl, C(O)R.sub.1, C(O)OR.sub.1,
SO.sub.2NR.sub.1'R.sub.1", C(O)NR.sub.1R.sub.1", SO.sub.2R.sub.1'".
With the limitation that when X is S, then R.sub.2 can only be
selected from H, alkyl, cycloalkyl, aryl, heterocycle and
aralkyl.
[0060] R.sub.3 is selected from the group consisting of H,
hydroxyl, alkyl, cycloalkyl, a heterocycle, aryl, aralkyl, acyl,
carbalkoxy, carboxamido, halogen, amine, substituted amine,
OR.sub.3', CH.sub.2OR.sub.3', CH.sub.2NR.sub.3'R.sub.3",
CH.sub.2SR.sub.3', OC(O)R.sub.3', OC(O)OR.sub.3",
OC(O)NR.sub.3'R.sub.3", OS(O).sub.2R.sub.3', and
OS(O).sub.2NR.sub.3'R.sub.3". The R.sub.3' and R.sub.3" groups are
each independently H, alkyl, aralkyl, heterocycle, cycloalkyl, or
aryl. R.sub.3' and R.sub.3" when taken together may form a
cycloalkyl, aryl, or heterocyclic group, any of which may be
optionally substituted.
[0061] When R.sub.3 is a carbalkoxy, acyl, or carboxainido group,
these groups are optionally substituted with one or two substituent
groups, each of which is independently H, alkyl, aralkyl,
heterocycle, cycloalkyl, or aryl. The substituent groups, when
taken together, may form a cycloalkyl, aryl, or heterocyclic group,
any of which may be optionally substituted.
[0062] The R.sub.2 and R.sub.3 groups may also be taken together to
form a cycloalkyl, aryl, or heterocyclic group, any of which may be
optionally substituted.
[0063] The R.sub.4 group is selected from the group consisting of
H, alkyl, cycloalkyl, aryl, heterocycle, aralkyl, R.sub.1'OC(O),
R.sub.1'C(O), R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS where n is an
integer of 1 or 2.
[0064] Y is selected from the group consisting of a valence bond,
O, S, and NR.sub.4', R.sub.4' is selected from H, alkyl, aralkyl, a
heterocycle, C(O)R.sub.1, C(O)OR.sub.1,
S(O.sub.2)NR.sub.1'R.sub.1", C(O)NR.sub.1'R.sub.1", and
S(O.sub.2)R.sub.1. With the limitation that when Y is S, then
R.sub.4 can only be selected from alkyl, cycloalkyl, aryl,
heterocycle and aralkyl.
[0065] In addition, when Y is a primary or secondard amine it can
be taken together with R.sub.3 to form a heterocyclic ring system
which may be optionally substituted.
[0066] R.sub.5 is selected from the group consisting of H, halogen,
cyano, alkyl, cycloalkyl, a heterocycle, aryl, aralkyl, acyl,
substituted alkylene group, R.sub.1'OC(O), R.sub.1'C(O),
R.sub.1"R.sub.1'NC(O), R.sub.1'"O(O).sub.2S,
R.sub.1'R.sub.1"N(O).sub.2S and R.sub.1'"(O).sub.nS where n is an
integer of 1 or 2.
[0067] Z is selected from the group consisting of a valence bond,
O, S, and NR.sub.5', with the conditions that when Z is a valence
bond, then R.sub.5 can only be selected from H, halogen, a
substituted alkylene group or a cyano group and when Z is S, then
R.sub.5 can only be selected from H, alkyl, cycloalkyl, aryl,
heterocycle and aralkyl. The group R.sub.5' is H, alkyl, aralkyl,
or a heterocycle.
[0068] Finally, R.sub.6 is selected from the group consisting of H,
alkyl, cycloalkyl, aryl, aralkyl, a heterocycle, acyl, carbalkoxy,
and carboxamido. The carbalkoxy, acyl, and carboxamido groups are
optionally substituted with one or two substituent groups, each of
which is independently H, alkyl, aralkyl, or a heterocycle.
[0069] Also included in the invention are the salts, solvates, and
stereoisomers enantiomers, and diastereomers of the compounds of
formula I.
[0070] All stereoisomers of the compounds of formula I are
contemplated, either in admixture or in pure or substantially pure
form. A compound of formula I may have asymmetric centers at any of
its non-aromatic carbon or nitrogen atoms, including those carbon
atoms in any of its substituents. Consequently, compounds of
formula I can exist in enantiomeric or diastereomeric forms or in
mixtures thereof. The processes for preparation can utilize
racemates, enantiomers or diastereomers as starting materials. When
diastereomeric or enantiomeric products are prepared, they may be
separated by conventional methods, for example, chromatographic or
fractional crystallization.
[0071] Preferred compounds of the invention are compounds of
formula I wherein Z is a valence bond and R.sub.5 is cyano.
[0072] In some preferred embodiments, R.sub.3 is an alkyl, aryl or
heteroaryl, and is preferably methyl.
[0073] In some preferred embodiments, Y is N. In still further
embodiments, X is a valence bond, O, or NR.sub.2'. R.sub.2' is
preferably R.sub.1'C(O) or --C(O)NR.sub.1'R.sub.2' or
--C(O)OR.sub.1'.
[0074] According to some embodiments of the present invention,
R.sub.1' and R.sub.1" are independently alkyl, cycloalkyl, or
heterocycloalkyl.
[0075] Other preferred compounds are compounds of formula I wherein
R.sub.4 is phenoxyaniline. Also preferred are compounds of formula
I wherein Y is NR.sub.4' wherein R.sub.4' is as defined above, or
wherein Y is O, and R.sub.4 is an aryl or heteroaryl group.
[0076] Other preferred compounds of the invention are compounds of
formula I wherein Z is a valence bond, R.sub.5 is cyano, and
R.sub.3 is methyl. Preferably, these compounds have one or more of
the following substituents: Y is N; R.sub.1' and R.sub.1" are
independently alkyl, cycloalkyl, or heterocycloalkyl; R.sub.4 is
alkyl, aryl or heteroaryl; X is a valenc bond, O, or NR.sub.2'; and
R.sub.2 is R.sub.1'C(O), --(C(O)NR.sub.1'R.sub.2', or
--C(O)OR.sub.1'.
[0077] Additional preferred compounds of formula I include those in
which Z is a valence bond, R.sub.5 is cyano, Y is NR.sub.4' wherein
R.sub.4' is as defined above, and R.sub.4 is a phenyl group or
heteroaryl group with one or more substitutions.
[0078] Further preferred compounds are illustrated in the examples
below.
[0079] Methods of Making the Compounds
[0080] Generally, compounds of formula I may be made by reacting a
pyrrole of formula II: 3
[0081] wherein X, R.sub.1, R.sub.2, and R.sub.3 are as previously
defined, with an aminating agent in the presence of a base to
produce the aminated pyrrole of formula III: 4
[0082] wherein X, R.sub.1, R.sub.2, and R.sub.3 are as previously
defined.
[0083] The compound of formula III is reacted with a carbonyl of
formula R.sub.6C(O)CH.sub.2ZR.sub.5 or an acetal of the formula
(RO).sub.2CR.sub.6CH.sub.2ZR.sub.5, wherein R is an alkyl group and
Z, R.sub.5, and R.sub.6 are as previously defined, under
ring-closure conditions to produce a compound of formula IV. 5
[0084] 4-Oxo-pyrrolopyridazines of formula IV may be reacted with a
reagent providing a leaving group, such as POCl.sub.3 or
POCl.sub.5, to yield a compound of formula V 6
[0085] wherein L is a leaving group.
[0086] The compound of formula V may be reacted with a compound of
the formula HYR.sub.4, wherein Y and R.sub.4 are as previously
defined, to produce a compound of formula I, above.
[0087] The compounds of formula I may be prepared by the processes
described in the following reaction schemes. Examples of suitable
reagents and procedures for conducting these reactions appear
hereinafter and in the working examples. Protection and
deprotection in the schemes herein may be carried out by procedures
generally known in the art. (See, for example, T. W. Greene &
P. G. M. Wuts, Protecting Groups in Organic Synthesis, 3rd Edition,
Wiley, (1999)). In schemes A though D, unless otherwise noted, X,
Y, Z, R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, and R.sub.6 are
as defined above. The variables L and L' represent leaving groups.
Variables designated with the subscript "a" or "b" have the same
scope as, but are chosen independently of, their parent variable.
For example, R.sub.2a, R.sub.2a', and R.sub.2a" are coextensive
with, but not necessarily identical to, R.sub.2, R.sub.2', and
R.sub.2", respectively. 7
[0088] Pyrroles of formula II may be obtained by the processes
described in Patent Cooperation Treaty (PCT) publication numberWO
00/71129, pending U.S. patent application Ser. No. 09/573829,
pending PCT Application Number US01/49982 and pending U.S. patent
application Ser. No. 10/036293 (all of which are herein
incorporated by reference in their entirety).
[0089] Treatment of a pyrrole of formula II with a base in a
suitable reaction medium followed by the addition of an amninating
reagent generates an aminopyrrole of formula III. Suitable bases
include sodium hydride (NaH), n-BuLi, t-BuLi, NaOH, lithium
diisopropylamide (LDA), and lithium hexamethyldisilazide (LiHMDS).
Suitable reaction media include tetrahydrofuran (THF),
CH.sub.2Cl.sub.2, dimethylformarnide (DMF), CH.sub.3CN and toluene.
Suitable aminating reagents include 2,4-dinitroaminophenol,
NH.sub.2OSO.sub.3H and ClNH.sub.2. Preferably the aminating reagent
is ClNH.sub.2.or 2,4-dinitroaminophenol. Preferably, the base is
NaH or LDA, the reaction medium is DMF or THF. More preferably, the
base is NaH, the reaction medium is DMF, and the aminating reagent
is 2,4-dinitroaminophenol.
[0090] Condensation of the compound of formula III with an acetal
followed by base induced cyclization in a suitable reaction medium
provides a pyrrolopyridazine of formula IV. Suitable bases include
NaOH, LDA, diisopropylethylamine (DIPEA),
1,8-diazoicyclo[5.4.0]undec-7-ene (DBU), and K.sub.2CO.sub.3.
Suitable reaction media include THF, CH.sub.2Cl.sub.2, DMF and
toluene. Preferably, the base is DBU, DIPEA or LDA and the reaction
medium is toluene or DMF. More preferably, the base is DBU or
DIPEA, and the reaction medium is toluene. Alternatively, compounds
of formula IV may be obtained by the reactions of Schemes H, J and
K, below.
[0091] Conversion of the oxo group at position 4 of the compound of
formula IV to a leaving group L, as in compounds of formula V, can
then be accomplished using a suitable halogenating reagent, such as
SOCl.sub.2, POCl.sub.3 or POCl.sub.5. More preferably, the reagent
is POCl.sub.3.
[0092] Treatment of a compound of formula V with a reagent of
formula HY-R.sub.4 in the presence of a base in a reaction medium
then provides compounds of formula VI, which are compounds of
formula I wherein R.sub.6 is H. Suitable bases include NaH,
Et.sub.3N, DIPEA, K.sub.2CO.sub.3 or Na.sub.2CO.sub.3 and suitable
reaction media include THF, DMF, CH.sub.2Cl.sub.2 or CH.sub.3CN.
Preferably, the base is NaH, Et.sub.3N or K.sub.2CO.sub.3 and the
solvent is CH.sub.3CN or DMF. More preferably, the base is
triethylamine and the reaction medium is acetonitrile. 8
[0093] Compounds of formula VIa, which are compounds of formula VI
wherein X is a valence bond, R.sub.2 represents CO.sub.2R.sub.2',
ZR.sub.5 represents CN, and R.sub.6 represents hydrogen, may be
saponified with a base to prepare carboxylic acids of formula VII
as shown above in Scheme B. Suitable bases NaOH, KOH, LiOH, and
Ba(OH).sub.2. Preferably, the base is an alkali metal hydroxide.
More preferably, the base is NaOH.
[0094] Compounds of formula VIII, which are compounds of formula VI
in which R.sub.2X is R.sub.2a'R.sub.2a"NC(O), ZR.sub.5 represents
CN, and R.sub.6 represents hydrogen, may be prepared via treatment
of compounds of formula VII with a coupling reagent and an amine of
formula NH.sub.2R.sub.2a'R.sub.2a" in a reaction medium. Suitable
coupling agents include PyBOP
[benzotriazol-1-yloxytripyrrolidino-phosphonium
hexafluorophosphate], BOP [benzotriazol-1-yloxytris(dimethylamino)
phosphonium hexafluorophosphate], CDI (N,N'-carbonyldiimidazole),
DCC (N,N'-dicyclohexylcarbodiimide), HBTU
[0-benzotriazol-1-yl-N,N,N',N'-tetr- anethyluronium
hexafluorophosphate], HOAt (1-hydroxy-7-azabenzotriazole) and HOBt
(1-hydroxybenzotriazole) and EDC [1-ethyl-3-(3-dimethylaminoprop-
yl) carbodimide]. Preferably, the coupling reagent is HOBt, PyBOP
or EDC. More preferably, the coupling reagent is HOBt or PyBOP.
[0095] Compounds of formula IX, which are compounds of formula VI
wherein XR.sub.2 is R.sub.2a'OC(O), ZR.sub.5 represents CN, and
R.sub.6 represents hydrogen, may be prepared via treatment of a
compound of formula VII with an acid or a base and an alcohol of
the formula R.sub.2a'OH. Suitable acids include HCl,
H.sub.2SO.sub.4, TsOH, 10-camphorsulfonic acid (CSA) and pyridinium
p-toluenesulfonates (PPTs). More preferably, the acid is
hydrochloric acid.
[0096] Compounds of formula X, which are compounds of formula VI
where XR.sub.2 is R.sub.2a'OC(O)NR.sub.2a", ZR.sub.5 represents CN,
and R.sub.6 represents hydrogen, may be prepared via treatment of
compounds of formula VII with an azidization reagent, that is, a
source of N.sub.3, followed by the addition of an alcohol of
formula R.sub.2a'OH. Suitable azidization reagents include
diphenylphosphorylazide (DPPA) and NaN.sub.3. Preferably, the
reagent is DPPA. 9
[0097] As shown in Scheme C, compounds of formula XII, which are
compounds of formula I wherein XR.sub.2 is NH(R.sub.2a').sub.2, may
be prepared via compounds of formula X where the carbalkoxy moiety
of the compound of formula X functions as a removable protecting
group. The intermediate compound of formula XI may be prepared by
removal of the carbalkoxy moiety of the compound of formula X.
Preferably, the carbalkoxy group will be a t-butoxycarbonyl (BOC)
or benzyloxycarbonyl (Cbz or Z). Suitable conditions for removing
these and other suitable protecting groups are disclosed in Green
and Wuts, supra. Preferably, the deprotection reaction is an acid
cleavage or a hydrogenation reaction.
[0098] Compounds of formula XII result from the reductive arination
of compounds of formula XI using an aldehyde of formula
R.sub.2a'CHO and a reducing agent in a suitable reaction medium.
Suitable reducing agents include NaBH.sub.4, LiBH.sub.4,
diisobutylaluminum hydride (DIBAL-H), lithium aluminum hydride
(LAH) and NaBH(OAc).sub.3. Preferably, the reducing agent is
NaBH(OAc).sub.3 or NaBH.sub.4. More preferably, the reducing agent
is NaBH(OAc).sub.3. Suitable reaction media include
1,2-dichloroethane, CH.sub.2Cl.sub.2, THF and CH.sub.3CN. Preferred
reaction media include 1,2-dichloroethane and CH.sub.2Cl.sub.2 and
more preferably, the reduction is carried out in
1,2-dichloroethane.
[0099] Alternatively, preparation of compounds of formula XII may
be accomplished via treatment of compounds of formula XI with a
base and a reagent of formula R.sub.2a'L. Suitable bases include
K.sub.2CO.sub.3, NaHCO.sub.3, Et.sub.3N, DIPEA, Cs.sub.2CO.sub.3,
DBU and pyridine. Preferably, the base is selected from the group
consisting of K.sub.2CO.sub.3, NaHCO.sub.3 and Et.sub.3N. More
preferably, the base is sodium bicarbonate. 10
[0100] Compounds of formula XV may be prepared via the method shown
in Scheme D. Net reduction of compounds of formula VI wherein
XR.sub.2 is R.sub.2'OC(O) provides aldehydes of formula XIII.
Suitable means of carrying out a net reduction include reaction
with a reducing agent or sequential reaction with a stronger
reducing agent and a weaker oxidizing agent. Suitable reducing
agents are generally known to those skilled in the art and can be
fuond in references such as Advanced Organic Chemistry III ed.,
Part B: Reactions and Synthesis, Francis A. Carey, Richard J.
Sundberg, Plenum Publishing Corp., NY (1993) and March's Advanced
Organic Chemistry: Reactions, Mechanisms and Structure. 5.sup.th
ed., Wiley-InterScience, John Wiley & Sons, Inc., NY (2001)
(both herein incorporated by reference).
[0101] Suitable combinations of oxidizing agents and reducing
agents include diisobutylaluminum hydride (DIBAL-H) with MnO.sub.2,
Preferably, net reduction is accomplished by sequential reduction
and oxidation with a reducing agent such as DIBAL-H, LAH,
NaBH.sub.4 or LiBH.sub.4, and an oxidizing agent such as MnO.sub.2,
SO.sub.3-pyridine, TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy,
free radical), Dess-Martin periodinane, or TPAP
(tetrapropylammonium perruthenate) and NMO
(N-methylmorpholine-N-oxide) in combination. More preferably, the
reduction is carried out first with DIBAL-H to produce an
intermediate compound, which is then oxidized with MnO.sub.2 to
yield the compound of formula XIII.
[0102] Subsequent treatment with an oxidizing agent in a suitable
reaction medium followed by etherification using a base in a
suitable reaction medium and a reagent of formula R.sub.2a'L yields
compounds of formula XV, which are compounds of formula VI wherein
XR.sub.2 is OR.sub.2a'. Suitable oxidizing agents include
m-chloroperbenzoic acid (m-CPBA) and H.sub.2O.sub.2. Suitable bases
include NaH, Et.sub.3N, DIPEA and K.sub.2CO.sub.3. Suitable
reaction media include THF, DMF, CH.sub.2Cl.sub.2 and CH.sub.3CN.
More preferably, the oxidizing agent is m-chloro perbenzoic acid
(m-CPBA), the base is NaH, and the reaction medium is
tetrahydrofuran (THF) or DMF. 11
[0103] Halogenation of the 5-methyl group of a compound of formula
V may be effected by treatment with a halogenating agent. Suitable
halogenating agents include, but are not limited to sulfuryl
choride, N-Iodosuccinimide, N-Bromosuccinimide,
N-chlorosuccinimide, oxalyl choride. Preferably the halogenating
agent is N-bromosuccinimide or sulfuryl chloride. The halogenation
can be performed under an inert atmosphere, such as N.sub.2, in the
presence of a catalyst, to produce a halogenated pyrrole
intermediate of formula XVI. Preferably, the catalyst is dibenzoyl
peroxide or 2,2'-azobisisobutyronitrile, or irradiation.
[0104] Treatment of a pyrrole of formula XVI with a thiol of
formula HSR.sub.3a', an alcohol intermediate of formula
HOR.sub.3a', or a primary or a secondary amine of formula
HNR.sub.3a'R.sub.3a" in the presence of a base affords a pyrrole of
formula XVII. Suitable bases include NaHCO.sub.3, diisopropyle
ethylamine DBU, KHCO.sub.2, and trimethylamine. Preferably, the
base is NaHCO.sub.3 or triethylamine. Acetonitrile is one suitable
reaction medium for this reaction.
[0105] Treatment of a pyrrole of formula XVII with a reagent of
formula HYR.sub.4, at room temperature in the presence of a base
yields the compound of formula XVIII. Preferably, the base is
NaHCO.sub.3 or triethylamine. Acetonitrile is one suitable reaction
medium for this reaction. Heating the pyrrole of formula XVII with
a reagent of formula HYR.sub.4 in the absence of base also affords
the compound of formula XVIII. 12
[0106] Compound XIX, which is a compound of formula VI wherein
R.sub.3 is --CH.sub.2SR.sub.3a' (see Scheme A, above), can be
oxidized to the sulfoxide of compound XX, wherein n=1, or the
sulfone of compound XX, wherein n=2. Suitable oxidizing agents
include m-chloroperbenzoic acid (MCPDA), tBu-OOH, H.sub.2O.sub.2
NaIO.sub.4, and dimethyldioxirane. Preferably, the oxidizing agent
is MCPDA. The number of equivalents of oxidizing agent added to the
reaction mixture will determine the final oxidation state of the
sulfur atom. A compound of formula XX wherein n=1 or 2 can be
heated with an excess of an alcohol of formula HOR.sub.3b' or a
primary or secondary amine of formula HNR.sub.3b'R.sub.3b" to yield
a compound of formula XXI. 13
[0107] Compounds of formula VII, from Scheme B, undergo a Wittig
reaction with a phosphonate in the presence of a base to afford a
compound of formula XXII. Suitable phosphonates known to those
skilled in the art may be used. Preferably, the phosphonate is
methyl diethylphosphonoacetate. Dichloroethane and the like are
suitable organic reaction media for Wittig reactions. Suitable
bases include KH, K2CO3, N-Butyllithium, sec-Butyllithium,
tert-Butyllithium, NaH, preferably NaH.
