WO 2004/048382 PCT/US2003/032747
QUINOLINYI-PYRROLOPYRAZOLES
The invention relates to new quionolinyl-pyrazole compounds and their use as
pharmaceutical agents, in particular their use as TGF-beta signal transduction inhibitors,
5
BACKGROUND OF THE INVENTION
The transforming growth factor-beta (TGF-beta) ("TGF-β") polypeptides
influence growth, differentiation, and gene expression in many cell types. The first
10 polypeptide of this family (hat was characterized, TGF-βl, has two identical 112 amino
acid subunits that are covalently linked. TGF-βl is a highly conserved protein with only
a single amino acid difference distinguishing humans from mice. There are two other
members of the TGF-β gene family that are expressed in mammals. TGF-β2 is 71%
homologous to TGF-βl (de Martin, et al. (1987) EMBO J. 6:3673-3677), whereas
15 TGF-β3 is 80% homologous to TGF-β1 (Derynck, et al. (1988) EMBO J 7:3737-3743).
The structural characteristics of TGF-β1 as determined by nuclear magnetic resonance
(Archer, el al. (1993) Biochemistry 32:1164-1171) agree with the crystal structure of
TGP-β2 (Daopin, et al. (1992) Science 257:369-374; Schlunegger and Grutter (1992)
Nature 358:430-434).
20 There are at leasl three different extracellular TGF-β receptors, Type I, II and III
that are involved in the biological functions of TGF-β1, -β2 and -β3 (For reviews, see
Derynek (1994) TIBS 19:548-553 and Massague (1990) Ann. Rev. Cell Biol. 6:597-641).
The Typt I and Type II receptors are trasmembrane serinethreonine kinases, which in
the presence of TGF-β form a heteromeric signaling complex (Wrana, et al (1992)
25 Cell 71: 1003-1014)-
The mechanism of activation of the heteromeric signaling complex at the cell
surface has been elucidated (Wrana, et al. (1994) Nature 370: 341-347), TGF-β first
binds the type II receptor that is a constitutively active transmembrane serine/threonine
30 kinase. The type I receptor is subsequently recruited into the complex, phoshoiylated at
the GS domain and activated to phosphorylate downstream signaling components (e.g.
Smad proteins) to initiate the intracellular signaling cascade. A constitutively active type I
receptor (T204D mutant) has been shown to effectively transduce TGF-β responses,thus

WO 2004/048382 PCT/US2003/032747
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bypassing the requirement for TGF-β and the type II receptor (Wieser, et at. (1995)
EMBO J 14; 2199-2208). Although no signaling function has been discovered for the
type III receptor, it does increase TGF-β2's affinity for the type II receptor making it
essentially equipotent with TGF-β1 and TGF-β3 (Lopez-Casillas, el al. (1993) Cell
5 73: 1435-1444).
Vascular endothelial cells lack the Type III reccplor. Instead endothelial cells
express a structurally related protein called endoglin (Cheifete, et al. (1392) J. Biol.
Chem. 267:19027-19030), which only binds TGF-βl and TGF-β3 with high affinity.
Thus, the relative potency of the TGF-β's reflects the type of receptors expressed in a cell
10 and organ system. In addition to the regulation of the components in the multi-factorial
signaling pathway, the distribution of the synthesis of TGF-β polypeptides also affects
physiological function. The distribution of TGF-β2 and TGF-β3 is more limited
(Derynck, et al. (1988) EMBO J 7:3737-3743) than TGF-β1, e.g., TGF-β3 is limited to
tissues of mesenchymal origin, whereas TGF-β1 is present in both tissues of
15 mesenchymal and epithelial origin,
TGF-βl is a multifunctional cytokine critical for tissue repair. High
concentrations of TGF-β1 are delivered to the site of injury by platelet granules (Assoian
and Spom (1986) J.Cell Biol. 102:1217-1223). TGF-βl initiates a series of events that
promote healing including chemo taxis of cells such as leukocytes, manocytes and
20 ibroblasts, and regulation of growth factors and cytokines involved in angiogenesis, cell
ivision associated with tissue repair and inflammatory responses. TGF-βl also
stimulates the synthesis of extracellular matrix components (Roberts, et al. (1986) Proc.
Natl. Acad. Sci. USA 83:4167-4171; Sporn, et al. (19E3) Science 219:1329-1330;
Massague (1987) Cell 49:437-438) and most importantly for understanding the
25 pathophysiology of TGF-βl, TGF-β1 autoregulates its own synthesis (Kim, et al. (19S9)
J. Biol. Chem. 264:7041-7045).
The compounds disclosed herein may also exhibit other kinase activity, such as
p3S kinase inhibition and/or KDR (VEGFR2) kinase inhibition. Assays to deteimine such
kinase activity are known in the art and one skilled in the art would be able to test the
30 disclosed compounds for such activity.

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SUMMARY OF THE INVENTION
The disclosed invention also relates to the select compound of Formula II:

5
Formula II
2-(6-methyl-pyridin-2-yl)-3-(6-arnido-quinolin-4-yl)-5,6-(dihydro-4H-pyrrolat [l,2-
b]pyrazole
10
and the pharmaceutically acceptable salts thereof.
The compound above is generically disclosed and claimed in FCT patent application
PCT7US02/11834, filed 13 May 2002, which claims priority from U.S. patent application
15 U.S.S-N. 60/293,464, filed 24 May 2001, and incorporated herein by reference. The
above compound has been selected for having a surprisingly superior toxicology profile
over the compounds specifically disclosed in application cited above.
DETAILED DESCRIPTION OF THE INVENTION
20
The term "effective amount" as used in "an effective amount of a compound of
Formula I," for example, refers to an amount of a compound of (he present invention that
is capable of inhibiting TGF-beta,
The term μM refers to micromolar.
25 The general chemical terms used heiein have their usual meanings.
The following abbreviations are used throughout the synthesis Schemes and
Examptes.
DMF refers to dimethyl formamide
THF refers to tetrahydrofuran
30 Ms refers to mesyl which is methylsulfonyl
THP refers to tetrahydropyran

WO 2004/048382 PCT/US2003/032747
4
The compounds disclosed herein can be made according to the following schemes
and examples. The examples should in no way be understood to be limiting in any way
as to how the compounds may be made.
5 The following scheme illuslrates the preparation of the compound of Formula 1.

10

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The following scheme illustrates the preparation of the compound of Formula II.

5
The following examples further illustrate the preparation of the compounds of this
invention ss shown schematically in Schemes land II.

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Example 1
Preparation of 7-(2-morpholin-4-yl-cthoxy)-4-(2-pyridin-2-yl-5,6-dihydro-4H-
pyrrolo [1,2-b] pyrazol -3-y l)-quinoline
5
A. Preparation of 4-{K2-pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l.2-b]pyrazol-3-yl)-
7-[2-(tetrahydropyran-2-yloxy)ethoxy] quinoline
Heat 4-(2-pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l,2-b)pyrazol-3-yl)-quinolin-7-o](376
10 mg, l.146 mmol), cesium carbonate (826 mg, 2.54 mmol), and 2-(2-
brornethoxy)tetralhydro-2H-pyran (380 μL, 2.52 mmol) in DMF (5 mL) at 120°C for 4
hours. Quench the reaction with saturated sodium chloride and then extract with
chloroform. Dry the organic layer over sodium sulfate and concentrate in vacuo. Purify
the reaction mixture on a silica gel column eluting with dichloromethane to 10%
15 methanol in dichloromethane to give the desited subtitled intermediate as a yellow oil
(424 mg, 81%). MS ES+m/e 457.0 (M+I).
B. Preparation of 2-[4-(pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l,2-b]pyrazol-3-
20 yl)-quinolin-7-yloxyl-ethanol
Heat a solution of 4-(2-pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l ,2-b]pyrazol-3-yl)-
7-[2-(tetrahydropyran-2-yloxy)ethoxy]quinoline (421 mg, 0.92 mmol) in acetic acid:
tetrahydrofuran: water (4:2:1) (20 mL). Remove the solvent in vacuo and recover the
25 residue with chloroform:isopropyl (3:1). Wash the organic layer with saturated sodium
bicarbonate and dry over sodium sulfate. Concentrate in vacuo. The residue will be pure
enough for the next step in the scheme (425 mg, 100%). MS ES+m/e 373.1 (M+1).
C. Preparation of methanesulfonic acid 2-[4-(2-pyridin-2-yl-5,6-(iihydro-4H-
30 pyrrolo[l,2-b]pyrazol-3-yl)-quinolin-7-yloxyl-ethyl ester
Stir a solution of 2-[4-(2-pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l,2-b]pyrazol-3-
yl)-quinolin-7-yloxy]-ethanol (293 mg, 0,78 nunol) and methane sulfonyl chloride (68
μL,0.81 ml)in dried pyridine(5 mL)for 2 hours. Remove the pyridine in vacuo and
35 recover the residue with chloroform. Wash the organic layer with saturated sodium
bicarbonate and dry over sodium sulfete to give the desired subtitled intermediate as a
white foam (425 mg, 100%). MSES+m/e451.1 (M+l).