[0108] The double bond in the R2 group of the compound of formula
XXII may be hydrogenated in the presence of a catalyst to yield a
compound of formula XXIII. Suitable catalysts include PtO2,
palladium on carbon (Pd/C), Pd(OH)2, and Raney Ni. Preferably, the
catalyst is Pd/C.
[0109] Esters of formula XXIII may be hydrolyzed by techniques well
known in the art, for example those taught in Green and Wuts,
supra, preferably base hydrolysis with NaOH. Subsequent coupling of
the resulting acid with an amine in the presence of a coupling
agent affords the amide of formula XXIV. Suitable coupling agents
are known to those skilled in the art and include those described
in The Practice of Peptide Synthesis, 2.sup.nd Ed., by Bodanszy,
Miklos, Springer-Velag (1993) (herein incorporated by reference).
Preferably the coupling agent is NAN'-dicyclohexylcarbodiimid- e
(DCC). 14
[0110] Scheme H depicts an alternative route to the synthesis of
compounds of formula IV (see Scheme A, above). Condensation of a
pyrrole of formula III with a reagent of formula
R.sub.6C(O)CH.sub.2ZR.sub.5, followed by base induced cyclization
in a suitable reaction medium, yields the intermediate of formula
IV. Suitable bases include DBU, NaH, BuLi, Et.sub.3N and DIPEA.
Suitable reaction media include toluene, THF, CH.sub.2Cl.sub.2,
DMF, toluene and CH.sub.3CN. Preferably the base is NaH, DBU, or
DIPEA and the reaction medium is DMF, toluene or THF. Reagents of
formula R.sub.6C(O)CH.sub.2ZR.sub.5, particularly those wherein
R.sub.6 is a substituted oxygen, nitrogen or an alkyl group and
ZR.sub.5 is a nitrile group, can be purchased from commerical
sources or else readily synthesized by those of skill in the art.
15
[0111] As shown in scheme I, a compound of formula XXV, which is a
compound of formula VI wherein ZR.sub.5 is a nitrile group (see
Scheme A, above), can be reduced in the presence of hydrogen and a
catalyst to yield a compound of formula XXVI. Suitable catalysts
include PtO.sub.2, Pd/C, Pd(OH).sub.2, and Raney Ni. Preferably,
the catalyst is palladium on carbon (Pd/C).
[0112] Compounds of formula XXVI, when combined with a reagent of
formula R.sub.5aL, wherein L is a leaving group, e.g., a halogen,
in the presence of a base, yield compounds of formula XXVII.
Suitable bases KH, K.sub.2CO.sub.3, N-Butyllithium,
sec-Butyllithium, tert-Butyllithium, and NaH. Preferably, the base
is NaH. Reagents of formula R.sub.5aL are readily available from
commercial sources.
[0113] Additionally, a compound of formula XXVI can be treated with
a reagent of formula (R.sub.5b-Z.sub.b-C(O)).sub.2O or
R.sub.5b-Z.sub.b-C(O)-L', wherein L' is a leaving group, e.g., a
halogen, in the presence of a base to yield a compound of formula
XXVII. Suitable bases include NaHCO.sub.3, diisopropylethylamine,
DBU, KHCO.sub.3, trimethylamine. Preferably, the base is
triethylamine. Reagents of formula R.sub.5b-Z.sub.b-C(O)-L' or
(R.sub.5b-Z.sub.b-C(O)).sub.2O are readily available from
commercial sources, or may be synthesized by those of skill in the
art. 16
[0114] Scheme J depicts the synthesis of a compound of formula XXX,
which is a compound of formula IV (see scheme A, above) wherein
R.sub.6 is hydrogen and ZR.sub.5 is a nitrile group. Treatment of a
pyrrole of formula III with a reactive intermediate in a high
boiling protic solvent yields an intermediate of formula XXIX.
Preferably, dimethylformamide (DMF) and dimethylacetamide are
used.
[0115] Treatment of the intermediate of formula XXIX with
acetonitrile in the presence of a base results in cyclization to
produce the compound of formula XXX. Suitable bases include, but
are not limited to KH, NaH, sec-butyllithium, and preferably
N-Butyllithium. As shown in scheme A, above, compounds of formula
XXX are intermediates in the synthesis of compounds of formula I
wherein R.sub.6 is hydrogen and ZR.sub.5 is a nitrile group. 17
[0116] The synthesis of compounds of formula XXXIV and XXXV is
shown in Scheme K. Compounds of formula XXXIV and XXXV are
intermediates of formula IV (see scheme A, above) wherein R.sub.6
is hydrogen and ZR.sub.5 is a nitrile group. Treatment of a pyrrole
of formula XXXI with a reactive intermediate of formula XXXII in a
high boiling solvent yields an intermediate of formula XXXIII.
Suitable solvents include but are not limited to xylene,
nitrobenzene and toluene, preferably toluene.
[0117] Further heating of an intermediate of formula XXXIII in a
high boiling solvent results in cyclization to yield intermediates
of formula XXXIV and XXXV. Suitable solvents include but are not
limited to DMF, DMA, N-methylpyrolidinone, preferably Dowtherm.TM.,
or toluene in a high pressure apparatus. 18
[0118] The synthesis of compounds of formula XXXVI is shown in
Scheme L. Treatment a compound of formula XXI, from Scheme F, with
a reactive intermediate of formula X'C(O)X' or X'C(O)OC(O)X', as
decribed in Scheme L, in presence of a base such as
diisopropylethyl amine or triethyl amine, with or without heating,
yields a compound of formula XXXVI. Suitable solvents include, but
are not limited to, methylene chloride, chloroform,
tetrahydrofurane or ethyl acetate.
[0119] As shown in scheme K, above, compounds of formula XXXIV and
XXXV are intermediates in the synthesis of compounds of formula I
wherein R.sub.6 is hydrogen and ZR.sub.5 is a nitrile group. The
reactive intermediate of formula XXXII and related reagents of this
structure are readily available from commercial sources, or may be
synthesized by those of skill in the art.
[0120] Schemes 1 through 5, below, summarize several preferred
methods of making some of the compounds of the invention. In
schemes 1 through 5, unless otherwise noted, X, Y, Z, R.sub.1,
R.sub.2, R.sub.2', R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are as
defined above, and L represents a leaving group. Variables
designated with the subscript "a" or "b" have the same scope as,
but are chosen independently of, their parent variable. 19
[0121] 3-Cyanopyrrolopyridazines of formula VI may be prepared in
accordance with Scheme 1. Pyrroles of formula II may be obtained by
the processes described in Patent Cooperation Treaty (PCT)
publication numberWO 00/71129, pending U.S patent application Ser.
No. 09/573829, pending PCT Application Number US01/49982 and
pending U.S. patent application Ser. No. 10/036293 (all of which
are herein incorporated by reference in their entirety).
[0122] Treatment of a pyrrole of formula II with a base such as
sodium hydride in a reaction medium such as DMF followed by the
addition of an aminating reagent such as 2,4-dinitroaminophenol
generates an aminopyrrole of formula III. Condensation with an
acetal such as 1,1-diethoxypropionitrile followed by base induced
cyclization employing a base such as DBU or diisopropylethylarnine,
in a reaction medium such as toluene, provides the
3-cyanopyrrolopyridazine of formula IV. Conversion to the 4-chloro
compounds of formula V can then be accomplished using a
chlorinating reagent such as POCl.sub.3 or POCl.sub.5. Treatment of
a compound of formula V with a reagent of formula HY-R.sub.4 in the
presence of a base such as triethylamine in a reaction medium such
as acetonitrile provides compounds of formula VI, which are
compounds of formula I wherein XR.sub.2 is C(O)OEt, Z is a valence
bond, R.sub.5 is CN, and R.sub.6 is H. 20
[0123] Compounds of formula VI may be saponified with a base such
as NaOH to prepare carboxylic acids of formula VII as shown above
in Scheme 2. Compounds of formula VIII, which are compounds of
formula I wherein XR.sub.2 is C(O)NHR.sub.2', Z is a valence bond,
R.sub.5 is CN, and R.sub.6 is H, may be prepared via treatment of
compounds of formula VII with a coupling reagent such as EDC and an
amine such as NHR.sub.2a'R.sub.2a", in a reaction medium such as
dichloromethane. Compounds of formula IX, which are compounds of
formula I wherein XR.sub.2 is C(O)OR.sub.2', Z is a valence bond,
R.sub.5 is CN, and R.sub.6 is H, may be prepared via treatment of
compound VII with an acid such as hydrochloric acid and an alcohol
of formula R.sub.2'OH. Compounds of formula X, which are compounds
of formula I where XR.sub.2 is NHC(O)OR.sub.2', Z is a valence
bond, R.sub.5 is CN, and R.sub.6 is H, may be prepared via
treatment of compounds of formula VII with a reagent such as DPPA
followed by the addition of an alcohol of the formula R.sub.2'OH.
21
[0124] As shown in Scheme 3, compounds of formula XII, which are
compounds of formula I wherein XR.sub.2 is NHR.sub.2a', Z is a
valence bond, R.sub.5 is CN, and R.sub.6 is H, may be prepared from
compounds of formula X where the carbalkoxy of the compound of
formula X is a removable protecting group (e.g., R.sub.2' is
t-butyl or benzyl). The compound of formula XI may be prepared by
deprotection, i.e., acid cleavage or hydrogenation, respectively.
Compounds of formula XII may then be prepared via reductive
amination of compounds of formula XI using an aldehyde of formula
R.sub.2a'CHO and a reducing agent such as NaBH(OAc).sub.3 in a
reaction medium such as 1,2-dichloroethane. Alternatively,
preparation of compounds of formula XII may be accomplished via
treatment of compounds of formula XI with a base such as
NaHCO.sub.3 and a reagent of formula R.sub.2a'L. 22
[0125] Compounds of formula XV may be prepared via the method shown
in Scheme 4. Reduction of compounds of formula VI with a reducing
agent such as DIBAL-H in reaction media such as dichloromethane or
toluene, followed by oxidation with an oxidizing agent such as
MnO.sub.2, provides aldehydes of formula XIII. Treatment of
compounds of formula XIII with a peracid such as m-CPBA in a
reaction medium such as dichloromethane followed by etherification
using a base such as sodium hydride in reaction mediums such as
tetrahydrofuran or DMF and a reagent of formula R.sub.2a'L yields
compounds of formula XV, which are compounds of formula I where
XR.sub.2 is OR.sub.2a', Z is a valence bond, R.sub.5 is CN, and
R.sub.6 is H. 23
[0126] As shown in Scheme 5, an intermediate of formula XXV, which
is a compound of formula I wherein R.sub.6 is H and ZR.sub.5 is a
nitrile group, can be reduced in the presence of hydrogen,
trifluoroacetic acid (TFA), and a catalyst such as Pd/C to yield an
compound of formula XXVI. The compound of formula XXVI can be
treated with intermediates of formula R.sub.5b-Z.sub.b-C(O)--Cl or
(R.sub.5b-Z.sub.b-C(O)).sub.2O in the presence of a base such as
triethylamine to yield the compound of formula XXVII. Intermediates
of formula R.sub.5b-Z.sub.b-C(O)--Cl and
(R.sub.5b-Z.sub.b-C(O)).sub.2O are readily available from
commercial sources, or may be synthesized by those of skill in the
art.
[0127] Solvates (e.g., hydrates) of the compounds of formula I are
also within the scope of the present invention. Methods of
solvation are generally known in the art. Accordingly, the
compounds of the instant invention may be in the free or solvated
form.
[0128] The compounds of formula I may be present as salts, in
particular pharmaceutically acceptable salts. Compounds of formula
I having, for example, at least one basic center can form acid
addition salts. These are formed, for example, with strong
inorganic acids, such as mineral acids, for example sulfuric acid,
phosphoric acid or a hydrohalic acid, with strong organic
carboxylic acids, such as alkanecarboxylic acids of 1 to 4 carbon
atoms which are unsubstituted or substituted, for example, by
halogen, for example acetic acid, such as saturated or unsaturated
dicarboxylic acids, for example oxalic, malonic, succinic, maleic,
fumaric, phthalic or terephthalic acid, such as hydroxycarboxylic
acids, for example ascorbic, glycolic, lactic, malic, tartaric or
citric acid, such as amino acids, (for example aspartic or glutamic
acid or lysine or arginine), or benzoic acid, or with organic
sulfonic acids, such as (C.sub.1-C.sub.4) alkyl or arylsulfonic
acids which are unsubstituted or substituted, for example by
halogen, for example methanesulfonic acid or p-toluenesulfonic
acid. Corresponding acid addition salts can also be formed having,
if desired, an additional basic center.
[0129] The compounds of formula I having at least one acid group
(for example COOH) can also form salts with bases. Suitable salts
with bases are, for example, metal salts, such as alkali metal or
alkaline earth metal salts, for example sodium, potassium or
magnesium salts, or salts with ammonia or an organic amine, such as
morpholine, thiomorpholine, piperidine, pyrrolidine, a mono-, di-,
or tri-lower alkylamine, for example ethyl, t-butyl, diethyl,
diisopropyl, triethyl, tributyl or dimethyl-propylamine, or a mono,
di, or trihydroxy lower alkylamine, for example mono, di or
triethanolamine.
[0130] Corresponding internal salts may furthermore be formed.
Salts which are unsuitable for pharmaceutical uses but which can be
employed, for example, for the isolation or purification of free
compounds of formula I or their pharmaceutically acceptable salts,
are also included within the scope of this invention.
[0131] Preferred salts of the compounds of formula I which include
a basic group include monohydrochloride, hydrogen sulfate,
methanesulfonate, phosphate or nitrate.
[0132] Preferred salts of the compounds of formula I which include
an acid group include sodium, potassium and magnesium salts and
pharmaceutically acceptable organic amines.
[0133] Methods of Using the Compounds
[0134] It has been discovered that pyrrolopyridazines of the
invention are inhibitors of protein kinases. More specifically,
certain pyrrolopyridazines inhibit the effects of receptor tyrosine
kinases and serine/threonine kinases, a property of value in the
treatment of disease states associated with hyperproliferation,
angiogenesis, increased vascular permeability, and inflammation,
such as cancer and inflammatory disease. In particular, the
compounds of formula I and their salts, solvates, and stereoisomers
are expected to inhibit the growth of primary and recurrent solid
tumors by antiproliferative and/or antiangiogenic mechanisms. The
solid tumors include, for example, cancers of the bladder, squamous
cell, head, colorectal, oesophageal, gynecological (such as
ovarian), pancreas, breast, prostate, lung, vulva, skin, brain,
genitourinary tract, lymphatic system (such as thyroid), stomach,
larynx and lung
[0135] In some embodiments of the present invention, methods are
provided for treating proliferative or inflammatory diseases
comprising administering to a patient in need thereof a
therapeutically effective amount of a compound having formula I, as
described above.
[0136] The methods optionally comprise administering at least one
other therapeutic agent such as angiogenesis inhibitors,
antiestrogens, progestogens, aromatase inhibitors, antihormones,
antiprogestogens, antiandrogens, LHRH agonists and antagonists,
testosterone 5.alpha.-dihydroreductase inhibitors, farnesyl
transferase inhibitors, anti-invasion agents, growth factor
inhibitors, antimetabolites, intercalating antitumour antibiotics,
platinum derivatives, alkylating agents, antimitotic agents,
topoisomerase inhibitors, cell cycle inhibitors, and biological
response modifiers, linomide, integrin .alpha.v.beta.3 function
inhibitors, angiostatin, razoxin, tamoxifen, toremifen, raloxifene,
droloxifene, iodoxyfene, megestrol acetate, anastrozole, letrazole,
borazole, exemestane, flutamide, nilutamide, bicalutamide,
cyproterone acetate, gosereline acetate, luprolide, finasteride,
metalloproteinase inhibitors, urokinase plasminogen activator
receptor function inhibitors, growth factor antibodies, growth
factor receptor antibodies, tyrosine kinase inhibitors,
serine/threonine kinase inhibitors, methotrexate, 5-fluorouracil,
purine, adenosine analogues, cytosine arabinoside, doxorubicin,
daunomycin, epirubicin, idarubicin, mitomycin-C, dactinomycin,
mithramycin, cisplatin, carboplatin, nitrogen mustard, melphalan,
chlorambucil, busulphan, cyclophosphamide, ifosfamide nitrosoureas,
thiotephan, vincristine, taxol, taxotere, epothilone analogs,
discodermolide analogs, eleutherobin analogs, etoposide,
teniposide, amsacrine, topotecan, flavopyridols, and biological
response modifiers. In some preferred embodiments, the additional
thereapeutic agent is selected from Erbitux.TM., taxol, paraplatin
and Ifex.
[0137] More generally, the compounds of formula I are useful in the
treatment of a variety of cancers, including, but not limited to,
the following:
[0138] carcinoma, including that of the bladder, breast, colon,
kidney, liver, lung, including small cell lung cancer, esophagus,
gall bladder, ovary, pancreas, stomach, cervix, thyroid, prostate,
and skin, including squamous cell carcinoma;
[0139] hematopoietic tumors of lymphoid lineage, including
leukemia, acute lymphocytic leukemia, acute lymphoblastic leukemia,
B-cell lymphoma, T-cell lymphoma, Hodgkins lymphoma, non-Hodgkins
lymphoma, hairy cell lymphoma and Burkett's lymphoma;
[0140] hematopoietic tumors of myeloid lineage, including acute and
chronic myelogenous leukemias, myelodysplastic syndrome and
promyelocytic leukemia;
[0141] tumors of mesenchymal origin, including fibrosarcoma and
rhabdomyosarcoma;
[0142] tumors of the central and peripheral nervous system,
including astrocytoma, neuroblastoma, glioma and schwannomas;
and
[0143] other tumors, including melanoma, seminoma, teratocarcinoma,
osteosarcoma, xenoderoma pigmentosum, keratoctanthoma, thyroid
follicular cancer and Kaposi's sarcoma.
[0144] The compounds of formula I are especially useful in
treatment of tumors having a high incidence of protein kinase
activity, such as colon, lung, prostate, breast and pancreatic
tumors. By the administration of a composition comprising a
compound of the invention, or a combination of such compounds,
development of tumors in a mammalian host is reduced.
[0145] Compounds of formula I may also be useful in the treatment
of diseases other than cancer that may be associated with signal
transduction pathways operating through growth factor receptors.
For example, due to the key role of kinases in the regulation of
cellular proliferation in general, kinase inhibitors could act as
reversible cytostatic agents which may be useful in the treatment
of any disease process which features abnormal cellular
proliferation, e.g., benign prostate hyperplasia, familial
adenomatosis polyposis, neuro-fibromatosis, atherosclerosis,
pulmonary fibrosis, arthritis, psoriasis, glomerulonephritis,
restenosis following angioplasty or vascular surgery, hypertrophic
scar formation, inflammatory bowel disease, transplantation
rejection, endotoxic shock, and fungal infections.
[0146] In addition, compounds of formula I may induce or inhibit
apoptosis. The apoptotic response is aberrant in a variety of human
diseases. Compounds of formula I, as modulators of apoptosis, will
be useful in the treatment of cancer (including but not limited to
those types mentioned hereinabove), viral infections (including but
not limited to herpevirus, poxvirus, Epstein-Barr virus, Sindbis
virus and adenovirus), prevention of AIDS development in
HIV-infected individuals, autoimmune diseases (including but not
limited to systemic lupus erythematosus, autoimmune mediated
glomerulonephritis, rheumatoid arthritis, psoriasis, inflammatory
bowel disease, and autoimmune diabetes mellitus), neurodegenerative
disorders (including but not limited to Alzheimer's disease,
AIDS-related dementia, Parkinson's disease, amyotrophic lateral
sclerosis, retinitis pigmentosa, spinal muscular atrophy and
cerebellar degeneration), myelodysplastic syndromes, aplastic
anemia, ischemic injury associated with myocardial infarctions,
stroke and reperfusion injury, arrhythmia, atherosclerosis,
toxin-induced or alcohol related liver diseases, hematological
diseases (including but not limited to chronic anemia and aplastic
anemia), degenerative diseases of the musculoskeletal system
(including but not limited to osteoporosis and arthritis)
aspirin-sensitive rhinosinusitis, cystic fibrosis, multiple
sclerosis, kidney diseases and cancer pain.