WO 2004/048382 PCT/US2003/032747
7
D. Preparation of 7-(2-morpholin-4-yl-ethoxy)-4-(2-pyridin-2-yl-5,6-dihydro-
4H-pyrrolo[l,2-b]pyrazol-3-yl)-quinoline

5
Heat methanesulfonic acid 2-[4-(2-pyridin-2-yl-5,6-dihydro-4H-pyrrolo[l,2-
b]pyrazol-3-yl)-quinolin-7-yloxy]-ethyl ester (87 mg, 0.19 mmol) with morpholine (l
mL.) at 50 °C for 4 hours. Remove the morpholine in vacua and then extract the product
with isopropyl alcohol:chloroform (1:3). Wash the organic layer with sodium chloride
10 and dry over sodium sulfate. Concentrate in vacuo to give the desired title product as a
slight yellow solid (83 mg, 100%). MS ES+ m/e 442.0 (M+l).
EXAMPLE 2
15
preparation of
2-(6-melhyl-pyridin-2-yl)-3-[6-amido-quinolin-4-yl)-5,6-dihydro-4H-pyrrolo[l,2-
b]pyrazote
20 A. Preparation of 6-hromo-4-methyl-quinoline
Stir a solution of 4-bromo-phenytamine (1 eq), in 1,4-dioxane and cool to
approximately 12 0C. Slowly add sulfuric acid (2 eq) and heat at reflux. Add
metbylvinyl ketone (1.5 eq) dropwiseinto the refluxing solution. Heat the solution for 1
25 hour after addition is complete. Evaporate the reaction solution to dryness and dissolve in
methylene chloride. Adjust the solution to pH 8 with 1 M sodium carbonate and extract
three times with water. Chromatograph the residue on SiO2 (70/30 hexane/ethyl acetate)
to obtain the desired subtitled intermediate.
MS ES+ m/e = 158.2 (M+l).
30

WO 2004/048382 PCT/US2003/032747
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B. Preparation of 6-methyl-pyridime-2-carboxylic acid melhyl ester
Suspend 6-methyl-pyridine-2-carboxylic acid (10 g, 72.9 mmol) in methylene
chloride (200 mL). Cool to 0°C. Add methanol(10mL), 4-dimethylaminopyriaine (11,6
5 g, 94.8mmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC)
(18.2 g, 94.8 mmol). Stir the mixture at room temperature for 6 hours, wash with water
and brine, and dry over sodium sulfate. Filter the mixture and concentrate in vacuo,
Chromatograph the residue on SiO2 (50% ethyl acetate/hexanes) to obtain the desired
subtitled intermediate, 9.66 g (92%), as a colorless liquid.
10 1H NMR (CDCI3) δ 7.93-7.88 (in, IH), 7.75-7.7 (m, 1H), 7.35-7.3 {m, 1H), 4.00 (s, 3H),
S.60 (s, 3H).
C. Preparation of 2-(6-bronw-quinol in-4-yl)-l-(6-methyl-pyridin-2-yl)-ethanone
15 Dissolve 6-bromo-4-methyl-quinoline (38.5 g, 153 mmol) in 600 mLdry THF.
Cool to-700C and beat with the dropwise addition of 0.5 M potassium
hexamethyldisilazane (KN(SiMe3)2 (400 mL, 200 mmol) over 2 hours while keeping the
temperature below -65 °C. Stir the resultant solution at -70°C for 1 hour and add a
solution of 6-methylpyridine-2-carboxylic add methyl ester (27.2, 180 mmol) in 100 mL
20 dry THF dropwise over 15 minutes. During the addition, the mixture will turn from dark
red to pea-green and farm a precipitate. Stir the mixture at -700C over 2 hours then allow
it to warm to ambient temperature with stirnng for 5 hours. Cool the mixture then quench
with 12NHCl to pH=l. Raise the pH to 9 with solid potassium carbonate. Decant the
solution from the solids and extract twice with 200 mL ethyl acetate. Cambine the
25 organic extracts, wash with water and dry over potassium carbonate. Stir the solid in
200 mL water and 200 mL ethyl acetate and treat with additional potassium carbonate.
Separate the organic portion and dry with the previous ethyl acetate extracts. Concentrate
the solution in vacuo to a dark oil. Pass the oil through a 300 mL silica plug with
methylene chloride then ethyl acetate. Combine the appropriate fractions and concentrate
30 in vacuo to yield an amber oil. Rinse the oil down the sides of the flask with methylene
chloride then dilute with hexane while swirling the flask to yield 38.5 g (73.8 %) of the
desired subtitled intermediate as a yellow solid.
MSES+ = 341(M+1).