[0147] As inhibitors of protein kinases, compounds of the present
invention have utility in treating conditions associated with
inappropriate kinase activity. Such conditions also include
diseases in which cytokine levels are modulated as a consequence of
intracellular signaling, and in particular, diseases that are
associated with an overproduction of such cytokines as IL-1, IL-4,
IL-8 and TNF-.alpha.. For example, compounds of the present
invention are useful in treating and preventing:
[0148] IL-1 mediated diseases such as, for example, rheumatoid
arthritis, osteoarthritis, stroke, endotoxemia and/or toxic shock
syndrome, inflammatory reaction induced by endotoxin, inflammatory
bowel disease, tuberculosis, atherosclerosis, muscle degeneration,
cachexia, psoriatic arthritis, Reiter's syndrome, gout, traumatic
arthritis, rubella arthritis, acute synovitis, diabetes, pancreatic
.beta.-cell disease and Alzheimer's disease;
[0149] IL-4 mediated diseases or conditions such as, for example,
allergic inflammatory processes including those that occur in
asthma,
[0150] IL-8 mediated diseases or conditions such as, for example,
those characterized by massive neutrophil infiltration, such as
psoriasis, inflammatory bowel disease, asthma, cardiac and renal
reperfusion injury, adult respiratory distress syndrome, thrombosis
and glomerulonephritis; and
[0151] TNF-mediated diseases or conditions such as rheumatoid
arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis
and other arthritic conditions, sepsis, septic shock syndrome,
adult respiratory distress syndrome, cerebral malaria, chronic
pulmonary inflammatory disease, silicosis, pulmonary sarcoisosis,
bone resorption disease, reperfusion injury, graft vs. host
reaction, allograft rejections, fever and myalgias due to
infection, cachexia secondary to infection, AIDS, ARC or
malignancy, meloid formation, scar tissue formation, Crohn's
disease, ulcerative colitis, pyresis, viral infections, such as
HIV, CMV, influenza and herpes; and veterinary viral infections,
such as lentivirus infections, including, but not limited to equine
infectious anemia virus; or retrovirus infections, including feline
immunodeficiency virus, bovine immunodeficiency virus, or canine
immunodeficiency virus.
[0152] Diseases mediated by p38 include rheumatoid arthritis (RA),
chronic obstructive pulmonary disease (COPD), asthma, Crohn's
disease, neurological diseases such as Alzheimer's disease and
stroke, and inflammatory bone diseases. A further discussion of
diseases mediated by p38 can be found in pending PCT Application
Number US01/49982 and pending U.S. patent application Ser. No.
10/036293 (both of which are herein incorporated by reference in
their entirety).
[0153] Inhibitors of protein kinase activity, such as the compounds
of the present invention, are useful in treating and preventing
other conditions and classes of conditions including, but not
limited to, inflammatory diseases, autoimmune diseases, destructive
bone disorders, proliferative disorders, angiogenic disorders,
infectious diseases, neurodegenerative diseases, viral diseases,
allergies, myocardial ischemia, reperfusion/ischemia in stroke
heart attacks, organ hyposia, vascular hyperplasia, cardiac
hypertrophy, thrombin-induced platelet aggregation, and conditions
associated with prostaglandin endoperoxidase synthase-2.
[0154] Inflammatory diseases which may be treated or prevented
include, but are not limited to, acute pancreatitis, chronic
pancreatitis, asthma, allergies and adult respiratory distress
syndrome.
[0155] Autoimmune diseases which may be treated or prevented
include, but are not limited to, glomerulonephritis, rheumatoid
arthritis, systemic lupus erythematosis, scleroderma, chronic
thyroiditis, Grave's disease, autoimmune gastritis, diabetes,
autoimmune hemolytic anemia, autoimmune neutropenia,
thrombocytopenia, atopic dermatitis, chronic active hepatitis,
myasthenia gravis, multiple sclerosis, inflammatory bowel disease,
ulcerative colitis, Crohn's disease, psoriasis, or graft vs. host
disease.
[0156] Destructive bone disorders which may be treated or prevented
include, but are not limited to, osteoporosis, osteoarthritis and
multiple myeloma-related bone disorder.
[0157] Proliferative diseases which may be treated or prevented
include, but are not limited to, acute myelogenous leukemia,
chronic myelogenous leukemia, metastatic melanoma, Kaposi's
sarcoma, and multiple myeloma.
[0158] Angiogenic disorders which may be treated or prevented
include hemangiomas, psoriasis, Kaposi's sarcoma, ocular
neovascularization, retinopathy of prematurity, macular
degeneration, diabetic retinopathy, diabetic nephropathy,
rheumatoid arthritis, endometriosis, atherosclerosis, tumor growth
and metastasis, myocardial ischemia, peripheral ischemia, cerebral
ischemia, impaired wound healing, certain female reproductive
disorders, organ hypoxia, and impaired ulcer healing.
[0159] Infectious diseases which may be treated or prevented
include, but are not limited to, sepsis, septic shock, and
Shigellosis.
[0160] Neurodegenerative diseases which may be treated or prevented
by the compounds of this invention include, but are not limited to,
Alzheimer's disease, Parkinson's disease, cerebral ischemias or
neurodegenerative disease caused by traumatic injury.
[0161] Viral diseases which may be treated or prevented include,
but are not limited to, acute hepatitis infection (including
hepatitis A, hepatitis B and hepatitis C), HIV infection and CMV
retinitis.
[0162] The compounds of formula I may also prevent blastocyte
implantation, and, therefore, may be used as contraceptives in
mammals.
[0163] In addition, protein kinase inhibitors of this invention
also exhibit inhibition of the expression of inducible
pro-inflammatory proteins such as prostaglandin endoperoxide
synthase-2 (PGHS-2), also referred to as cyclooxygenase-2 (COX-2).
Accordingly, additional conditions which may be treated or
prevented by appropriate administration of compounds of the
invention include edema, analgesia, fever and pain, such as
neuromuscular pain, headache, pain caused by cancer, dental pain
and arthritis pain.
[0164] In the field of medical oncology, it is normal practice to
combine different agents for treatment of patients with cancer.
Thus, a compound of formula I may optionally be combined with other
components, such as antiproliferative, antiangiogenic and/or
vascular permeability reducing agents. Additionally, surgery,
radiotherapy or chemotherapy may optionally be utilized in
conjunction with administration of compounds of formula I.
Accordingly, the compound of formula I may be administered alone or
combined with the administration of one or more other therapeutic
agents, substances and/or treatments.
[0165] Such conjoint treatment may be achieved by way of the
simultaneous, sequential or separate administration of the
individual components of the treatment. When not administered
simultaneously, the component therapies may be administered in any
order. If formulated as a fixed dose, such combination products
employ the compounds of this invention within the dosage range
described below and the other pharmaceutically active agent within
its approved dosage range. Dosage ranges of many pharmaceutically
active agents may be found in the Physician's Desk Reference,
55.sup.th Edition, Medical Economics Company (2001). Compounds of
formula I may also be used sequentially with known anticancer or
cytotoxic agents and treatment, including radiation, when a
combination formulation is inappropriate.
[0166] In general, there are three main categories of
chemotherapeutic agents:
[0167] (i) antiangiogenic agents, for example, linomide, inhibitors
of integrin .alpha.v.beta.3 function, angiostatin, and razoxin;
[0168] (ii) cytostatic agents such as antiestrogens (for example
tamoxifen, toremifen, raloxifene, droloxifene, iodoxyfene),
progestogens (for example megestrol acetate), aromatase inhibitors
(for example anastrozole, letrazole, borazole, exemestane),
antihormones, antiprogestogens, antiandrogens (for example,
flutamide; nilutamide; bicalutamide; cyproterone acetate;
(R)-2,3,4,5-tetrahydro-1-(1H-imidazole-
-4-ylmethyl)-3-(phenylmethyl)-4-(2-theienylsufonyl)-1H-1,4-benzodiazepine--
7-carbonitrile, mesylate salt;
N-[5-[[[5-(1,1-Dimethylethyl)-2-oxazolyl]me-
thyl]thio]-2-thiazolyl]-4-piperidinecarboxamide, hemi L-Tartaric
acid salt; cetuximab; molecules disclosed in pending U.S. patent
application Ser. No. 10/025,116 (herein incorporated by reference),
LHRH agonists and antagonists (for example gosereline acetate,
luprolide), inhibitors of testosterone 5.alpha.-dihydroreductase
(for example finasteride), farnesyl transferase inhibitors,
anti-invasion agents (for example metalloproteinase inhibitors like
marimastat and inhibitors of urokinase plasminogen activator
receptor function) and inhibitors of growth factor function, (such
growth factors include for example EGF, FGF, platelet derived
growth factor and hepatocyte growth factor such inhibitors include
growth factor antibodies, growth factor receptor inhibitors);
and
[0169] (iii) antiproliferative/antineoplastic drugs and
combinations thereof, as used in medical oncology, such as
antimetabolites (for example antifolates like methotrexate,
fluoropyrimidines like 5-fluorouracil, purine and adenosine
analogues, cytosine arabinoside); intercalating antitumour
antibiotics (for example anthracyclines like doxorubicin,
daunomycin, epirubicin and idarubicin, mitomycin-C, dactinomycin,
mithramycin); platinum derivatives (for example cisplatin,
carboplatin); alkylating agents (for example nitrogen mustard,
melphalan, chlorambucil, busulphan, cyclophosphamide, ifosfamide
nitrosoureas, thiotephan); antimitotic agents (for example vinca
alkaloids like vincristine and taxoids like taxol, taxotere and
newer microbtubule agents such as epothilone analogs,
discodermolide analogs, and eleutherobin analogs); topoisomerase
inhibitors (for example epipodophyllotoxins like etoposide and
teniposide, amsacrine, topotecan); cell cycle inhibitors (for
example flavopyridols); and biological response modifiers.
Particular compounds could include
N-[5-[[[5-(1,1-Dimethylethyl)-2-oxazolyl]methyl]thio]-2-thiazolyl]-4-pipe-
ridinecarboxamide, hemi L-Tartaric acid salt.
[0170] The compounds of formula I and the pharmaceutical
compositions comprising compounds of formula I may be administered
by any means suitable for the condition to be treated, which may
depend on the need for site-specific treatment or quantity of drug
to be delivered. The compounds may be administered in a dosage
range of about 0.05 to 200 mg/kg/day, preferably less than 100
mg/kg/day, in a single dose or in 2 to 4 divided doses.
[0171] Topical administration is generally preferred for
skin-related diseases, and systematic treatment is preferred for
cancerous or pre-cancerous conditions, although other modes of
delivery are contemplated. For example, the compounds and
compositions may be delivered orally, such as in the form of
tablets, capsules, granules, powders, or liquid formulations
including syrups; topically, such as in the form of solutions,
suspensions, gels or ointments; sublingually; bucally;
parenterally, such as by subcutaneous, intravenous, intramuscular
or intrasternal injection or infusion techniques (e.g., as sterile
injectable aqueous or non-aqueous solutions or suspensions);
nasally such as by inhalation spray; topically, such as in the form
of a cream or ointment; rectally such as in the form of
suppositories; or liposomally.
[0172] Dosage unit formulations containing non-toxic,
pharmaceutically acceptable carriers, vehicles or diluents may be
administered. The compounds and compositions may be administered in
a form suitable for immediate release or extended release.
Immediate release or extended release may be achieved with suitable
pharmaceutical compositions or, particularly in the case of
extended release, with devices such as subcutaneous implants or
osmotic pumps. Further techniques for formulation and
administration of the compounds and compositions of the instant
application may be found in "Remington's Pharmaceutical Sciences,"
18.sup.th Ed. (1990, Mack Publishing Co., Easton, Pa.).
[0173] Abbreviations
[0174] The following abbreviations are among those used herein:
[0175] .DELTA.=heat
[0176] Ac=acetyl
[0177] AcOH=acetic acid
[0178] aq.=aqueous
[0179] ATP=adenosine triphosphate
[0180] BOP=benzotriazol-1-yloxytris(dimethylamino)-phosphonium
[0181] BSA=Bovine serum albumin
[0182] DBU=1,8-diazabicyclo[5.4.0]undec-7-ene
[0183] DCC=Dicyclohexylcarbodiimide
[0184] DCE=dichloroethane
[0185] DEAD=diethyl azodicarboxylate
[0186] DIBAL-H=diisobutylaluminum hydride
[0187] DIPEA=N,N-diisopropylethylamine
[0188] DMA=dimethylacetamide
[0189] DME=1,2-dimethoxyethane
[0190] DMF=dimethylformamide
[0191] DMSO=dimethylsulfoxide
[0192] DPPA=Diphenylphosphoryl azide
[0193] DTT=Dithiothreitol
[0194] EDC=1-(3-dimethylaminopropyl)-3-ethylcarbodiimide
hydrochloride
[0195] EDTA=ethylenediamine tetracetic acid
[0196] Et=ethyl
[0197] Et.sub.2O=diethyl ether
[0198] EtOAc=ethyl acetate
[0199] EtOH=ethanol
[0200] GST=gluetithione S-transferase
[0201] h=hours
[0202] Hexafluorophosphate
[0203] HOAt=1-hydroxy-7-azabenzotriazole
[0204] HOBt=1-hydroxybenzotriazole
[0205] Hunig's Base=N,N-diisopropylethylamine
[0206] KOtBu=potassium tert-butoxide
[0207] LC=liquid chromatography
[0208] LDA=lithium diisopropylamide
[0209] MBP=Myelin basic protein
[0210] mCPBA=m-chloroperoxybenzoic acid
[0211] Me=methyl
[0212] MeI=methyl iodide
[0213] MeOH=methanol
[0214] MS(ES)=Electro-Spray Mass Spectrometry
[0215] n-BuLi=n-butyllithium
[0216] Pd/C=palladium on activated charcoal
[0217] Ph=phenyl
[0218] PhCH.sub.3=toluene
[0219] pTSA=para-toluenesulfonic acid
[0220] RT=retention time
[0221] rt=room temperature
[0222] sat.=saturated
[0223] t-Bu=tert-butyl
[0224] TCA=trichloroacetic acid
[0225] TEA=triethylamine
[0226] TFA=trifluoroacetic acid
[0227] THF=tetrahydrofuran
[0228] TLC=thin layer chromatography
[0229] Tris-HCl=Tris[hydroxymethyl]aminomethane hydrochloride
[0230] Ts=tosyl
[0231] TsCl=tosyl chloride
[0232] TsOH=tosic acid
[0233] The following examples are provided to describe the
invention in further detail. These examples are intended to
illustrate and not to limit the invention. All temperatures are
given in centigrade degrees (.degree. C.) unless otherwise noted.
The YMC Co., Ltd., a supplier of HPLC columns, is located in Kyoto,
Japan, and may be reached through the Waters Co. in Milford,
Mass.
EXAMPLE 1
Preparation of
3-Cyano-4-(cyclohexylamino)-5-methylpyrrolo[1,2-b]pyridazin-
e-6-carboxylic acid ethyl ester (1E)
[0234] 24
[0235] A. Preparation of 3-Methyl-1H-pyrrole-2,4-dicarboxylic acid
diethyl ester (1A) 25
[0236] To a solution of ethyl isocyanoacetate (38.1 mL, 0.34 mol)
and DBU (50.8 mL, 0.34 mol) in THF (400 mL) at 50.degree. C. was
added a solution of acetaldehyde (9.5 mL, 0.17 mol) in THF (100 mL)
over 25 min. The reaction mixture was stirred at 55.degree. C. for
17 h, cooled to 25.degree. C. and acetic acid (20 mL) was slowly
added. The resulting mixture was concentrated in vacuo and the
resulting residue was dissolved in ethyl acetate (800 mL) and
washed with HCl (1 N, 3.times.300 mL). The combined aqueous washes
were extracted with ethyl acetate (3.times.200 mL) and the combined
organic layers were washed with NaHCO.sub.3 (sat. aq., 3.times.200
mL), water (100 mL) and brine (100 mL) and then concentrated in
vacuo to afford a dark brown oil. Elution of this oil through a
silica pad using ethyl acetate/hexanes (1:1) and the concentration
in vacuo provided compound 1A (16 g, 42% yield) as a yellow solid.
HPLC: 100% at 3.536 min (retention time) (YMC S5 ODS column,
4.6.times.50 mm, eluting with 10-90% aqueous methanol over 4 min
containing 0.2% phosphoric acid, 4 mL/min, monitoring at 220 nm).
MS (ES): m/z 226.0 [M+H].sup.+.
[0237] B. Preparation of
1-Amino-3-methyl-1H-pyrrole-2,4-dicarboxylic acid diethyl ester
(1B) 26
[0238] To a suspension of NaH (60% suspension in mineral oil, 213
mg, 5.33 mmol) in DMF (15 mL) at 0.degree. C. was added compound 1A
(1.0 g, 4.44 mmol), portionwise. The reaction mixture was stirred
at 0.degree. C. for 5 min and then warmed to 25.degree. C. and
stirred for an additional 1 h. The reaction mixture was then cooled
to 10.degree. C. and 2,4-dinitro-aminophenol (972 mg, 4.88 mmol)
was added in two portions. The resulting mixture was warmed to
25.degree. C., stirred for 12 h, poured onto water (40 mL) and
dichloromethane (50 mL), and the layers were separated. The aqueous
phase was extracted with dichloromethane (3.times.20 mL), and the
combined organic extracts were washed with NaOH (1N, 3.times.20
mL), water (20 mL), and brine (20 mL) and then dried over
MgSO.sub.4, filtered, and concentrated in vacuo. The resulting
reddish-brown residue was further concentrated for 12 h under high
vacuum to yield 800 mg (75% yield) of compound 2B which was used
without further purification. HPLC: 100% at 3.488 min (retention
time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with 10-90%
aqueous methanol over 4 min containing 0.2% phosphoric acid, 4
mL/min, monitoring at 220 nm). MS (ES): m/z 241.17 [M+H].sup.+.
[0239] C. Preparation of
3-Cyano-1,4-dihydro-5-methyl-4-oxopyrrolo[1,2-b]p-
yridazine-6-carboxylic acid ethyl ester (1C) 27
[0240] To a solution of
1-amino-3-methyl-1H-pyrrole-2,4-dicarboxylic acid diethyl ester
(1.08 g, 4.50 mmol) in toluene (15 mL) were added
1,1-diethoxypropionitrile (2.02 mL, 1.93 g, 13.5 mmol) and
TsOH-H.sub.2O (171 mg, 0.90 mmol). The reaction mixture was heated
at reflux for 12 h and then cooled to 25.degree. C. DBU (0.81 mL,
0.822 g, 5.40 mmol) was added and the resulting dark brown mixture
was heated at 80.degree. C. for 1 h and then cooled to room
temperature. The reaction mixture was poured onto dichloromethane
(50 mL) and NH.sub.4Cl (sat. aq., 50 mL) and the layers were
separated. The aqueous layer was extracted with dichloromethane
(2.times.30 mL) and the combined organic extracts were washed with
water (30 mL), dried over MgSO.sub.4, filtered, and concentrated in
vacuo. The crude product was purified by silica gel column
chromatography (10-30% methanol/dichloromethane) to provided 441 mg
(40%) of compound 1C as a brown solid. HPLC: 100% at 3.383 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.2% phosphoric acid,
4 mL/min, monitoring at 220 nm). MS (ES): m/z 246.09
[M+H].sup.+.
[0241] D. Preparation of
4-Chloro-3-cyano-5-methylpyrrolo[1,2-b]pyridazine- -6-carboxylic
acid ethyl ester (1D) 28
[0242] A 15 mL round bottom flask containing the compound 1C (370
mg, 1.51 mmol) was charged with POCl.sub.3 (1 mL) and heated to
75.degree. C. for 2 h. The reaction mixture was concentrated in
vacuo and the resulting yellow residue was dissolved in
dichloromethane (10 mL) and added, via pipette, to a saturated
aqueous solution of NaHCO.sub.3 with stirring at 0.degree. C. The
heterogeneous mixture was stirred for 10 min at 0.degree. C. then
warmed to room temperature and stirred for an additional 1 h. The
mixture was poured into a separatory funnel and the layers were
separated. The aqueous phase was extracted with dichloromethane
(2.times.20 mL) and the combined organic extracts were washed with
NaHCO.sub.3 (sat. aq., 1.times.20 mL), dried over Na.sub.2SO.sub.4,
filtered and concentrated in vacuo. The resulting residue was
dissolved in EtOAc (20 mL) and filtered through a pad of silica
using EtOAc (100 mL) to wash the silica pad. The filtrate was
concentrated in vacuo to afford compound 1D as a yellow solid which
was used without further purification. HPLC: 100% at 4.160 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.2% phosphoric acid,
4 mUmin, monitoring at 220 nm).
[0243] E. Preparation of
3-Cyano-4-(cyclohexylamino)-5-methylpyrrolo[1,2-b-
]pyridazine-6-carboxylic acid ethyl ester (1E)
[0244] To a solution of compound 1D (20 mg, 0.076 mmol) in
acetonitrile (1 mL) were added Et.sub.3N (32 .mu.L, 0.228 mmol) and
cyclohexylamine (10 .mu.L, 0.084 mmol) and the reaction mixture was
stirred at 25.degree. C. After 24 h, an additional 10 .mu.L of
cyclohexylamine was added and the reaction mixture was stirred for
an additional 1.5 h after which time it was poured onto NaHCO.sub.3
(sat. aq., 20 mL) and dichloromethane. The layers were separated
and the aqueous layer was extracted with dichloromethane
(2.times.10 mL). The combined organic layers were washed with water
(20 mL), dried over MgSO.sub.4, filtered, and concentrated in vacuo
to yield 18 mg (75%) of compound 1E as a yellow solid, which was
used without further purification. HPLC: 100% at 4.60 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.2% phosphoric acid,
4 mL/min, monitoring at 220 nm). MS (ES): m/z 327.2
[M+H].sup.+.
EXAMPLE 2
Preparation of 3-Cyano-5-methyl-4-phenoxypyrrolo
[1,2-b]pyridazine-6-carbo- xylic acid ethyl ester
[0245] 29
[0246] To a solution of compound 1D (18 mg, 0.068 mmol) in
acetonitrile (0.5 mL) at room temperature were added Et.sub.3N (21
uL, 0.205 mmol) and phenol (7 mg, 0.075 mmol). The reaction mixture
was stirred for 24 h and then poured onto dichloromethane (10 mL)
and NaHCO.sub.3 (sat. aq., 10 mL). The layers were separated, the
aqueous phase was extracted with dichloromethane (3.times.5 mL),
and the combined organic extracts were washed with water, dried
over MgSO.sub.4, filtered, and concentrated in vacuo to afford
compound 2 (15 mg, 68%) as a yellow solid. HPLC: 100% at 4.35 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.2% phosphoric acid,
4 mL/min, monitoring at 220 nm). MS (ES): m/z 340.0
[M+NH.sub.4].sup.+.
EXAMPLE 3
Preparation of
6-(Methoxymethyl)-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrol-
o[1,2-b]pyridazine-3-carbonitrile (3C)
[0247] 30
[0248] A. Preparation of
3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrol-
o[1,2-b]pyridazine-6-carboxylic acid ethyl ester (3A) 31
[0249] To a solution of compound 1D (26 mg, 0.10 mmol) in DMF (2
mL) were added K.sub.2CO.sub.3 (138 mg, 1.00 mmol) and
p-phenoxyaniline (20 mg, 0.11 mmol) at 25.degree. C. The reaction
mixture was stirred for 12 h and then diluted with dichloromethane
(15 mL) and washed with water (10 mL) and brine (10 mL). The
organic phase was dried over Na.sub.2SO.sub.4 and concentrated and
the resulting residue was triturated with methanol to afford 31 mg
(76% yield) of the desired compound as a yellow solid. HPLC: 100%
at 4.62 min (retention time) (YMC S5 ODS column, 4.6.times.50 mm,
eluting with 10-90% aqueous methanol over 4 min containing 0.2%
phosphoric acid, 4 mL/min, monitoring at 220 nm). MS (ES): m/z
413.12 [M+H].sup.+.
[0250] Compound 3A can also be prepared as follows: To a solution
of ID (1.00 g, 3.79 mmol) in THF (10 mL) were added
4-phenoxyaniline (0.84 g, 4.53 mmol) and triethylamine (1.06 mL,
7.58 mmol). The reaction mixture was heated at 60.degree. C. for 3
days, after which time it was cooled to room temperature and
diluted with MeOH (50 mL). The resulting solids were filtered,
washed with MeOH and dried to yield 1.50 g (96% yield) of 3A as a
yellow powder.
[0251] B. Preparation of
6-(Hydroxymethyl)-5-methyl-4-[(4-phenoxyphenyl)am- ino]
pyrrolo[1,2-b]pyridazine-3-carbonitrile (3B) 32
[0252] To a solution of compound 3A (41 mg, 0.10 mmol) in THF (2
mL) at -78.degree. C. was added DIBAL-H (1.5 M in toluene, 0.13 mL,
0.20 mmol). The reaction mixture was stirred for 6 h at -78.degree.
C, warmed to 0.degree. C. and stirred for an additional 2 h. The
reaction mixture was quenched by the addition of methanol (3 mL)
and sat. aq. Na.sub.2CO.sub.3 (3 mL) and then poured onto
dichloromethane (20 mL). The layers were separated, the aqueous
phase was extracted with dichloromethane (2.times.15 mL), and the
combined organic extracts were dried over MgSO.sub.4, filtered, and
concentrated in vacuo. Purification via preparative HPLC (YMC S5
ODS 20.times.100 mm, eluting with 30-100% aqueous methanol over 15
min containing 0.1% TFA, 20 mL/min) afforded 33 mg (90% yield) of
compound 3B as a yellow solid. HPLC: 100% at 3.98 min (retention
time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with 10-90%
aqueous methanol over 4 min containing 0.2% phosphoric acid, 4
mL/min, monitoring at 220 nm). MS (ES): m/z 371.19 [M+H].sup.+.
[0253] C. Preparation of
6-(Methoxymethyl)-5-methyl-4-[(4-phenoxyphenyl)am- ino]
pyrrolo[1,2-b]pyridazine-3-carbonitrile (3C)
[0254] To a solution of compound 3B (9.0 mg, 0.025 mmol) in DMF:THF
(1:1, 1 mL) at 0.degree. C. was added KOtBu (1.5 M in THF, 0.025
mL, 0.038 mmol). After stirring for 45 min at 0.degree. C., methyl
iodide (2 .mu.L, 0.025 mmol) was added and the reaction mixture was
stirred for an additional 1 h , warmed to 25.degree. C., and
stirred for 3 h. No reaction was observed during this time. The
reaction mixture was cooled once more to 0.degree. C., additional
KOtBu (1.5 M in THF, 0.25 mL, 0.38 mmol) was added and the reaction
mixture was stirred for 30 min, after which time additional methyl
iodide (20 .mu.L, 0.25 mmol) was added. After stirring for an
additional 2 h at 0.degree. C., the reaction was quenched by the
addition of saturated aqueous NH.sub.4Cl (10 mL) and
dichloromethane (10 mL). The layers were separated, the aqueous
phase was extracted with dichloromethane (2.times.10 mL) and the
combined organic extracts were dried over MgSO.sub.4, filtered,
concentrated, and purified by preparative HPLC (YMC S5 ODS
20.times.100 mm, eluting with 30-100% aqueous methanol over 15 min
containing 0.1% TFA, 20 mL/min) to afford the desired product as a
yellow semi-solid. HPLC: 100% at 4.27 min (retention time) (YMC S5
ODS column, 4.6.times.50 mm, eluting with 10-90% aqueous methanol
over 4 min containing 0.2% phosphoric acid, 4 mL/min, monitoring at
220 nm). MS (ES): m/z 385.21 [M+H].sup.+.
EXAMPLE 4
Preparation of
4-[(2-Chloro-4-iodophenyl)amino]-3-cyano-5-methylpyrrolo[1,-
2-b]pyridazine-6-carboxylic acid
[0255] 33
[0256] To a solution of
4-[(2-chloro-4-iodophenyl)amino]-3-cyano-5-methylp-
yrrolo[1,2-b]pyridazine-6-carboxylic acid, ethyl ester (59 mg,
0.123 mmol, prepared as described in Example 1) in THF (1 mL) was
added NaOH (1N, 1 mL). The reaction mixture was stirred at
25.degree. C. for 72 h and then poured onto NaHCO.sub.3 (30 mL) and
EtOAc (30 mL). The layers were separated and the aqueous phase was
acidified to pH=2 and extracted with dichloromethane (2.times.20
mL). The combined organic extracts were washed with brine, dried
over MgSO.sub.4, filtered and concentrated in vacuo to afford 20 mg
(36% yield) of compound 4 which was used without further
purification. .sup.1H NMR (DMSO-d.sub.6) .delta. 8.99 (s, 1H), 8.18
(s, 1H), 8.03 (s, 1H), 7.97 (s, 1H), 7.75 (d, 1H), 7.29 (d, 1H),
2.78 (s, 3H). HPLC: 100% at 4.04 min (retention time) (YMC S5 ODS
column, 4.6.times.50 mm, eluting with 10-90% aqueous methanol over
4 min containing 0.2% phosphoric acid, 4 mL/min, monitoring at 220
nm).
EXAMPLE 5
Preparation of
4-[(2-Chloro-4-iodophenyl)amino]-3-cyano-5-methyl-N-(2-meth-
ylpropoxy)pyrrolo[1,2-b]pyridazine-6-carboxamide
[0257] 34
[0258] To a solution of compound 4 (20 mg, 0.442 mmol) in
THF:dichloromethane (1:1, 1 mL) were added isopropylhydroxylamine
HCl (7 mg, 0.053 mmol), Hunig's base (18 .mu.L, 0.106 mmol) and
PyBOP (benzotriazol-1-yl oxytripyrrolidinophosphonium
hexafluorophosphate) (28 mg, 0.0531 mmol). The reaction mixture was
stirred for 1 h , concentrated in vacuo and diluted with 10% HCl
(15 mL) and Et.sub.2O (15 mL). The layers were separated and the
organic phase was washed with 1N NaOH (15 mL) and brine (15 mL),
dried over MgSO.sub.4, filtered and concentrated in vacuo. The
aqueous phase was made basic by the addition of 1N NaOH and
extracted with EtOAc (2.times.15 mL). The organic extracts were
dried over MgSO.sub.4, filtered, concentrated in vacuo, and added
to the combined organic layers of the first extractions.
Preparative HPLC (YMC S5 ODS 20.times.100 mm, eluting with 30-100%
aqueous methanol over 15 min containing 0.1% TFA, 20 mL/min)
provided 1.1 mg of the compound 5. HPLC: 100% at 3.68 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.1% TFA, 4 mL/min,
monitoring at 220 nm). MS (ES): m/z 524.02 [M+H].sup.+.
EXAMPLE 6
Preparation of
3-Cyano-4-[[5-[(methoxyamino)carbonyl]-2-methylphenyl]amino-
]-5-methylpyrrolo[1,2-b]pyridazine-6-carboxylic acid
[0259] 35
[0260] To a solution of
3-cyano-4-[[5-[(methoxyamino)carbonyl]-2-methylphe-
nyl]amino]-5-methylpyrrolo[1,2-b]pyridazine-6-carboxylic acid,
ethyl ester (160 mg, 0.393 mmol, prepared as described in Example
1) in THF (2 mL) was added 1N NaOH (4 mL). The reaction mixture was
stirred at 25.degree. C. for 2 days and then neutralized with 1M
citric acid and poured onto dichloromethane. The organic phase was
separated and extracted with dichloromethane, and the combined
organic extracts were dried over Na.sub.2SO.sub.4, filtered, and
concentrated in vacuo. The resulting residue was purified via
preparative HPLC (YMC S5 ODS 20.times.100 mm, eluting with 30-100%
aqueous methanol over 15 min containing 0.1% TFA, 20 mL/min) to
afford 36 mg (39% yield) of compound 6. HPLC: 100% at 3.33 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm, eluting with
10-90% aqueous methanol over 4 min containing 0.2% phosphoric acid,
4 mL/min, monitoring at 220 nm). MS (ES): m/z 380.24
[M+H].sup.+.
EXAMPLE 7
Preparation of
3-Cyano-4-[[5-[(methoxyamino)carbonyl]-2-methylphenyl]amino-
]-5-methyl-N-[(1S)-1-phenylethyl]pyrrolo[1,2-b]pyridazine-6-carboxamide
[0261] 36
[0262] To a solution of compound 6 (19 mg, 0.05 mmol) in DMF (2 mL)
were added EDC (14.4 mg, 0.075 mmol), HOBt (10.1 mg, 0.075 mmol)
and DIPEA (12.9 mg, 0.10 mmol) and the reaction mixture was stirred
for 30 min at 25.degree. C. (S)-Methylbenzylamine (7.3 mg, 0.06
mmol) was then added and the reaction mixture was stirred for an
additional 16 h at 25.degree. C. The reaction was then diluted with
dichloromethane and poured into water. The layers were separated
and the organic phase was dried over Na.sub.2SO.sub.4, filtered and
concentrated in vacuo. The residue was purified via preparative
HPLC (YMC S5 ODS 20.times.100 mm, eluting with 30-100% aqueous
methanol over 15 min containing 0.1% TFA, 20 mL/min) to afford 21
mg (87% yield) of compound 7 as a yellow semi-solid. HPLC: 100% at
3.79 min (retention time) (YMC S5 ODS column, 4.6.times.50 mm,
eluting with 10-90% aqueous methanol over 4 min containing 0.2%
phosphoric acid, 4 mI/min, monitoring at 220 nm). MS (ES): m/z
483.36 [M+H].sup.+.
EXAMPLE 8
Preparation of
6-Formyl-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[1,2-b]p-
yridazine-3-carbonitrile
[0263] 37
[0264] To a solution of compound 3B (266 mg, 0.72 mmol) in
dichloroethane (30 mL) was added MnO.sub.2 (200 mg, 2.0 mmol). The
reaction mixture was heated at 60.degree. C. for 3 h, cooled to
25.degree. C, diluted with dichloromethane and filtered through
Celite. The filtrate was concentrated in vacuo and purified by
column chromatography on silica gel (0.5% MeOH/CH.sub.2Cl.sub.2) to
afford 238 mg (90% yield) of compound 8 as a yellow solid. HPLC:
100% at 3.37 min (retention time) (YMC S5 ODS column, 4.6.times.50
mm, eluting with 10-90% aqueous methanol over 4 min containing 0.2%
phosphoric acid, 4 mL/min, monitoring at 220 nm). MS (ES): m/z
369.08 [M+H].sup.+.
EXAMPLE 9
Preparation of
3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[1,2-b]py-
ridazine-6-carboxylic acid
[0265] 38
[0266] To a solution of compound 3A (1.50 g, 3.64 mmol) in THF (50
mL) were added NaOH (1 M, 20.0 mL) and EtOH (25 mL). The reaction
was heated at 80.degree. C., effectively evaporating the THF, and,
after 1 h , the reaction mixture became homogeneous. Heating was
continued for an additional 6 h, after which time the reaction was
cooled to rt and neutralized with HCl (1 M, 20.0 mL). The resulting
solids were filtered, washed with water and dried to afford
compound 9 (1.33 g, 95% yield) as a yellow solid. HPLC: 100% at
1.84 min (retention time) (YMC S5 ODS column, 4.6.times.50 mm
eluting with 10-90% MeOH/H.sub.2O over 2 minutes containing 0.1%
TFA; 4 mL/min, monitoring at 220 nm). MS (ES): m/z 489.0
[M+H].sup.+.
EXAMPLE 10
Preparation of an Amide Library
Preparation of
3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]-N-phenylpyrrolo-
[1,2-b]pyridazine-6-carboxamide
[0267] 39
[0268] To a solution of compound 9 (11.5 mg, 0.030 mmol) and HOAt
(6.1 mg, 0.045 mmol) in THF (0.60 mL) was added a solution of
aniline (14 mg, 0.15 mmol) in THF (0.15 mL) followed by a solution
of EDC (11.5 mg, 0.06 mmol) in chloroform (0.30 mL). The reaction
mixture was heated at 60.degree. C. overnight and then cooled to rt
and diluted with MeOH (0.4 mL). The resulting mixture was purified
by elution through a SCX/SAX cartridge (500 mg/500 mg) SCX SAX
silica bound ionexchange cartridges, supplied by United Chemical
Technologies, Inc., Bristol, Pa., with MeOH, followed by
concentration of the solvent in vacuo, to afford 13.2 mg (96%
yield) of compound 10 as a yellow solid. HPLC: 100% at 2.05 min
(retention time) (YMC S5 ODS column 4.6.times.50 mm eluting with
10-90% MeOH/H.sub.2O over 4 minutes containing 0.2% phosphoric
acid, 4 mL/min, monitoring at 220 nm). MS (ES): m/z 460.0
[M+H].sup.+.
[0269] The above procedure was utilizeded to prepare a library of
68 amide compounds by substituting other amines for the aniline
reactant. Compounds were purified using the above method or by
preparative HPLC (Shimadzu VP-ODS 20.0.times.50.0 mm eluting with
25-90% MeOH/H.sub.2O over 7 minutes containing 0.1% TFA, 10 mL/min,
monitoring at 220 nm).
EXAMPLE 11
Preparation of
6-Amino-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[1,2-b]py-
ridazine-3-carbonitrile (11B)
[0270] 40
[0271] A. Preparation of
[3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]pyrro-
lo[1,2-b]pyridazin-6-yl]carbamic acid, phenylmethyl ester (11A)
41
[0272] To a solution of compound 9 (192 mg, 0.50 mmol) in dioxane
(anhydrous, 4 mL) under an N.sub.2 atmosphere were added
triethylamine (0.140 mL, 1.00 mmol) and DPPA (0.216 mL, 1.00 mmol),
and the mixture was stirred overnight. Benzyl alcohol (0.310 mL,
3.00 mmol) was then added and the reaction mixture was heated at
75.degree. C. for 4 h, concentrated in vacuo and purified by column
chromatography on silica gel. (20 to 30% EtOAc/hexanes) to yield
compound 11A as a yellow oil (172 mg, 70%). HPLC: 100% at 4.01 min
(retention time) (YMC S5 ODS column, 4.6.times.50 mm eluting with
10-90% MeOH/H.sub.2O over 4 minutes containing 0.1% TFA; 4 mL/min,
monitoring at 220 nm). MS (ES): m/z 490.0 [M+H].sup.+.
[0273] B. Preparation of
6-Amino-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrol-
o[1,2-b]pyridazine-3-carbonitrile (11B)
[0274] To a solution of compound 11A (40 mg, 0.082 mmol) in MeOH (4
mL) was added Pd/C (12 mg), and the reaction mixture was stirred
under hydrogen (1 atm) for 30 minutes, after which time HCl (4M in
dioxane, 0.1 mL) was added. The reaction mixture was filtered and
the filtrate concentrated in vacuo to provide 32 mg of compound 11B
as an orange solid (quantitative yield as HCl salt). HPLC: 100% at
2.90 min (retention time) (YMC S5 ODS column, 4.6.times.50 mm
eluting with 10-90% MeOH/H.sub.2O over 4 minutes containing 0.1%
TFA; 4 mL/min, monitoring at 220 nm). MS (ES): m/z 356.0
[M+H].sup.+.
EXAMPLE 12
Preparation of
N-[3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[1,2-b-
]pyridazin-6-yl]acetamide
[0275] 42
[0276] To a solution of compound 11B (HCl salt, 32 mg, 0.082 mmol)
in THF (2 mL) was added acetic anhydride (11 mg, 0.11 mmol)
followed by triethylamine (33 mg, 0.33 mmol) and the reaction was
stirred at rt for 30 min. After quenching with MeOH, the reaction
mixture was stirred for an additional 30 min, concentrated in vacuo
and purified by flash chromatography on silica gel (40 to 50%
EtOAc/dichloromethane) to furnish compound 12 as a yellow oil (30
mg, 92% yield). HPLC: 100% at 3.46 min (retention time) (YMC S5 ODS
column, 4.6.times.50 mm eluting with 10-90% MeOH/H.sub.2O over 2
minutes containing 0.1% TFA; 4 mL/min, monitoring at 220 nm). MS
(ES): m/z 398.0 [M+H].sup.+.