WO 2004/048382 PCT/US2003/032747
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D. Preparation of l-[2-(6-bromo-quinolin-4-yl)-l-(6-methyl-pyridin-2-yl)-
ethylideneaminol-pyrrolidin-2-one
Stir a mixture of 2-(6-bromo-quinolin-4-yl)-l-(6-methyl-pyridin-2-yl)-ethanone
5 (38.5 g, 113 mmol) and l-aminopyrrolidnione hydrochloride (20 g, 147 mmol) in 115
mL pyridine at ambient temperature for 10 hours. Add about 50 g4 Åunactivated
sieves. Continue stirring an additional 13 h and add 10-15 g silica and filter the
mixture through a 50 g silica plug. Elute the silica plug with 3 L ethyl acetate.
Combine the filtrates aud concentrate in vacuo. Collect the hydrazone precipitate by
10 filtration and suction dry to yield 33.3 g (69.7%) of the desired subtitled intermediate
as an off-white solid
MSES+ = 423(M+1).
E. Preparation of 6-bromo-4-[2-(6-methyl-pyridin-2-yl)-5,6-dihydro-4H-
15 pyrrolo[l,2-b]pyrazol-3-yl]-quinoline
To a mixture of (12 eq,) cesium carbonate and 1-[2-(6-bromo-qumolm-4-yl)-l-
(6'methyl-pyridiri-2-yl)-cthylideneamino]-pyrrolidin'2-otie (33.3 g, 78.7 mmol) acid
300 mL dry N,N-dimethylformamide. Stir the mixture 20 hours at 1000C. The
20 mixture may turn dark during the reaction. Remove the N,N-dimethylformamide in
vacno. Partition the residue between water and methylene chloride. Exrtract. the
aqueous portion with additional methylene chloride. Filter the organic soiutions
through a 300 mL silica plug, eluting with 1.5 L methylene chloride, 1.5 L ethyl
acetate and 1.5 L acetone- Combine the appropriate fractions and concentrate in
25 vacuo. Collect the resultin gprecipitate by filtration to yield 22.7 g (71.2%) of the
desired subtitled intetmediate as an off-white solid.
M5ES+ = 405(M+1).
F. Preparation of 4-[2-(6-incthyl-pyridin-.2-yl)-5,6-dihydro-4H-pyrrolo[l,2-
30 b]pyrazol-3-yl]-quino1ine-6-carboxylic acid methyl ester
Add 6-bromo-4-[2-(6-mothyl-pyridin-2-yl)-5,6-dihydro-4H-pyrrolo[l,2-
b]pyrazol-3-yl]-quinoline (22.7 g. 45 mmol) to a mixture of sodium acetate (19 g, 230
mmol) and the palladium catalyst [1, ].
35 bis(diphenylphosphino)ferraceneJdichloropalladium(II), complex with

WO 2004/048382 PCT/US2003/032747
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dichloromethane (1:1) (850 mg, 1.04 mmol) in 130 mL methanol. Place the mixture
under 50 psi carbon monoxide atmosphere and stir while warming to 90o C over 1 hour
and -with constant charging with additional carbon monoxide. Allow the mixture to
cool over 8 hours, recharge again with carbon monoxide and beat to 90 °C. The
5 pressure may rise lo about 75 PSI. The reaction is complete in about an hour when the
pressure is etable and tle (1:1 tolnenelacetone) snows no remaining bromide. Partition
the mixture between methylene chloride (600 mL) and water (1 L). Extract the
aqueous portion with an additional portion of methylene chloride (400 mL.) Filter the
organic solution through a 300 mL silica plug and wash with 500 mL meihytene
10 chloride. 1200 mL ethyl acetate and 1500 mL acetone. Discard the acetone portion.
Combine appropriate fractions and concentrate to yield 18.8 g(87.4%) of the desired
subtitled intermediate as a pink powder,
MSES+=385(M+1).
15 G. Preparation of 2-(6-methyl-pyridin-2-yl)-3-[6-amido-quinolin-4-yl)-5,6-
dlhydro-4H-pyrrolo[l,2-b]pyrazole