EXAMPLE 13
Preparation of
3-(Aminomethyl)-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[-
1,2-b]pyridazine-6-carboxylic acid
[0277] 43
[0278] To a solution of compound 9 (27 mg, 0.070 mmol) in MeOH:THF
(2:1 v/v, 6 ml) was added TFA (30 mg) followed by Pd/C (10 mg). The
reaction mixture was stirred under hydrogen (1 atm) overnight and
then filtered through a pad of Celite. The filtrate was
concentrated in vacuo and the residue was purified by several
azeotropic distillations with MeOH to remove excess TFA. This
procedure afforded compound 13 as a yellow solid (35 mg,
quantitative). HPLC: 100% at 2.96 min (retention time) (YMC S5 ODS
column, 4.6.times.50 mm eluting with 10-90% MeOH/H.sub.2O over 4
minutes containing 0.1% TFA; 4 mL/min, monitoring at 220 nm). MS
(ES): m/z 389.0 [M+H].sup.+.
EXAMPLE 14
Preparation of
3-[(Acetylamino)methyl]-5-methyl-4-[(4-phenoxyphenyl)amino]-
pyrrolo[1,2-b]pyridazine-6-carboxylic acid
[0279] 44
[0280] To a solution of compound 13 (10 mg, 0.02 mmol) in THF was
added triethylamine (1 drop, 10 mg) followed by acetic anhydride (1
drop, 10 mg). The reaction was stirred at rt for 10 min and then
concentrated in vacuo. The residue was redissolved in THF, and NaOH
(1M, 2 drops) was added. The resulting mixture was stirred at rt
for 2 hours, neutralized with HCl (1M), and purified by preparative
HPLC (Shimadzu VP-ODS 20.0.times.50.0 mm eluting with 25-90%
MeOH/H.sub.2O over 7 minutes containing 0.1% TFA, 10 mL/min,
monitoring at 220 nm) to give compound 14 as a yellow solid (7 mg,
81% yield). HPLC: 100% at 3.46 min (retention time) (YMC S5 ODS
column, 4.6.times.50 mm eluting with 10-90% MeOH/H.sub.2O over 4
minutes containing 0.1% TFA; 4 mL/min, monitoring at 220 nm). MS
(ES): m/z 431.0 [M+H].sup.+.
EXAMPLE 15
Preparation of
N-[3-Cyano-5-methyl-4-[(4-phenoxyphenyl)amino]pyrrolo[1,2-b-
]pyridazin-6-yl]-N'-methylurea
[0281] 45
[0282] A solution of compound 9 (19 mg, 0.05 mmol), triethylamine
(0.014 mL, 0.10 mmol) and DPPA (0.022 mL, 0.10 mmol) in dry dioxane
(1 mL) was stirred under N.sub.2 for 12 h. The reaction was then
heated to 80.degree. C. for 1 h and then allowed to cool to
25.degree. C. on standing. Methylamine (2.0M THF solution, 0.30 mL,
0.60 mmol) was added and the reaction was stirred at 25.degree. C.
for 1 h , concentrated in vacuo and purified by flash
chromatography on a silica gel column (50 to 70%
EtOAc/dichloromethane) to give compound 15 (15 mg, 73%) as a yellow
solid. HPLC: 100% at 3.44 min (retention time) (YMC S5 ODS column,
4.6.times.50 mm eluting with 10-90% MeOH/H.sub.2O over 4 minutes
containing 0.1% TFA; 4 mL/min, monitoring at 220 nm). MS (ES): m/z
413.0 [M+H].sup.+.
EXAMPLE 16
Preparation of
3-Cyano-5-hydroxymethyl-4-(4-phenoxy-phenylamiino)-pyrrolo[-
1,2-b]pyridazine-6-carboxylic acid ethyl ester (16C)
[0283] 46
[0284] A.
5-Bromomethyl-4-chloro-3-cyano-pyrrolo[1,2-b]pyridazine-6-carbox-
ylic acid ethyl ester (16A) 47
[0285] A suspension of compound 1D (79 mg, 0.30 mmol), NBS (59 mg,
0.33 mmol) and benzoyl peroxide (5 mg, 0.02 mmol) in CCl.sub.4 (2
mL) was heated at 77.degree. C. for 3 hours. After cooling to room
temperature, the reaction was purified by a short silica gel column
(eluted with CH.sub.2Cl.sub.2) to give compound 16A as a yellow
solid (102 mg, 99%).
[0286] B.
4-Chloro-3-cyano-5-hydroxymethyl-pyrrolo[1,2-b]pyridazine-6-carb-
oxylic acid ethyl ester (16B) 48
[0287] To a solution of compound 16A (102 mg, 0.30 mmol) in THF (12
mL) was added water (3 mL) dropwise. The reaction was kept at room
temperature for 3 days and then heated to 50.degree. C. for 3
hours. Upon cooling to room temperature, NaHCO.sub.3 (70 mg) was
added to the reaction mixture. The reaction was concentrated to
dryness, redissolved in CH.sub.2Cl.sub.2 and filtered. The filtrate
was concentrated to give compound 16B as a yellow solid (84 mg,
100%). This compound was used in the following steps without
further purification.
[0288] C.
3-Cyano-5-hydroxymethyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]-
pyridazine-6-carboxylic acid ethyl ester (16C)
[0289] A solution of compound 16B (84 mg, 0.30 mmol),
4-phenoxyaniline (72 mg, 0.39 mmol) and triethylamine (0.083 mL,
0.60 mmol) in THF (2.5 mL) was heated at 70.degree. C. for 30 min.
After cooled to room temperature, the reaction was concentrated to
about 0.5 mL, diluted with MeOH (2 mL) and filtered. The solid was
washed with MeOH and dried to give compound 16C as a yellow solid
(118 mg, 92%). HPLC: 92% at 2.12 min (retention time) (PrimeSphere
5u C18-HC column, 4.6.times.30 mm eluting with 10-90% MeOH/H.sub.2O
over 2 minutes containing 0.1% TFA; 5 mL/min, monitoring at 220
nm). MS (ES): m/z 429.0 [M+H].sup.+.
EXAMPLE 17
Preparation of
3-Cyano-5-methoxymethyl-4-(4-phenoxy-phenylamino)-pyrrolo[1-
,2-b]pyridazine-6-carboxylic acid ethyl ester (17B)
[0290] 49
[0291] A.
4-Chloro-3-cyano-5-methoxymethyl-pyrrolo[1,2-b]pyridazine-6-carb-
oxylic acid ethyl ester (17A) 50
[0292] To a solution of compound 16A (31 mg, 0.09 mmol) in 1:1
MeOH:CH.sub.2Cl.sub.2 (2 mL) was added NaHCO.sub.3 (30 mg, 0.36
mmol). The reaction was kept at room temperature for 3 h, heated to
70.degree. C. for 1 h, cooled to room temperature, concentrated to
dryness, redissolved in CH.sub.2Cl.sub.2 and filtered. The filtrate
was concentrated to give compound 17A as a yellow solid (26 mg,
98%).
[0293] B.
3-Cyano-5-methoxymethyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]-
pyridazine-6-carboxylic acid ethyl ester (17B)
[0294] Compound 17B was made in acordance with the procedure
described in Example 16C. HPLC: 96% at 2.24 min (retention time)
(PrimeSphere 5u C18-HC column, 4.6.times.30 mm eluting with 10-90%
MeOH/H.sub.2O over 2 minutes containing 0.1% TFA; 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 443.0 [M+H].sup.+.
EXAMPLE 18
Preparation of
3-Cyano-5-formyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]py-
ridazine-6-carboxylic acid ethyl ester (18)
[0295] 51
[0296] To a solution of compound 16C (21.4 mg, 0.05 mmol) in
chloroform (1 mL) was added MnO.sub.2 (<5 micron, activated, 17
mg, 0.20 mmol). The reaction was heated at 55.degree. C. overnight,
cooled to room temperature and purified by flash chromatography on
a silica gel column (0-2% EtOAc/CH.sub.2Cl.sub.2) to give compound
18 as a yellow solid (20 mg, 94%). HPLC: 94% at 2.20 min (retention
time) (PrimeSphere 5u C18-HC column, 4.6.times.30 mm eluting with
10-90% MeOH/H.sub.2O over 2 minutes containing 0.1% TFA; 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 427.0 [M+H].sup.+.
EXAMPLE 19
Preparation of
1-[3-Cyano-5-methyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b-
]pyridazin-6-yl]-3-(2-morpholin-4-yl-ethyl)-urea (19-1) &
5-Methyl-6-(5-oxo-4,5-dihydro-tetrazol-1-yl)-4-(4-phenoxy-phenylamino)-py-
rrolo[1,2-b]pyridazine-3-carbonitrile (19-2)
[0297] 52
[0298] A solution of compound 9 (115 mg, 0.30 mmol), triethylamine
(0.063 mL, 0.45 mmol) and DPPA (0.097 mL, 0.45 mmol) in dioxane (5
mL) was stirred overnight. The next day TMS-azide (0.080 mL, 0.60
mmol) was added and the reaction temperature was brought to
80.degree. C. The reaction was heated at 80.degree. C. for 2 hours,
cooled to room temperature and 4-(2-aminoethyl)morpholine (0.079
mL, 0.60 mmol) was added. The reaction was stirred at room
temperature for 1 h, concentrated and purified by flash
chromatography on a silica gel column (3-6% MeOH/CH.sub.2Cl.sub.2)
to give a mixture of compounds 19-1 and 19-2 as a yellow oil. This
oil was recrystalized from MeOH to give compound 19-1 as a yellow
solid (116 mg, 76%). 19-1: HPLC: 97% at 3.11 min (retention time)
(YMC S5 ODS column, 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 4 minutes containing 0.1% TFA; 4 mL/min,
monitoring at 220 nm). MS (ES): m/z 512.0 [M+H].sup.+.
[0299] The mother liquor from the above recrystalization was passed
through a SCX cartridge (500 mg) and eluted with MeOH (5 mL). The
elutant was concentrated to give compound 19-2 as a yellow solid (9
mg, 7%). 19-2: HPLC: 94% at 1.93 min (retention time) (PrimeSphere
5u Cl 8-HC column, 4.6.times.30 mm eluting with 10-90%
MeOH/H.sub.2O over 2 minutes containing 0.1% TFA; 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 424.0 [M+H].sup.+.
EXAMPLES 20 TO 144
[0300] Further compounds of the present invention were prepared by
procedures analogous to those described above. Table 1 provides the
name and structure or representative compounds and their retention
times, as well as the Example number of the procedure on which the
preparation of the compound was based. The chromatography
techniques used to determine the retention times of the compounds
listed in Table 1 are as follows:
[0301] LCMS=YMC S5 ODS column, 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 4 minutes containing 0.1% TFA; 4 mL/min,
monitoring at 220 nm.
[0302] LCMS*=YMC S5 ODS column, 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 2 minutes containing 0.1% TFA; 4 mL/min,
monitoring at 220 nm.
[0303] LCMS-1=PrimeSphere 5u C18-HC column, 4.6.times.30 mm eluting
with 10-90% MeOH/H.sub.2O over 2 minutes containing 0.1% TFA; 5
mL/min, monitoring at 220 nm.
[0304] LC=YMC S5 ODS column 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 4 minutes containing 0.2% phosphoric acid, 4
mL/min, monitoring at 220 nm.
[0305] LC*=YMC S5 ODS column 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 4 minutes containing 0.2% phosphoric acid, 4
mL/min, monitoring at 220 nm.
[0306] The molecular mass of the compounds listed in Table 1 were
determined by MS (ES) by the formula m/z.
1TABLE 1 Retention Time Min./ Ex. Compound Molecular Proc. No.
Compound Structure Name Mass of Ex. 20 53 4-(6-Amino-1H-
indazol-1-yl)-3- cyano-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 3.42 LC [M + H].sup.+ =361.0 1 21 54
4-(6-Amino-1H- indazol-1-yl)-3- cyano-5,7- dimethylpyrrolo[1,2-
b]pyridazine-6- carboxylic acid, ethyl ester 3.59 LC [M + H].sup.+
=375.0 1 22 55 3-Cyano-4-(1H- imidazol-1-yl)-5- methylpyrrolo[1 2-
b]pyridazine-6- carboxylic acid, ethyl ester 2.79 LC [M + H].sup.+
=296.0 1 23 56 3-Cyano-4- (dimethylamino)- 5,7-
dimethylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
4.27 LC [M + H].sup.+ =287.0 1 24 57 3-Cyano-4-(1H-
indazol-6-ylamino)- 5,7- dimethylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 4.06 LC [M + H].sup.+ =375.0 1 25 58
3-Cyano-5-methyl- 4-[(2-methyl-1H- indol-5- yl)amino]pyrrolo[1,
2-b]pyridazine-6- carboxylic acid, ethyl ester 4.22 LC [M +
H].sup.+ =374.0 1 26 59 3-Cyano-5-methyl- 4- (phenylamino)
pyrrolo[1,2- b]pyridazine- 6-carboxylic acid, ethyl ester 4.07 LC
[M + H].sup.+ =321.0 1 27 60 3-Cyano-5-methyl- 4-[(2-methyl-1H-
indol-5- yl)oxy]pyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl
ester 3.80 LCMS [M + H].sup.+ =375.0 2 28 61 3-Cyano-5-methyl-
4-[[1- (phenylmethyl)-1H- indazol-5- yl]amino]pyrrolo[1,
2-b]pyridazine-6- carboxylic acid, ethyl ester 4.31 LC [M +
H].sup.+ =451.0 1 29 62 3-Cyano-5-methyl- 4-(1H-indazol-1-
yl)pyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester 4.04
LC [M + H].sup.+ =346.0 1 30 63 3-Cyano-4-[(4- fluoro-2-methyl-
1H-indol-5- yl)oxy]-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 3.86 LC [M + H].sup.+ =393.0 1 31 64
3-Cyano-4-[[2- fluoro-5- [(methoxyamino) carbonyl]phenyl]amino-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
4.23 LC [M + H].sup.+ =412.0 1 32 65 3-Cyano-4-[[5- [(methoxyamino)
carbonyl]-2- methylphenyl]amino]-5- methylpyrrolo[1,2-
b]pyridazine-6- carboxylic acid, ethyl ester 4.19 LC [M + H].sup.+
=408.0 1 33 66 4-[(4-Bromo-2- fluorophenyl)amino]- 3-cyano-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
4.32 LC 1 34 67 4-[(4-Chloro-2- iodophenyl)amino]- 3-cyano-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
4.37 LC [M + H].sup.+ =481.0 1 35 68 4-[(2-Chloro-4-
iodophenyl)amino]- 3-cyano-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 4.51 LC [M + Na.sup.+].sup.+ =503.0 36
69 3-Cyano-4-[[2- fluoro-5- [(methoxyamino)
carbonyl]phenyl]amino]-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 4.24 LC [M + H].sup.+ =412.0 1 37 70
3-Cyano-N-ethyl-5- methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.85 LC* [M + H].sup.+ =412.0 10 38 71
1-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[1,2-
b]pyridazin-6- yl]carbonyl]-4- methylpiperazine 1.53 LC* [M +
H].sup.+ =467.0 10 39 72 3-Cyano-5-methyl- 4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.75
LC* [M + H].sup.+ =384.0 10 40 73 3-Cyano-N,5- dimethyl-4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.79
LC* [M + H].sup.+ =398.0 10 41 74 3-Cyano-N,N,5- trimethyl-4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.78
LC* [M + H].sup.+ =412.0 10 42 75 N-Butyl-3-cyano-5- methyl-4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- - b]pyridazine-6- carboxamide 2.00
LC* [M + H].sup.+ =440.0 10 43 76 3-Cyano-5-methyl- 4-[(4-
phenoxyphenyl) amino]-N- (phenylmethyl) pyrrolo[1,2-
b]pyridazine-6- carboxamide 2.00 LC* [M + H].sup.+ =474.0 10 44 77
3-Cyano-N-[(4- methoxyphenyl) methyl]-5-methyl-4-
[(4-phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6- carboxamide
1.98 LC* [M + H].sup.+ =504.0 10 45 78 3-Cyano-N-[(3-
methoxyphenyl) methyl]-5-methyl-4- [(4-phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.01 LC* [M +
H].sup.+ =504.0 10 46 79 N-[(4-Chlorophenyl) methyl]-3-cyano-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 2.09 LC* [M + H].sup.+ =508.0 10 47 80
N-[(3-Chlorophenyl) methyl]-3-cyano-5- methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.08 LC* [M +
H].sup.+ =508.0 10 48 81 N-[(2-Chlorophenyl) methyl]-3-cyano-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 2.06 LC* [M + H].sup.+ =508.0 10 49 82
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-(2-
phenylethyl)pyrrolo [1,2-b]pyridazine-6- carboxamide 2.04 LC* [M +
H].sup.+ =488.0 10 50 83 3-Cyano-N-(2- furanylmethyl)-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.92 LC* [M + H].sup.+ =464.0 10 51 84
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-(2- thienylmethyl)
pyrrolo[1,2- b]pyridazine- 6-carboxamide 1.97 LC* [M + H].sup.+
=480.0 10 52 85 3-Cyano-N- cyclopropyl-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.86
LC* [M + H].sup.+ =424.0 10 53 86 3-Cyano-N- cyclopentyl-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.99 LC* [M + H].sup.+ =452.0 10 54 87 3-Cyano-N-
cyclohexyl-5- methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 2.07 LC* [M + H].sup.+ =466.0 10 55 88
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-[(tetrahydro-2-
furanyl)methyl]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.89 LC* [M
+ H].sup.+ =468.0 10 56 89 3-Cyano-N-(2- ethoxyethyl)-5-
methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.90 LC* [M + H].sup.+ =456.0 10 57 90
3-Cyano-5-methyl- N-(2-phenoxyethyl)-4- [(4-phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.05 LC* [M +
H].sup.+ =504.0 10 58 91 3-Cyano-N-(2,3- dihydroxypropyl)-
5-methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.69 LC* [M + H].sup.+ =458.0 10 59 92 3-Cyano-N-(6-
hydroxyhexyl)-5- methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[- 1,2-
b]pyridazine-6- carboxamide 1.87 LC* [M + H].sup.+ =484.0 10 60 93
N-[2-(Acetylamino) ethyl]-3-cyano-5- methyl-4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.74
LC* [M + H].sup.+ =469.0 10 61 94 3-Cyano-5-methyl- N-[3-(2-oxo-1-
prrolidinyl) propyl]-4-[4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.81 LC*[M + H].sup.+ =509.0 10 62 95
3-Cyano-N,N- diethyl-5-methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.90 LC* [M +
H].sup.+ =440.0 10 63 96 1-[[3-Cyano-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6-
yl]carbonyl]pyrrolidine 1.88 LC* [M + H].sup.+ =438.0 10 64 97
1-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazin-6- yl]carbonyl]piperidine 1.96 LC* [M + H].sup.+ =452.0
10 65 98 4-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl)
amino]pyridazin-6- yl]carbonyl]morpholine 1.76 LC* [M + H].sup.+
=454.0 10 66 99 1-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazin-6- yl]carbonyl]-4- hydroxypiperidine
1.74 LC* [M + H].sup.+ =468.0 10 67 100 1-[[3-Cyano-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6-
yl]carbonyl]-4- (hydroxymethyl) piperidine 1.76 LC* [M + H].sup.+
=482.0 10 68 101 1-Acetyl-4-[[3- cyano-5-methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6-
yl]carbonyl]piperazine 1.69 LC* [M + H].sup.+ =495.0 10 69 102
4-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[1,2-
b]pyridazin-6- yl]carbonyl]-1- piperazinecarboxylic acid, ethyl
ester 1.85 LC* [M + H].sup.+ =525.0 10 70 103 1-[[3-Cyano-5-
methyl-4-[(4- phenoxyphenyl)amino]pyr- rolo[1,2- b]pyridazin-6-
yl]carbonyl]-3- piperidinecarboxamide 1.72 LC* [M + H].sup.+ =495.0
10 71 104 (2S)-1-[[[3-Cyano-5- methyl-4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazin-6- yl]carbonyl]-2-
(hydroxymethyl) pyrrolidine 1.79 LC* [M + H].sup.+ =468.0 10 72 105
1-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazin-6- yl]carbonyl]-3- hydroxypyrrolidine 1.70 LC* [M +
H].sup.+ =454.0 10 73 106 3-Cyano-N,N-bis(2- hydroxyethyl)-5-
methyl-4-[(4- phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.62 LC* [M + H].sup.+ =472.0 10 74 107
N-(2-Chlorophenyl)-3- cyano-5-methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.09 LC* [M +
H].sup.+ =494.0 10 75 108 N-(3-Chlorophenyl)-3-
cyano-5-methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 2.18 LC* [M + H].sup.+ =494.0 10 76 109
N-(4-Chlorophenyl)-3- cyano-5-methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.17 LC* [M +
H].sup.+ =494.0 10 77 110 3-Cyano-N-(4- methoxyphenyl)-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 2.03 LC* [M + H].sup.+ =490.0 10 78 111
3-Cyano-5-methyl- N-(4-phenoxyphenyl)- 4-[(4-phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.26 LC* [M +
H].sup.+ =552.0 10 79 112 N-[4-(Acetylamino) phenyl]-3-cyano-5-
methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6-
carboxamide 1.91 LC* [M + H].sup.+ =517.0 10 80 113
3-Cyano-5-methyl- N-1-naphthalenyl- 4-[(4-phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 2.09 LC* [M +
H].sup.+ =510.0 10 81 114 1-[[3-Cyano-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6- yl]carbonyl]-4-
phenylpiperazine 2.02 LC* [M + H].sup.+ =529.0 10 82 115
1-[[3-Cyano-5- methyl-4-[(4- phenoxyphenyl) amino]pyridazin-6-
yl]carbonyl]-4- (phenylmethyl) piperazine 1.64 LC* [M + H].sup.+
=543.0 10 83 116 3-Cyano-N-[2- (dimethylamino)
ethyl]-5-methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.60 LC* [M + H].sup.+ =455.0 10 84 117
3-Cyano-N-[2- (diethylamino) ethyl]-5-methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.62 LC* [M +
H].sup.+ =483.0 10 85 118 N-[2-[Bis(1- methylethyl)amino]ethyl]-3--
cyano-5- methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.66 LC* [M + H].sup.+ =511.0 10 86 119
3-Cyano-5-methyl-4-[(4- phenoxyphenyl)amino]-
pyrrolidinyl)ethyl]pyrrolo[- 1,2- b]pyridazine-6- carboxamide 1.62
LC* [M + H].sup.+ =481.0 10 87 120 3-Cyano-5-methyl-4-[(4-
phenoxyphenyl)amino]- N-[2-(1-piperidinyl) ethyl]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.64 LC* [M + H].sup.+ =495.0 10 88 121
3-Cyano-5-methyl-4-[(4- phenoxyphenyl)amino]- N-[2-(4-morpholinyl)
ethyl]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.60 LC* [M +
H].sup.+ =497.0 10 89 122 3-Cyano-5-methyl- N-[3-(4-
morpholinyl)propyl]- 4-[(4-phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.60 LC* [M + H].sup.+ =511.0 10 90 123
3-Cyano-N-[3- (dimethylamino)propyl]- 5-methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.62
LC* [M + H].sup.+ =469.0 10 91 124 3-Cyano-5-methyl-
4-[(4-phenoxyphenyl) amino]-N-[1- (phenylmethyl)-3-
pyrrolidinyl]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.71 LC* [M +
H].sup.+ =543.0 10 92 125 3-Cyano-5-methyl- 4-[(4-phenoxyphenyl)
amino]-N-[1- (phenylmethyl)-4- piperidinyl]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.71 LC* [M + H].sup.+ =557.0 10 93 126
3-Cyano-N-(1-ethyl-3- piperidinyl)-5- methyl-4-[(4- phenoxyphenyl)
amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.65 LC* [M +
H].sup.+ =495.0 10 94 127 3-Cyano-N-[(1-ethyl-2- pyrrolidinyl)
methyl]-5- methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.65 LC* [M + H].sup.+ =495.0 10 95 128
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-(2-
pyridinylmethyl) pyrrolo[1,2- b]pyridazine-6- carboxamide 1.63 LC*
[M + H].sup.+ =475.0 10 96 129 3-Cyano-5-methyl-
4-[(4-phenoxyphenyl) amino]-N-(3- pyridinylmethyl) pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.61 LC* [M + H].sup.+ =475.0 10 97 130
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-(4-
pyridinylmethyl) pyrrolo[1,2- b]pyridazine-6- carboxamide 1.61 LC*
[M + H].sup.+ =475.0 10 98 131 3-Cyano-N-[2-(1H- imidazol-4-
yl)ethyl]-5-methyl- 4-[(4-phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxamide 1.61 LC* [M + H].sup.+ =478.0 10 99 132
3-Cyano-N-[3-(1H- imidazol-1- yl)propyl]-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.62
LC* [M + H].sup.+ =492.0 10 100 133 3-Cyano-5-methyl-
4-[(4-phenoxyphenyl) amino]-N-3- pyridinylpyrrolo[1,
2-b]pyridazine-6- carboxamide 1.70 LC* [M + H].sup.+ =461.0 10 101
134 3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-2-
phenoxyphenyl)amino]-N-2-pyridinylpyrrolo[1, 2-b]pyridazine-6-
carboxamide 1.74 LC* [M + H].sup.+ =461.0 10 102 135
3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-3-
quinolinylpyrrolo[1, 2-b]pyridazine-6- carboxamide 1.96 LC* [M +
H].sup.+ =511.0 10 103 136 3-Cyano-N-ethyl-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazine-6- carboxamide 1.85
LC* [M + H].sup.+ =412.0 10 104 137 (2R)-1-[[3-Cyano-
5-methyl-4-[(4- phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6-
yl]carbonyl]-2- (hydroxymethyl) pyrrolidine 1.79 LC* [M + H].sup.+
=468.0 10 105 138 3-Cyano-5-methyl- 4-[(4-phenoxyphenyl) amino]-N-
phenylpyrrolo[1,2- b]pyridazin-6- carboxamide 2.05 LC* [M +
H].sup.+ =460.0 10 106 139 1-[[3-Cyano-5- methyl-4-[(4-
phenoxyphenyl) amino]pyrrolo[1,2- b]pyridazin-6- yl]carbonyl]-4-
methylpiperazine 1.53 LC* [M + H].sup.+ =467.0 10 107 140
3-Cyano-5-methyl-4-[(3- phenoxyphenyl) amino]pyrrolo[1,2-
b]pyridazine-6- carboxylic acid, ethyl ester 2.01 LCMS* [M +
H].sup.+ =413.0 10 108 141 4-[[4-(4-Chlorophenoxy) phenyl]amino]-3-
cyano-5- methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl
ester 2.13 LCMS* [M + H].sup.+ =447.0 10 109 142 3-Cyano-5-methyl-
4-[[4-(phenylamino) phenyl]amino]pyrrolo[1, 2-b]pyridazine-6-
carboxylic acid, ethyl ester 1.95 LCMS* [M + H].sup.+ =412.0 10 110
143 3-Cyano-5-methyl- 4-[[4-(phenylmethyl) phenyl]amino]pyrrolo[1,
2-b]pyridazine-6- carboxylic acid, ethyl ester 2.05 LCMS* [M +
H].sup.+ =411.0 10 111 144 4-[(4-Benzoylphenyl) amino]-3-cyano-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
1.91 LCMS* [M + H].sup.+ =425.0 10 112 145 4-[(3-Benzoylphenyl)
amino]-3-cyano-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic
acid, ethyl ester 1.92 LCMS* [M + H].sup.+ =425.0 10 113 146
3-Cyano-4- (diethylamino)-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 1.85 LCMS* [M + H].sup.+ =301.0 10 114
147 3-Cyano-5-methyl- 4-(4-phenoxyphenoxy) pyrrolo[1,2-
b]pyridazine-6- carboxylic acid, ethyl ester 2.19 LCMS* [M +
H].sup.+ =414.0 10 115 148 3-Cyano-5-methyl- 4-[[4-phenoxyphenyl)
methyl]amino]pyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl
ester 4.20 LCMS [M + H].sup.+ =427.0 10 116 149 4-[(4-
Butylphenyl)amino]- 3-cyano-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 2.10 LCMS* [M + H].sup.+ =377.0 10 117
150 3-Cyano-4-[[4-(1,1- dimethylethyl) phenyl]amino]-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
2.07 LCMS* [M + H].sup.+ =377.0 10 118 151 4-([1,1-Biphenyl]-
4-ylamino)-3- cyano-5- methylpyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 2.04 LCMS* [M + H].sup.+ =397.0 10 119
152 3-Cyano-4-[(9- ethyl-9H-carbazol- 3-yl)amino]-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
2.05 LCMS* [M + H].sup.+ =438.0 10 120 153 3-Cyano-5-methyl- 4-[[4-
(phenylmethoxy)phenyl]amino]- pyrrolo[1,2- b]pyridazine-6-
carboxylic acid, ethyl ester 2.03 LCMS* [M + H].sup.+ =427.0 10 121
154 [3-Cyano-5-methyl- 4-[(4- phenoxyphenyl)amino]pyrrolo[1,2-
b]pyridazin-6- yl]carbamic acid, methyl ester 3.65 LCMS* [M +
H].sup.+ =414.0 11A 122 155 [3-Cyano-5-methyl- 4-[(4-
phenoxyphenyl)amino]pyrrolo[1,2- b]pyridazin-6-
yl]carbamic acid, 1,1-dimethylethyl ester 3.99 LCMS [M + H].sup.+
=456.0 11A 123 156 3-Cyano-4-(1H- indazol-6- ylamino)-5-
methylpyrrolo[1,2- b]pyridazine-6- carboxylic acid, ethyl ester
3.79 LC [M + H].sup.+ =361.13 1 124 157 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
3-phenyl-urea 3.90 LCMS [M + H].sup.+ =475.0 19 125 158
4-[6-(4-Bromo- phenoxy)-pyridin- 3-ylamino]-3- cyano-5-methyl-
pyrrolo[1,2- b]pyridazine-6- carboxylic acid ethyl ester 4.01 LCMS
[M + H].sup.+ =492.0 1 126 159 3-Cyano-5-methyl- 4-[4-(pyrimidin-2-
yloxy)- phenylamino]- pyrrolo[1,2- b]pyridazine-6- carboxylic acid
ethyl ester 3.38 LC [M + H].sup.+ =415.0 1 127 160 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
benzamide 1.91 LCMS-1 [M + H].sup.+ =460.0 12 128 161
3-Cyano-5-methyl- 4-(6-phenoxy- pyridin-3-ylamino)- pyrrolo[1,2-
b]pyridazine-6- carboxylic acid ethyl ester 3.72 LCMS [M + H].sup.+
=414.0 1 129 162 3-Cyano-5- ethoxymethyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazine-6- carboxylic acid ethyl
ester 2.30 LCMS-1 [M + H].sup.+ =457.0 17 130 163 2-Acetylamino-N-
[3-cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- acetamide 1.70 LCMS-1 [M + H].sup.+ =455.0 12
131 164 3-Acetylamino-N- [3-cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- propionamide 1.73
LCMS-1 [M + H].sup.+ =469.0 12 132 165 4-Acetylamino-N-
[3-cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- butyramide 1.76 LCMS-1 [M + H].sup.+ =483.0 12
133 166 4-Acetylamino-N- [3-cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- benzamide 1.85 LCMS-1
[M + H].sup.+ =517.0 12 134 167 3-Acetylamino-N- [3-cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
benzamide 1.87 LCMS-1 [M + H].sup.+ =517.0 12 135 168 3-Cyano-5-
methoxymethyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazine-6- carboxylic acid 3.86 LCMS [M + H].sup.+ =415.0 4,
17 136 169 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- urea 3.31 LCMS [M + H].sup.+ =399.0
19 137 170 3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- 1,1-dimethyl-urea 3.38 LCMS [M +
H].sup.+ =427.0 19 138 171 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- 3-ethyl-urea 3.54
LCMS [M + H].sup.+ =427.0 19 139 172 N-(2-{3-[3-Cyano-
5-methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- ureido}-ethyl)- acetamide 3.35 LCMS [M +
H].sup.+ =484.0 19 140 173 3-Cyano-5- hydroxymethyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazine-6- carboxylic acid 3.66
LCMS [M + H].sup.+ =401.0 4, 16 141 174 [3-Cyano-5-
methoxymethyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid methyl ester 1.99 LCMS-1 [M +
H].sup.+ =444.0 11, 17 142 175 3-Cyano-5- methoxymethyl-4-
(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazine-6- carboxylic
acid (2- morpholin-4-yl- ethyl)-amide 1.78 LCMS-1 [M + H].sup.+
=527.0 12, 17 143 176 1-[3-Cyano-5- methoxymethyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-morpholin-4-yl-
ethyl)-urea 1.73 LCMS-1 [M + H].sup.+ =542.0 17, 19 144 177
3-(Methane- sulfonylamino-methyl)-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazine-6- carboxylic acid 3.35
LCMS [M + H].sup.+ =467.0 14
EXAMPLE 145
6-(1-Hydroxy-1-methyl-ethyl)-5-methyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,-
2-b]pyridazine-3-carbonitrile
[0307] 178
[0308] To a solution of compound 3A (124 mg, 0.30 mmol) in THF (5
mL) at 0.degree. C. was slowly added a 3.0 M solution of MeMgBr in
ether (0.40 mL, 1.20 mmol). The reaction was warmed to room
temperature and then heated at 50.degree. C. for 1 h. After cooling
to room temperature, the reaction was quenched with EtOAc (20 mL)
and saturated aqueous NH.sub.4Cl (20 mL) was added. The resulting
two layers were separated and the organic layer washed with brine,
dried over Na.sub.2SO.sub.4 and concentrated to an orange oil. This
crude oil was purified by silica gel flash chromatography (eluted
with 14-17% EtOAc/CH.sub.2Cl.sub.2) to give compound 145 as a
yellow solid (76 mg, 64%). HPLC: 97% at 1.97 min (retention time)
(Phenom-Prime S5 C18 column, 4.6.times.30 mm, eluting with 10-90%
aqueous methanol over 2 min containing 0.1% TFA, 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 399 [M+H].sup.+.
EXAMPLE 146
6-Hydroxy-5-methyl-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]pyridazine-3-ca-
rbonitrile
[0309] 179
[0310] To a mixture of H.sub.2O.sub.2 (50% wt in H.sub.2O, 0.0115
mL, 0.20 mmol) and CH.sub.2Cl.sub.2 (2 mL) at -5.degree. C. was
added BF.sub.3.OEt.sub.2. The reaction was stirred at -5.degree. C.
for 40 min before a solution of compound 145 (56 mg, 0.14 mmol) in
CH.sub.2Cl.sub.2 (3 mL) was added. The reaction was kept at
-5.degree. C. for 10 min and quenched with an aqueous solution of
Na.sub.2SO.sub.3 (2 g, 10 mL). The reaction was diluted with
CH.sub.2Cl.sub.2 and the two layers were separated. The aqueous
layer was extracted with CH.sub.2Cl.sub.2 (2.times.10 mL). The
CH.sub.2Cl.sub.2 layers were combined and concentrated in vacuo to
give a brown oil. This crude oil was purified by silica gel flash
chromatography (eluted with 10% EtOAc/CH.sub.2Cl.sub.2) to give
compound 146 as a yellow solid (32 mg, 64%). HPLC: 90% at 1.89 min
(retention time) (Phenom-Prime S5 C18 column, 4.6.times.30 mm,
eluting with 10-90% aqueous methanol over 2 min containing 0.1%
TFA, 5 mL/min, monitoring at 220 nm). MS (ES): m/z 357
[M+H].sup.+.
EXAMPLE 147
5-Methyl-6-(2-morpholin-4-yl-ethoxy)-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-
-b]pyridazine-3-carbonitrile
[0311] 180
[0312] To a solution of compound 146 (8.9 mg, 0.025 mmol) PPh.sub.3
(13.1 mg, 0.05 mmol) and 4-(2-hydroxyethyl)-morpholine (6.6 mg,
0.05 mmol) in dry THF (0.3 mL) under N.sub.2 at 0.degree. C. was
added DEAD (8.7 mg, 0.05 mmol). The reaction was stirred at
0.degree. C. for 5 min, warmed to room temperature for 2 h,
concentrated to dryness, and purified by silica gel flash
chromatography (eluted with 1-5% MeOH/CH.sub.2Cl.sub.2) to give
compound 147 as a yellow oil (10 mg, 85%). HPLC: 96% at 1.69 min
(retention time) (Phenom-Prime S5 C18 column, 4.6.times.30 mm,
eluting with 10-90% aqueous methanol over 2 min containing 0.1%
TFA, 5 mL/min, monitoring at 220 nm). MS (ES): ml/z 470
[M+H].sup.+.
EXAMPLE 148
5-Cyano-7-oxo-6-(4-phenoxy-phenyl)-6,7-dihydro-9H-8-oxa-2a,3,6-triaza-benz-
o[cd]azulene-1-carboxylic acid ethyl ester
[0313] 181
[0314] To a solution of compound 16C (26 mg, 0.061 mmol) and DIPEA
(63 mg, 0.485 mmol) in CH.sub.2Cl.sub.2 (6 mL) at -50.degree. C.
was added triphosgene (30 mg, 0.101 mmol). The reaction was slowly
warmed up to 10.degree. C. over 1 h, quenched with MeOH (1 mL),
concentrated to dryness in vacuo and purified by silica gel flash
chromatography (eluted with 1-2% EtOAc/CH.sub.2Cl.sub.2) to give
compound 148 as a yellow solid (16 mg, 58%). HPLC: 91% at 2.09 min
(retention time) (Phenom-Prime S5 C18 column, 4.6.times.30 mm,
eluting with 10-90% aqueous methanol over 2 min containing 0.1%
TFA, 5 mUrmin, monitoring at 220 nm). MS (ES): m/z 455
[M+H].sup.+.
EXAMPLE 149
5-Azidomethyl-3-cyano-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]pyridazine-6-
-carboxylic acid ethyl ester
[0315] 182
[0316] To a solution of compound 148 (21 mg, 0.05 mmol) in THF (0.6
mL) was added DPPA (22 mg, 0.08 mmol) followed by DBU (9 mg, 0.06
mmol). The reaction was stirred at room temperature for 4 h,
concentrated and purified by flash chromatography on a silica gel
column (0.5-1% EtOAc/CH.sub.2Cl.sub.2) to give compound 149 as a
yellow oil (14 mg, 63%). HPLC: 99% at 2.16 min (retention time)
(PrimeSphere 5u C18-HC column, 4.6.times.30 mm, eluting with 10-90%
aqueous methanol over 2 min containing 0.1% TFA, 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 454 [M+H].sup.+.
EXAMPLE 150
5-Andnomethyl-3-cyano-4-(4-phenoxy-phenylamino)-pyrrolo[1,2-b]pyridazine-6-
-carboxylic acid ethyl ester
[0317] 183
[0318] To a solution of compound 149 (12 mg, 0.026 mmol) in a
mixture of 1:2 THF:MeOH (3 mL) was added Pd/C (5 mg). The reaction
was hydrogenated under a hydrogen balloon at room temperature for
30 min and filtered. The filtrate was concentrated to give 8 as a
yellow solid (9 mg, 80%). No further purification was required.
HPLC: 92% at 1.71 min (retention time) (PrimeSphere 5u C18-HC
column, 4.6.times.30 mm, eluting with 10-90% aqueous methanol over
2 min containing 0.1% TFA, 5 mL/min, monitoring at 220 nm). MS
(ES): m/z 428 [M+H].sup.+.
EXAMPLE 151
5-Cyano-7-oxo-6-(4-phenoxy-phenyl)-6,7,8,9-tetrahydro-2a,3,6,8-tetraaza-be-
nzo[cd]azulene-1-carboxylic acid ethyl ester
[0319] 184
[0320] To a solution of compound 150 (7 mg, 0.016 mmol) and DIPEA
(17 mg, 0.13 mmol) in CH.sub.2Cl.sub.2 (1.5 mL) at -70.degree. C.
was added triphosgene (9.5 mg, 0.032 mmol). The reaction was slowly
warmed up to -5.degree. C. over 1 h, quenched with MeOH (0.5 mL),
concentrated to dryness and purified by silica gel flash
chromatography (eluted with 6-8% EtOAc/CH.sub.2Cl.sub.2) to give 9
as a yellow solid (6 mg, 81%). HPLC: 98% at 1.97 min (retention
time) (PrimeSphere 5u C 18-HC column, 4.6.times.30 mm, eluting with
10-90% aqueous methanol over 2 min containing 0.1% TFA, 5 mL/min,
monitoring at 220 nm). MS (ES): m/z 454 [M+H].sup.+.
EXAMPLES 152 TO 367
[0321] Further compounds of the present invention were prepared by
procedures analogous to those described above. Table 2 provides the
name and structure of representative compounds and their retention
times, as well as the Example number of the procedure on which the
preparation of the compound was based. The chromatography
techniques used to determine the retention times of the compounds
listed in Table 2 are as follows:
[0322] LC=YMC S5 ODS column, 3.6.times.50 mm, eluting with 10-90%
aqueous methanol over 2 min containing 0.1% TFA, 5 mL/min,
monitoring at 220 nm
[0323] LC*=YMC S5 ODS column 4.6.times.50 mm eluting with 10-90%
MeOH/H.sub.2O over 4 minutes containing 0.2% phosphoric acid, 4
mL/min, monitoring at 220 nm.