20 Warm a mixture of 4-[2-(6-methyl-pyridin-2-yl)-5,6-dihydro-4H-pyrrolo[l,2-
b]pyrazol-3-yl]-quinoline-6-carboxylic acid methyl ester in 60 mL 7 N ammonia in
methanol to 90 oC in a stainless steel pressure vessel for 66 hours. The pressure will rise
to about 80 PSI. Maintain the pressure for the duration of the reaction. Cool the vessel
and concentrate the brown mixture in vacuo. Purify the residual solid on two 12g Redi-
25 Pak cartridges coupled in series eluting with acetone. Combine appropriate fractions and
coocentrate in vacuo. Suspend the resulting nearly white solid in methylene chloride,
dilute with hexane, and filter. The collected off-white solid yields 1.104 g (63.8%) of the
desired title product.
MS ES+ = 370(M+1)

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The compounds disclosed herein ware tested by the following protocols for TGF-
ß inhibition, as described below in the protocol description.
TGF-ß RECEPTOR 1 PURIFICATION AND IN VITRO KINASE
5 REACTIONS
For TGF-ß Type 1 (RIT204D) Receptors:
The 6X-HIS tagged cytoplasmic kinasc domain of each receptor was expressed
and purified from S19 insect cell lysates as briefly described below:
10 Cell pellets after 48-72 hours of infection were lysed in lysis buffer (L8: 50 mM
Tris pH 7.5, 150 mM NaCI, 50 mM NaF, 0.5% NP40 with freshly added 20 mM
ß-mercaptoelhanol, 10 mM imidazole, 1 mM PMSF, IX EDTA-free Complete Protease
Inhibitor(Boehringer Marunheim)
Cell lysates were clarified by centrifugation and 0.45 uM filtered prior to
15 purification by Ni/NTA affinity chromatography (Qiagen).
Chromatography Protocol:
Eiquilibrate with 10 CV of LB. load sample, wash with 10 CV RJPA buffer (50
20 mM Tris pH 7.5, 150 mM NaCI, 1% NP40, lmM EDTA, 0.25% sodium dcoxycholate,
added ftcsh 20 mM ß-mercaploethanol, 1 mM PMSF), wash with 10 CV LB, wash with
10 CV IX KB (50 mM Tris pH 7.5, 150 mM NaCI, 4 mM MgCl2, 1 mM NaF, 2 mM
ß-mercaptoethanol), clute with a linear gradient of IX KB containing 200 mM Imidazole.
Both enzymes were approximately 90% pure and had autopltosphorylation
25 activity.
Reactions: 170-200 nM enzyme in IX KB, compound dilution series in IX
KB/16% DMSO (20 µM to 1 nM final concentration wilh 4% DMSO final
concentaition), reactions started by adding ATP mix (4 µM ATP/1 µCi 33P-γ-ATP final
concentaitions) in IX KB.
30 Reactions were incubated at 30 °C for 1 hour. Reactions were stopped and
quantitated using standard TCA/BSA precipitation onto Millipore FB glass fiber filter
plates arfd by liquid scintillation counting on a Micro Beta JET,