[0324] The molecular mass of the compounds listed in Table 2 were
determined by MS (ES) by the formula m/z.
2TABLE 2 Retention Time Min./ Ex. Compound Molecular Proc. No.
Compound Structure Name Mass of Ex. 152 185 1-(2-Chloro-ethyl)-
3-[3-cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- urea 1.74 LC [M + H].sup.+ =416.2 19 153 186
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-pyrrolidin-1-yl- ethyl)-urea 1.51 LC [M +
H].sup.+ =496.2 19 154 187 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- [3-(2-oxo-
pyrrolidin-1-yl)- propyl]-urea 1.67 LC [M + H].sup.+ =524.2 19 155
188 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- [2-(1H-imidazol-4- yl)-ethyl]-urea 1.51 LC [M
+ H].sup.+ =493.2 19 156 189 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- [2-(1H-indol-3-yl)-
ethyl]-urea 1.86 LC [M + H].sup.+ =542.2 19 157 190 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (3-morpholin-4-yl- propyl)-urea 1.51 LC [M +
H].sup.+ =526.2 19 158 191 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-pyridin-2-yl-
ethyl)-urea 1.53 LC [M + H].sup.+ =504.2 19 159 192 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-piperidin-1-yl- ethyl)-urea 1.55 LC [M +
H].sup.+ =510.2 19 160 193 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- [3-(2-methyl-
piperidin-1-yl)- propyl]-urea 1.57 LC [M + H].sup.+ =538.3 19 161
194 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-fluoro-ethyl)- urea 1.69 LC [M + H].sup.+
=455.2 19 162 195 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-dimethylamino- ethyl)-urea 1.5
LC [M + H].sup.+ =470.2 19 163 196 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(2-diethylamino- ethyl)-urea 1.54 LC [M + H].sup.+ =498.2 19 164
197 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-methoxy-ethyl)- urea 1.7 LC [M + H].sup.+
=457.2 19 165 198 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- [2-(2-hydroxy-
ethoxy)-ethyl]-urea 1.64 LC [M + H].sup.+ =487.2 19 166 199
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- propyl-urea 1.78 LC [M + H].sup.+ =441.2 19
167 200 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (3-dimethylamino- propyl)-urea
1.52 LC [M + H].sup.+ =484.2 19 168 201 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(3-ethoxy-propyl)- urea 1.79 LC [M + H].sup.+ =485.2 19 169 202
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (3-hydroxy- propyl)-urea 1.62 LC [M + H].sup.+
=457.2 19 170 203 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (4-hydroxy-butyl)- urea 1.63 LC
[M + H].sup.+ =471.3 19 171 204 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- pentyl-urea 1.92 LC
[M + H].sup.+ =467.2 19 172 205 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (5-hydroxy-pentyl)-
urea 1.7 LC [M + H].sup.+ =485.2 19 173 206 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (6-hydroxy-hexyl)- urea 1.75 LC [M + H].sup.+
=499.2 19 174 207 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (3-imidazol-1-yl- propyl)-urea
1.53 LC [M + H].sup.+ =507.2 19 175 208 1-(3-Butoxy-
propyl)-3-[3-cyano- 5-methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.91 LC [M + H].sup.+ =513.2
19 176 209 1-Butyl-3-[3- cyano-5-methyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.85 LC [M +
H].sup.+ =455.2 19 177 210 3-{3-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-propionic
acid ethyl ester 1.77 LC [M + H].sup.+ =499.2 19 178 211
6-{3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- ureido}-hexanoic acid methyl ester 1.81 LC [M +
H].sup.+ =527.2 19 179 212 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- [3-(4-methyl-
piperazin-1-yl)- propyl]-urea 1.44 LC [M + H].sup.+ =539.2 19 180
213 1-(2-Cyano-ethyl)- 3-[3-cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.64 LC [M +
H].sup.+ =452.2 19 181 214 1-{2-[Bis-(2- hydroxy-ethyl)-
amino]-ethyl}-3-[3- cyano-5-methyl-4- (4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.46 LC [M + H].sup.+ =530.2
19 182 215 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (3-methoxy- propyl)-urea 1.71 LC
[M + H].sup.+ =471.2 19 183 216 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(2-diisopropylamino- ethyl)-urea 1.58 LC [M + H].sup.+ =526.2 19
184 217 1-(3-Azepan-1-yl- propyl)-3-[3-cyano- 5-methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.59 LC
[M + H].sup.+ =538.2 19 185 218 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (3-piperidin-1-yl-
propyl)-urea 1.57 LC [M + H].sup.+ =524.2 19 186 219 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-ethoxy-ethyl)- urea 1.76 LC [M + H].sup.+
=471.2 19 187 220 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- [2-(1-methyl-1H- imidazol-4-yl)-
ethyl]-urea 1.52 LC [M + H].sup.+ =507.2 19 188 221 1-(3-Chloro-
propyl)-3-[3-cyano- 5-methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.8 LC [M + H].sup.+ =475.2 19
189 222 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-pyridin-4-yl- ethyl)-urea 1.53
LC [M + H].sup.+ =504.2 19 190 223 3-{3-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
ureido}-propionic acid methyl ester 1.7 LC [M + H].sup.+ =485.2 19
191 224 1-{3-[Bis-(2- hydroxy-ethyl)- amino]-propyl}-3-
[3-cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- urea 1.48 LC [M + H].sup.+ =544.2 19 192 225
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (4-dimethylamino- butyl)-urea 1.53 LC [M +
H].sup.+ =498.2 19 193 226 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (6-dimethylamino-
hexyl)-urea 1.6 LC [M + H].sup.+ =526.2 19 194 227 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-diisobutylamino- ethyl)-urea 1.73 LC [M +
H].sup.+ =552.1 19 195 228 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-thiophen-2-yl-
ethyl)-urea 1.87 LC [M + H].sup.+ =509.1 19 196 229
N-(4-{3-[3-Cyano- 5-methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-butyl)- acetamide 1.66 LC
[M + H].sup.+ =512.2 19 197 230 3-{3-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-
propionamide 1.59 LC [M + H].sup.+ =470.2 19 198 231 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (3-propoxy- propyl)-urea 1.85 LC [M + H].sup.+
=499.2 19 199 232 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-methyl-butyl)- urea 1.9 LC [M
+ H].sup.+ =469.2 19 200 233 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- [3-(methyl-phenyl-
amino)-propyl]- urea 1.61 LC [M + H].sup.+ =546.2 19 201 234
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-isopropoxy- ethyl)-urea 1.82 LC [M +
H].sup.+ =485.2 19 202 235 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-pyridin-3-yl-
ethyl)-urea 1.53 LC [M + H].sup.+ =504.2 19 203 236 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2,2,2-trifluoro- ethyl)-urea 1.76 LC [M +
H].sup.+ =481.1 19 204 237 4-{3-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-butyric acid
methyl ester 1.71 LC [M + H].sup.+ =499.2 19 205 238
(3-{3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- ureido}-propyl)- methyl-carbamic acid tert-butyl
ester 1.9 LC [M + H].sup.+ =570.3 19 206 239 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (1-ethyl-pyrrolidin- 2-ylmethyl)-urea 1.55 LC
[M + H].sup.+ =510.2 19 207 240 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(tetrahydro-furan-2- ylmethyl)-urea 1.77 LC [M + H].sup.+ =483.2 19
208 241 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-hydroxy-2- phenyl-ethyl)-urea
1.79 LC [M + H].sup.+ =519.2 19 209 242 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(2-hydroxy- propyl)-urea 1.66 LC [M + H].sup.+ =457.2 19 210 243
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2,3-dihydroxy- propyl)-urea 1.58 LC [M +
H].sup.+ =473.2 19 211 244 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- isobutyl-urea 1.85
LC [M + H].sup.+ =455.2 19 212 245 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(2-dimethylamino- propyl)-urea 1.52 LC [M + H].sup.+ =484.2 19 213
246 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- cyclopropylmethyl- urea 1.8 LC [M + H].sup.+
=453.2 19 214 247 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-hydroxy-butyl)- urea 1.73 LC
[M + H].sup.+ =471.2 19 215 248 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2-hydroxy-
propyl)-urea 1.66 LC [M + H].sup.+ =457.2 19 216 249 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-hydroxy- propyl)-urea 1.66 LC [M + H].sup.+
=457.2 19 217 250 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (tetrahydro-furan-2-
ylmethyl)-urea 1.77 LC [M + H].sup.+ =483.2 19 218 251
4-{3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- ureido}-butyric acid ethyl ester 1.79 LC [M +
H].sup.+ =513.2 19 219 252 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(1-ethyl-pyrrolidin- 2-ylmethyl)-urea 1.56 LC [M + H].sup.+ =510.2
19 220 253 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- (tetrahydro-furan-2-
ylmethyl)-urea 1.77 LC [M + H].sup.+ =483.2 19 221 254
1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2-hydroxy- cyclohexylmethyl)- urea 1.83 LC [M
+ H].sup.+ =511.2 19 222 255 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2,2-dimethyl-
propyl)-urea 1.9 LC [M + H].sup.+ =469.2 19 223 256 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2,3-dihydroxy- propyl)-urea 1.59 LC [M +
H].sup.+ =473.2 19 224 257 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (2,3-dihydroxy-
propyl)-urea 1.59 LC [M + H].sup.+ =473.2 19 225 258 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- (2,2-dimethyl- [1,3]dioxolan-4- ylmethyl)-urea
1.58 LC [M + H].sup.+ =511.2 19 226 259 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(1-hydroxy- cyclohexylmethyl)- urea 1.84 LC [M + H].sup.+ =511.2 19
227 260 {3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-acetic acid methyl ester
1.66 LC [M + H].sup.+ =471.2 19 228 261 {3-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
ureido}-acetic acid ethyl ester 1.72 LC [M + H].sup.+ =485.2 19 229
262 2-{3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- ureido}-acetamide 1.58 LC [M +
H].sup.+ =456.2 19 230 263 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- (3-hydroxy-2,2-
dimethyl-propyl)- urea 1.77 LC [M + H].sup.+ =485.2 19 231 264
2-{3-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- ureido}-N-methyl- acetamide 1.59 LC [M +
H].sup.+ =470.2 19 232 265 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- furan-2-ylmethyl-
urea 1.79 LC [M + H].sup.+ =479.2 19 233 266 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- naphthalen-1- ylmethyl-urea 1.94 LC [M +
H].sup.+ =539.1 19 234 267 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- thiophen-2-
ylmethyl-urea 1.83 LC [M + H].sup.+ =495.2 19 235 268
1-Benzo[1,3]dioxol- 5-ylmethyl-3-[3- cyano-5-methyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.84 LC [M +
H].sup.+ =533.1 19 236 269 1-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3- pyridin-2-ylmethyl-
urea 1.53 LC [M + H].sup.+ =490.2 19 237 270 1-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- pyridin-3-ylmethyl- urea 1.52 LC [M + H].sup.+
=490.2 19 238 271 1-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- pyridin-4-ylmethyl- urea 1.52 LC
[M + H].sup.+ =490.2 19 239 272 1-Benzyl-3-[3- cyano-5-methyl-4-
(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.85
LC [M + H].sup.+ =489.2 19 240 273 1-(4-Amino-2- methyl-pyrimidin-
5-ylmethyl)-3-[3- cyano-5-methyl-4- (4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- urea 1.53 LC [M + H].sup.+ =520.1
19 241 274 6-Methoxy-5- methyl-4-[methyl- (4-phenoxy-
phenyl)-amino]- pyrrolo[1,2- b]pyridazine-3- carbonitrile 2.17 LC
[M + H].sup.+ =385.2 154 242 275 1-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-3-
(2-hydroxy-ethyl)- urea 1.77 LC [M + H].sup.+ =443.2 19 243 276
3-Cyano-5-(2- methoxy- ethoxymethyl)-4- (4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazine-6- carboxylic acid ethyl ester 2.19 LC [M
+ H].sup.+ =487.2 17B 244 277 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.50 LC [M + H].sup.+ =513.2 11A 245
278 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- methoxy-ethyl ester 1.71 LC [M
+ H].sup.+ =458.2 11A 246 279 3-Cyano-4-(2,4- dichloro-
phenylamino)-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid
ethyl ester 4.39 LC* [M + H].sup.+ =390.0 1E 247 280 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- piperidin-1-yl- propionamide 1.52 LC [M +
H].sup.+ =495.2 12 248 281 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-
dimethylamino- ethyl ester 1.55 LC [M + H].sup.+ =471.2 11A 249 282
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- diethylamino-ethyl ester 1.52
LC [M + H].sup.+ =499.2 11A 250 283 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- phenoxy-ethyl ester 1.90 LC [M + H].sup.+ =520.1
11A 251 284 Acetic acid 2-[3- cyano-5-methyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6- ylcarbamoyloxy]- ethyl
ester 1.72 LC [M + H].sup.+ =486.2 11A 252 285 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- isopropoxy-ethyl ester 1.74 LC [M + H].sup.+
=486.4 11A
253 286 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- ethoxy-ethyl ester 1.77 LC [M +
H].sup.+ =472.2 11A 254 287 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-(3-
methoxy-phenyl)- ethyl ester 1.92 LC [M + H].sup.+ =534.2 11A 255
288 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 3- methoxy-butyl ester 1.81 LC [M
+ H].sup.+ =486.2 11A 256 289 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 3-
dimethylamino- propyl ester 1.56 LC [M + H].sup.+ =485.2 11A 257
290 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 3- diethylamino- propyl ester 1.58
LC [M + H].sup.+ =513.2 11A 258 291 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 3- (3,4-dimethoxy- phenyl)-propyl ester 1.90 LC [M +
H].sup.+ =578.2 11A 259 292 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 3-(4-
methoxy-phenyl)- propyl ester 1.96 LC [M + H].sup.+ =548.2 11A 260
293 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2-(1- methyl-pyrrolidin-
2-yl)-ethyl ester 1.59 LC [M + H].sup.+ =511.2 11A 261 294
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- pyrrolidin-1-yl- ethyl ester
1.56 LC [M + H].sup.+ =497.2 11A 262 295 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid [1,3]dioxolan-4- ylmethyl ester 1.70 LC [M + H].sup.+
=486.2 11A 263 296 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid tetrahydro-furan-3-
yl ester 1.72 LC [M + H].sup.+ =470.2 11A 264 297
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- thiophen-2-yl- ethyl ester 1.90
LC [M + H].sup.+ =510.1 11A 265 298 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- (1,3-dioxo-1,3- dihydro-isoindol-2- yl)-ethyl
ester 1.80 LC [M + H].sup.+ =573.2 11A 266 299 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- pyridin-2-yl-ethyl ester 1.57 LC [M + H].sup.+
=505.2 11A 267 300 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 3-
pyridin-3-yl-propyl ester 1.61 LC [M + H].sup.+ =519.2 11A 268 301
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 1- methyl-piperidin-2- ylmethyl
ester 1.60 LC [M + H].sup.+ =511.2 11A 269 302 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 1- methyl-piperidin-3- ylmethyl ester 1.60 LC [M +
H].sup.+ =511.2 11A 270 303 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-
piperidin-1-yl- ethyl ester 1.54 LC [M + H].sup.+ =511.2 11A 271
304 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- diisopropylamino- ethyl ester
1.59 LC [M + H].sup.+ =527.2 11A 272 305 3-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6- ylcarbamoyloxy]-
2,2-dimethyl- propionic acid methyl ester 1.83 LC [M + H].sup.+
=514.2 11A 273 306 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-(2- methyl-5-nitro-
imidazol-1-yl)- ethyl ester 1.70 LC [M + H].sup.+ =553.2 11A 274
307 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
b]pyridazin-6-yl]- carbamic acid 2- thiophen-3-yl- ethyl ester 1.95
LC [M + H].sup.+ =510.2 11A 275 308 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- [(2-dimethylamino- ethyl)-methyl- amino]-ethyl
ester 1.43 LC [M + H].sup.+ =528.2 11A 276 309 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 3-(6- methyl-pyridin-2- yl)-propyl ester 1.61 LC [M +
H].sup.+ =533.2 11A 277 310 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-(2-
oxo-pyrrolidin-1- yl)-ethyl ester 1.68 LC [M + H].sup.+ =511.2 11A
278 311 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- (methyl-phenyl- amino)-ethyl
ester 1.72 LC [M + H].sup.+ =533.2 11A 279 312 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- azepan-1-yl-ethyl ester 1.56 LC [M + H].sup.+
=525.2 11A 280 313 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2- dimethylamino-2-
methyl-propyl ester 1.57 LC [M + H].sup.+ =499.2 11A 281 314
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 1- methyl-2-piperidin- 1-yl-ethyl
ester 1.58 LC [M + H].sup.+ =525.2 11A 282 315 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 3- piperidin-1-yl- propyl ester 1.60 LC [M + H].sup.+
=525.2 11A 283 316 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 5- oxo-tetrahydro-
furan-2-ylmethyl ester 1.70 LC [M + H].sup.+ =498.2 11A 284 317
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 3- pyridin-2-yl-propyl ester 1.60
LC [M + H].sup.+ =519.2 11A 285 318 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 3-(2- oxo-pyrrolidin-1- yl)-propyl ester 1.72 LC [M +
H].sup.+ =525.2 11A 286 319 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-
propionylamino- ethyl ester 1.68 LC [M + H].sup.+ =499.2 11A 287
320 [3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2-(2- dimethylamino- ethoxy)-ethyl
ester 1.58 LC [M + H].sup.+ =515.2 11A 288 321 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- [(pyridine-4- carbonyl)-amino]- ethyl ester 1.58
LC [M + H].sup.+ =548.2 11A 289 322 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- (2,5-dioxo- pyrrolidin-1-yl)- ethyl ester 1.64 LC
[M + H].sup.+ =523.2 11A 290 323 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid 2-
pyridin-4-yl-ethyl ester 1.57 LC [M + H].sup.+ =505.2 11A 291 324
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 5- hydroxymethyl-3H- imidazol-4-
ylmethyl ester 1.49 LC [M + H].sup.+ =510.2 11A 292 325
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid 2- imidazol-1-yl- ethyl ester 1.55
LC [M + H].sup.+ =494.2 11A 293 326 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 2- (isopropyl-methyl- amino)-ethyl ester 1.58 LC [M +
H].sup.+ =499.2 11A 294 327 3-Cyano-4-[4-(2- methoxy-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid
ethyl ester 1.86 LC [M + H].sup.+ =443.2 1E 295 328 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-3- methoxy- propionamide 1.64 LC [M + H].sup.+
=442.2 12 296 329 4-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6- ylcarbamoyl]- butyric acid methyl ester
1.69 LC [M + H].sup.+ =484.2 12 297 330 N-[3-Cyano-5- methyl-4-(4-
phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6- yl]-3-hydroxy-
propionamide 1.55 LC [M + H].sup.+ =428.2 12 298 331 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-
yl]-3-ethoxy- propionamide 1.71 LC [M + H].sup.+ =456.2 12 299 332
N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6- yl]-3- (1H-indol-3-yl)- propionamide 1.8 LC [M +
H].sup.+ =527.2 12 300 333 N-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6- yl]-3- pyridin-3-yl-
propionamide 1.49 LC [M + H].sup.+ =489.2 12 301 334 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-
yl]-3- diethylamino- propionamide 1.54 LC [M + H].sup.+ =483.3 12
302 335 1-Methyl-1,2,5,6- tetrahydro- pyridine-3- carboxylic acid
[3- cyano-5-methyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- amide 1.51 LC [M + H].sup.+ =479.3 12 303 336
N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- butyramide 1.73 LC [M + H].sup.+ =426.2 12 304
337 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-4- dimethylamino- butyramide 1.55 LC [M +
H].sup.+ =469.3 12 305 338 N-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-2- pyridin-2-yl-
acetamide 1.5 LC [M + H].sup.+ =475.3 12 306 339 N-[3-Cyano-5-
methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-2- pyridin-3-yl- acetamide 1.48 LC [M + H].sup.+
=475.3 12 307 340 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-2- pyridin-4-yl- acetamide 1.47 LC
[M + H].sup.+ =475.3 12 308 341 N-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-2- thiophen-2-yl-
acetamide 1.76 LC [M + H].sup.+ =480.2 12 309 342 Pyridine-2-
carboxylic acid [3- cyano-5-methyl-4- (4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- amide 1.87 LC [M + H].sup.+ =461.2
12 310 343 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- nicotinamide 1.59 LC [M + H].sup.+
=461.2 12 311 344 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- isonicotinamide 1.57 LC [M +
H].sup.+ =461.2 12 312 345 N-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- 2-dimethylamino-
acetamide 1.5 LC [M + H].sup.+ =441.2 12 313 346 2-Cyano-N-[3-
cyano-5-methyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- acetamide 1.64 LC [M + H].sup.+ =423.2 12 314
347 2-tert-Butyl-5- methyl-2H- pyrazole-3- carboxylic acid [3-
cyano-5-methyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- amide 1.85 LC [M + H].sup.+ =520.2 12 315 348
5-Methyl-pyrazine- 2-carboxylic acid [3-cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- amide
1.83 LC [M + H].sup.+ =476.3 12 316 349 1,5-Dimethyl-1H-
pyrazole-3- carboxylic acid [3- cyano-5-methyl-4- (4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- amide 1.78 LC [M +
H].sup.+ =478.3 12 317 350 N-[3-Cyano-5- methyl-4-(4- phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-2- fluoro-3-pyridin-3-
yl-acrylamide 1.6 LC [M + H].sup.+ =505.2 12 318 351 4-Methyl-
[1,2,3]thiadiazole- 5-carboxylic acid [3-cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- amide
1.74 LC [M + H].sup.+ =482.2 12 319 352 1-Methyl-1H- imidazole-2-
carboxylic acid [3- cyano-5-methyl-4- (4-phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]- amide 1.7 LC [M + H].sup.+ =464.3
12 320 353 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-3- dimethylamino- benzamide 1.62 LC
[M + H].sup.+ =503.3 12 321 354 Isoxazole-5- carboxylic acid [3-
cyano-5-methyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- amide 1.67 LC [M + H].sup.+ =451.2 12 322 355
N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-6- methyl- nicotinamide 1.52 LC [M + H].sup.+
=475.3 12 323 356 N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)-
pyrrolo[1,2- b]pyridazin-6-yl]-2- methyl- nicotinamide 1.5 LC [M +
H].sup.+ =475.3 12 324 357 1-Methyl-1H- pyrrole-2- carboxylic acid
[3- cyano-5-methyl-4- (4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- amide 1.76 LC [M + H].sup.+ =463.3 12 325 358
N-[3-Cyano-5- methyl-4-(4- phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]-4- methoxy- butyramide 1.67 LC [M + H].sup.+
=456.2 12 326 359 {3-Cyano-4-[4-(2- methoxy-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic
acid 2- methoxy-ethyl ester 1.64 LC [M + H].sup.+ =488.4 11A 327
360 {3-Cyano-4-[4-(2- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.42 LC [M + H].sup.+ =543.4 11A 328
361 1-{3-Cyano-4-[4- (2-methoxy- phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- 3-(2-methoxy- ethyl)-urea
1.39 LC [M + H].sup.+ =542.4 19 329 362 1-{3-Cyano-4-[4-
(2-methoxy- phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazin-6-yl}- 3-(2-morpholin-4- yl-ethyl)-urea 1.58 LC [M +
H].sup.+ =487.2 19 330 363 {3-Cyano-4-[4-(2- methoxy-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic
acid benzyl ester 1.83 LC [M + H].sup.+ =520.2 11A 331 364
3-Cyano-4-[4-(2- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid 1.66 LC [M +
H].sup.+ =415.2 9 332 365 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid
furan-2-ylmethyl ester 1.80 LC [M + H].sup.+ =480.2 11A 333 366
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid oxiranylmethyl ester 1.67 LC [M +
H].sup.+ =456.3 11A 334 367 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid
furan-3-ylmethyl ester 1.84 LC [M + H].sup.+ =480.3 11A 335 368
[3-Cyano-5-methyl- 4-(4-phenoxy- phenylamino)- pyrrolo[1,2-
b]pyridazin-6-yl]- carbamic acid tetrahydro-furan-2- ylmethyl ester
1.76 LC [M + H].sup.+ =484.3 11A 336 369 [3-Cyano-5-methyl-
4-(4-phenoxy- phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]-
carbamic acid 3- methyl-oxetan-3- ylmethyl ester 1.75 LC [M +
H].sup.+ =484.2 11A 337 370 [3-Cyano-5-methyl- 4-(4-phenoxy-
phenylamino)- pyrrolo[1,2- b]pyridazin-6-yl]- carbamic acid
tetrahydro-furan-3- ylmethyl ester 1.75 LC [M + H].sup.+ =484.3 11A
338 371 3-Cyano-4-[4-(4- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid ethyl ester
1.93 LC [M + H].sup.+ =431.2 1E 339 372 3-Cyano-4-[4-(3-
fluoro-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazine-6- carboxylic acid ethyl ester 1.92 LC [M + H].sup.+
=431.2 1E 340 373 3-Cyano-4-[4-(2- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid ethyl ester
1.86 LC [M + H].sup.+ =431.2 1E 341 374 3-Cyano-4-[4-(3-
methoxy-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazine-6- carboxylic acid ethyl ester 1.96 LC [M + H].sup.+
=443.3 1E 342 375 3-Cyano-4-[4-(4- methoxy-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid
ethyl ester 1.89 LC [M + H].sup.+ =443.2 1E 343 376
3-Cyano-4-[4-(2- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid 1.65 LC [M +
H].sup.+ =403.3 9 344 377 3-Cyano-4-[4-(3- fluoro-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid
1.70 LC [M + H].sup.+ =403.3 9 345 378 3-Cyano-4-[4-(4-
fluoro-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazine-6- carboxylic acid 1.69 LC [M + H].sup.+ =403.3 9 346
379 3-Cyano-4-[4-(3- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid 1.74 LC [M +
H].sup.+ =415.2 9 347 380 3-Cyano-4-[4-(4- methoxy-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazine-6- carboxylic acid
1.66 LC [M + H].sup.+ =415.2 9 348 381 1-{3-Cyano-4-[4-
(2-fluoro-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazin-6-yl}- 3-(2-morpholin-4- yl-ethyl)-urea 1.46 LC [M +
H].sup.+ =530.3 19 349 382 1-{3-Cyano-4-[4- (3-fluoro-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazin-6-yl}-
3-(2-morpholin-4- yl-ethyl)-urea 1.53 LC [M + H].sup.+ =530.3 19
350 383 1-{3-Cyano-4-[4- (4-fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- 3-(2-morpholin-4-
yl-ethyl)-urea 1.50 LC [M + H].sup.+ =530.3 19 351 384
1-{3-Cyano-4-[4- (3-methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- 3-(2-morpholin-4-
yl-ethyl)-urea 1.51 LC [M + H].sup.+ =542.4 19 352 385
1-{3-Cyano-4-[4- (4-methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- 3-(2-morpholin-4-
yl-ethyl)-urea 1.49 LC [M + H].sup.+ =542.4 19 353 386
1-{3-Cyano-4-[4- (2-fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl]- 3-(2-methoxy- ethyl)-urea
1.64 LC [M + H].sup.+ =475.3 19 354 387 1-{3-Cyano-4-[4-
(3-fluoro-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazin-6-yl}- 3-(2-methoxy- ethyl)-urea 1.69 LC [M + H].sup.+
=475.3 19 355 388 1-{3-Cyano-4-[4- (4-fluoro-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazin-6-yl}-
3-(2-methoxy- ethyl)-urea 1.68 LC [M + H].sup.+ =475.3 19 356 389
1-{3-Cyano-4-[4- (3-methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- 3-(2-methoxy- ethyl)-urea
1.68 LC [M + H].sup.+ =487.3 19 357 390 1-{3-Cyano-4-[4-
(4-methoxy-phenoxy)- phenylamino]-5- methyl-pyrrolo[1,2-
b]pyridazin-6-yl}- 3-(2-methoxy- ethyl)-urea 1.64 LC [M + H].sup.+
=487.4 19 358 391 {3-Cyano-4-[4-(2- fluoro-phenoxy)-
phenylamino]-5- methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic
acid 2- morpholin-4-yl- ethyl ester 1.47 LC [M + H].sup.+ =531.3
11A 359 392 {3-Cyano-4-[4-(3- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.53 LC [M + H].sup.+ =531.3 11A 360
393 {3-Cyano-4-[4-(4- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.51 LC [M + H].sup.+ =531.3 11A 361
394 {3-Cyano-4-[4-(3- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.52 LC [M + H].sup.+ =543.4 11A 362
395 {3-Cyano-4-[4-(4- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
morpholin-4-yl- ethyl ester 1.48 LC [M + H].sup.+ =543.4 11A 363
396 {3-Cyano-4-[4-(2- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
methoxy-ethyl ester 1.68 LC [M + H].sup.+ =476.3 11A 364 397
{3-Cyano-4-[4-(3- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
methoxy-ethyl ester 1.68 LC [M + H].sup.+ =476.3 11A 365 398
{3-Cyano-4-[4-(4- fluoro-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
methoxy-ethyl ester 1.72 LC [M + H].sup.+ =476.3 11A 366 399
{3-Cyano-4-[4-(3- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
methoxy-ethyl ester 1.78 LC [M + H].sup.+ =488.3 11A 367 400
{3-Cyano-4-[4-(4- methoxy-phenoxy)- phenylamino]-5-
methyl-pyrrolo[1,2- b]pyridazin-6-yl}- carbamic acid 2-
methoxy-ethyl ester 1.68 LC [M + H].sup.+ =488.3 11A
EXAMPLE 368
VEGFR-2 and FGFR-1 Kinase assays
[0325]
3 Reagents Final Concentration Stock Solution VEGFR-2 FGFR-1 Tris
pH 7.0 20 mM 20 mM BSA 10 mg/ml 25 .mu.g/ml 25 .mu.g/ml MnCl.sub.2
(1 M) 1.5 mM 0.5 mM MgCl.sub.2 (1 M) -- 0.5 mM DTT (1 M) 0.5 mM 0.5
mM Enzyme Stock in 10% glycerol 5 ng/rxn 20 ng/rxn (1 mg/ml)
Polyglu/tyr (10 mg/ml) 80 .mu.g/ml 30 .mu.g/ml ATP (1 mM) 2.5 .mu.M
1.0 .mu.M .gamma.-ATP (10 .mu.Ci/.mu.l) 0.5 .mu.Ci/ml 0.5
.mu.Ci
[0326] Incubation mixtures employed for VEGFR-2 or FGFR-1 assay
contained the synthetic substrate polyGlu:Tyr, (4:1), ATP,
ATP-.gamma.-.sup.33P and buffer containing Mn.sup.++ and/or
Mg.sup.++, dithiothreitol (DTT), bovine serum albumin (BSA), and
Tris buffer. The reaction was initiated by addition of enzyme and
after 60 minutes was terminated by the addition of trichloroacetic
acid (TCA) to a concentration of 30% on a volume percent basis.
Inhibitors in accordance with the invention were brought to a
concentration of 10 mM in DMSO. Assays were prepared in a 96 well
format. Compounds were diluted 1:500 in DMSO and then 1: 10 in
water for a final DMSO concentration of 10%. Aliquots of 10 .mu.L
were added to rows B-H in a 96 well format of 10% DMSO. Aliquots of
20 .mu.l of inhibitor solution were added to row A at a
concentration 5 fold higher than running conditions. A 10 .mu.L
aliquot was transferred to each row with 10 pipetting phases for
mixing, and at row F a 10 .mu.L aliquot was discarded. Row G was a
control with no compound and row H was a no-compound and no-emzyme
control. Enzyme and substrate were delivered using a Tomtec Quadra
station.
[0327] Plates were covered with sticky plate tops, incubated at
27.degree. C. for 60 minutes, and then acid precipitated with TCA
for 20 minutes on ice. The precipitate was transferred to
UniFilter-96, GF/C microplates using either a Tomtec or Packard
FilterMate harvester. Activity was determined by quantifying the
incorporated radioactivity using a Packard TopCount Microplate
Scintillation Counter following the addition of Microscint-20
cocktail into each dried well of the UniFilter microplates.
[0328] Tested compounds of formula I inhibited VEGFR-2 and FGFR-1
kinases with IC.sub.50 values .ltoreq.80 .mu.M.
EXAMPLE 369
HER.sub.1, HER.sub.2 or HER.sub.4 Kinase assays
[0329] Compounds of interest were assayed in a kinase buffer that
contained 20 mM Tris.HCl, pH 7.5, 10 mM MnCl.sub.2, 0.5 mM
dithiothreitol, bovine serum albumin at 0.1 mg/ml, poly(glu/tyr,
4:1) at 0.1 mg/ml, 1 .mu.M ATP, and 4 .mu.Ci/ml
[.gamma.-.sup.33P]ATP. Poly(glu/tyr, 4:1) is a synthetic polymer
that serves as a phosphoryl acceptor and is purchased from Sigma
Chemicals. The kinase reaction is initiated by the addition of
enzyme and the reaction mixtures were incubated at 26.degree. C.
for 1 h. The reaction is terminated by the addition of EDTA to 50
mM and proteins are precipitated by the addition of trichloroacetic
acid to 5%. The precipitated proteins are recovered by filtration
onto Packard Unifilter plates and the amount of radioactivity
incorporated is measured in a Topcount scintillation counter.
[0330] For the preparation of recombinant HER1 and HER.sub.4, the
cytoplasmic sequences of the receptors were expressed in insect
cells as GST fusion proteins, which were purified by affinity
chromatography. The cytoplasmic sequence of HER.sub.2 was subcloned
into the baculovirus expression vector pBlueBac4 (Invitrogen) and
was expressed as an untagged protein in insect cells. The
recombinant protein was partially purified by ion-exchange
chromatography.
[0331] The instant compounds inhibit HER.sub.1, HER.sub.2, and
HER.sub.4 kinases with IC50 values between 0.001 25 .mu.M.
Preferred compounds have IC.sub.50 values between 0.001-5.0 .mu.M.
More preferred compounds have IC.sub.50 values between 0.001-1.0
.mu.M. Most preferred compounds have IC.sub.50 values between
0.001-0.1 .mu.M.
[0332] A HERG potassium channel assay may be used to screen
compounds for HERG activity (see Caballero R.sub.1et al., "Direct
Effects of Candesartan and Eprosartan on Human Cloned Potassium
Channels Involved in Cardiac Repolarization," Molecular
Pharmacology, 59(4), 825-36, (2001)). Accordingly, preferred
compounds have lower HERG assay activity.
[0333] For the preparation of recombinant HER.sub.1, the
cytoplasmic sequences of the receptor were expressed in insect
cells as a GST fusion protein, which was purified by affinity
chromatography. The cytoplasmic sequence of HER.sub.2 was subcloned
into the baculovirus expression vector pBlueBac4 (Invitrogen) and
was expressed as an untagged protein in insect cells. The
recombinant protein was partially purified by ion-exchange
chromatography.
[0334] Tested compounds of formula I inhibited HER-1 and HER-2
kinases with IC50 values .ltoreq.100 .mu.M.
EXAMPLE 370
MEK-ERK Kinase Cascade Assay
[0335] An in vitro 96-well plate assay described in Example 388,
above, was adopted with several modifications. Each well contained
30 l assay buffer (Tris-HCl, pH 7.5, MgCl.sub.2, DTT, BSA, Myelin
basic protein (MBP), ATP and [.gamma.-.sup.33P]ATP), 10 .mu.l
inhibitor dilutions or empty DMSO solvent and 10 .mu.l enzyme
mixture (5-10 ng MEK-EE and 100-200 ng ERK). The final
concentrations in the assay were 20 mM Tris-HCl, pH 7.5, 10 mM
MgCl.sub.2, 0.3 mM DTT, 50 .mu.g BSA, 50 .mu.g Myelin basic protein
(MBP), 10 .mu.M ATP and 200 nCi [.gamma.-.sup.33P]ATP. The plates
were incubated at room temperature for 60 min and reactions were
terminated by the addition of 10 .mu.l stop mixture containing 300
mM EDTA and 25 .mu.g BSA. The samples were subjected to
precipitation with TCA containing ATP (final concentrations: TCA,
3.2% and ATP, 2.3 mM). The samples were transferred to a Packard
GF/C 96-well Unifilter plates using a Packard Filter Mate 196
Harvester. Following drying under light, the radioactivity of the
residue in the wells was counted with a Packard Top Count
microplate counter.
[0336] Since this assay is a cascade assay, inhibitors of both MEK
and ERK are expected to be identified. Further in vitro analysis is
required to determine whether the "hits" were attributable to the
inhibition of MEK (using MEK and kinase deficient ERK) or ERK
(using activated ERK and MBP) inhibitors. Nevertheless, tested
compounds of formula I inhibited MEK and/or ERK with IC.sub.50
values .ltoreq.10 .mu.M.
EXAMPLE 371
Generation of p38 Kinases
[0337] cDNAs of human p38.alpha., .beta. and .gamma. isozymes were
cloned by PCR. These cDNAs were subcloned in the pGEX expression
vector (Pharmacia). GST-p38 fusion protein was expressed in E. coli
and purified from bacterial pellets by affinity chromatography
using glutathione agarose. p38 fusion protein was activated by
incubating with constitutively active MKK6. Active p38 was
separated from MKK6 by affinity chromatography. Constitutively
active MKK6 was generated according to Raingeaud et al. [Mol. Cell.
Biol., 1247-1255 (1996)].
EXAMPLE 372
TNF-.alpha. Production by LPS-Stimulated PBMCs
[0338] Heparinized human whole blood was obtained from healthy
volunteers. Peripheral blood mononuclear cells (PBMCs) were
purified from human whole blood by Ficoll-Hypaque density gradient
centrifugation and resuspended at a concentration of
5.times.10.sup.6/ml in assay medium (RPMI medium containing 10%
fetal bovine serum). 50 ul of cell suspension was incubated with 50
ul of test compound (4.times. concentration in assay medium
containing 0.2% DMSO) in 96-well tissue culture plates for 5
minutes at RT. 100 ul of LPS (200 ng/ml stock) was then added to
the cell suspension and the plate was incubated for 6 hours at
37.degree. C. Following incubation, the culture medium was
collected and stored at -20.degree. C. TNF-.alpha. concentration in
the medium was quantified using a standard ELISA kit
(Pharmingen-San Diego, Calif.). Concentrations of TNF-.alpha. and
IC.sub.50 values for test compounds (concentration of compound that
inhibited LPS-stimulated TNF-.alpha. production by 50%) were
calculated by linear regression analysis.
EXAMPLE 373
p38 Assay
[0339] The assays were performed in V-bottomed 96-well plates. The
final assay volume was 60 .mu.l prepared from three 20 .mu.l
additions of enzyme, substrates (MBP and ATP) and test compounds in
assay buffer (50 mM Tris pH 7.5, 10 mM MgCl.sub.2, 50 mM NaCl and 1
mM DTT). Bacterially expressed, activated p38 was pre-incubated
with test compounds for 10 min. prior to initiation of reaction
with substrates. The reaction was incubated at 25.degree. C. for 45
min. and terminated by adding 5 .mu.l of 0.5 M EDTA to each sample.
The reaction mixture was aspirated onto a pre-wet filtermat using a
Skatron Micro96 Cell Harvester (Skatron, Inc.), then washed with
PBS. The filtermat was then dried in a microwave oven for 1 min.,
treated with MeltilLex A scintillation wax (Wallac), and counted on
a Microbeta scintillation counter Model 1450 (Wallac). Inhibition
data were analyzed by nonlinear least-squares regression using
Prizm (GraphPadSoftware). The final concentration of reagents in
the assays are ATP, 1 .mu.M; [.gamma.-.sup.33P]ATP, 3 nM,; MBP
(Sigma, #M1891), 2 .mu.g/well; p38, 10 nM; and DMSO, 0.3%.
EXAMPLE 374
TNF-.alpha. Production by LPS-Stimulated Mice
[0340] Mice (Balb/c female, 6-8 weeks of age, Harlan Labs;
n=8/treatment group) were injected intraperitoneally with 50 ug/kg
lipopolysaccharide (LPS; E coli strain 0111:B4, Sigma) suspended in
sterile saline. Ninety minutes later, mice were sedated by
CO.sub.2:O.sub.2 inhalation and a blood sample was obtained. Serum
was separated and analyzed for TNF-alpha concentrations by
commercial ELISA assay per the manufacturer's instructions (R&D
Systems, Minneapolis, Minn.).
[0341] Test compounds were administered orally at various times
before LPS injection. The compounds were dosed either as
suspensions or as solutions in various vehicles or solubilizing
agents.
[0342] The entire disclosures of the publications cited above are
incorporated herein by reference. While certain preferred
embodiments of the present invention have been described and
specifically exemplified above, it is not intended that the
invention be limited to such embodiments. Various modifications may
be made to the invention without departing from the scope and
spirit thereof as set forth in the following claims.
* * * * *