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The compounds disclosed herein inhibit the TGF-βType I (RJT204D) receptor
kinase domain with IC50 values <20 βM, while exhibiting less toxicity in vivo than
structurally related compounds as disclosed in PCT patent application PCT/US02/11884
identified above.
5 Conditions "characterized by enhanced TGF-β activity" include those wherein
TGF-β synthesis is stimulated so that TGF-β is present al increased levels or wherein
TGF-β latent protein is undesirably activated or converted to active TGF-β protein or
wherein TGF-β receptors are upregulated or wherein the TGF-β protein shows enhanced
binding to cells or extracellular matrix in the location of the disease. Thus, in either case
10 "enhanced activity" refers to any condition wherein the biological activity of TGF-β is
undesirably high, regardless of the cause.
A Dumber of diseases have been associated with TGF-βl over prcduction
Inhibitors of TGF-β intracellular signaling pathway are useful treatments for
fibroproliferative diseases. Specifically, fibroproliferative diseases include kidney
15 disorders associated with unregulated TGF-β activity and excessive fibrosis including
glomerulonephritis (GN), such as mesangial proliferative GN, immune GN, and
crescertic GN. Other renal conditions include diabetic nephropathy, renal intenstitial
fibrosis, renal fibrosis in transplant patients receiving cyclosporin, and HIV -associated
nephropalhy, Coltagen vascular disorders include progressive systemic clerosis,
20 polymyositis scleroderma, dennatoroyositis, cosinophitic fascritis, morphea, or Those
associated with the occurrence of Raynaud's syndrome. Lung fibroses resulting from
excessive TGF- β activity include adult respiratory distress syndrome, idiopathic
pumonary fihtroais, aud interstitial pulmanary fibrosia often associated with autuimimune
disorders, such as systemic lupus erythematosus and scleroderma, chemical contact, or
25 a lergies Another autoirnmune disorder associated with fibroproliferative characteristics
is rheumatoid arthrrtis.
Eye diseases associated with a fibroproliferative condition include retinal
reattachment surgery accompanying proliferative vitreoretinopathy, cataracl evtraction
with intracular lens implantation, and post glaucoma drainage surgery are associated
30 w TGF-β overproduction.

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Fibrotic diseases associated with TGF-βl overproduction can be divided into
chronic conditions Such as fibrosis of the kidney, lung and liver and more acute
condition such as dennal scarring and restenosis (Chamobentain, J. Cardiovascular Drug
Reviews, 19(4 );329-344). Synthesis and secretion of TGF-β1 by tumor cells can also
5 lead to irnmime suppression such, as seen in patients with aggressive brain or breast
tumors (Artcaga, el al. (1993) J. Clin. Invest. 92:2569-2576). The course of Leishmarnial
infection in mire is drastically altered by TGF-β1 (Barrel-Netlo, et al. (1992) Science
257:545-547). TGF.β1 exacerbated .the disease, whereas TGF-β1 antibodies halted the
progression of the disease in genetically susceptible mice. Genetically resistant mice
10 became susceptible to Leishmanial infection upon administiration of TGF-β1.
The profound effects of TGF-βl on extracellular matrix deposition have been
reviewed (Rocco and Ziyadeh (1991) in Contemporary Issues in Nephrology γ.23,
Hormones, autocoids and the kidney, ed. Jay Stein, Churchill Livingston^ New York
pp391-410; Roberts, et al. (1988) Rec. Prog. Hormone Res. 44: 157-197) and include the
15 stimulation of the synthesis and the inhibition of degradation of extracellular matrix
componenls. Since the structure and filtration properties of the glomenulus are largely
determined by the extracellular matrix composition of the messangium and glomerular
membrane, it is not surprising that TGF-βl has profound effects on the kidney. The
accumulation of mesangial matrix in protiferative glorneruloephritis (Border, et al.
20 (1990) Kidney Int. 37.689-695) and diabetic nephropathy (Mauer, el al. (1984) J. Clin.
Invest. 74:143-1155) are clear and dominant pathological features of the diseases. TGF-
β1 levels are elevated in human diabetic glomerulosclerosis (advanced neuropathy)
(Yamaraoto, et al. (1993) Proc. Natl. Acad. Sci. 90:1814-1818), TGF-β1 is an important
mediator in the genesis of renal fibrosis in a nomber of aninmal models (Phan, et al. (1990)
25 Kidney Int. 37:426; Okuda, et al. (1390) J. clin. Invest. 86:453). Suppression of
experimentally induced glomerulonephritis in rats has been demonstrated by antiserum
against TGF-βl (Border, et al. (1990) Nature 346:371) and by an extracellular matrix
protein, decorin, which can bind TGF-β1 (Border, et al. (1992) Nature 360:361-363).
Too much TGF-βl leads to dermal scar-tissue formation. Neutralizing TGF-β]
30 antibodies injected into the margins of healing wounds in rats have been ahown to inhibit
scaning without interfering with the rale of wound healing or the tensile strength of the

WO 2004/048382 PCT/US2003/032747
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woond (Shah, et al. (1992) Lancet 339:213-314). At the same time there was reduced
angiogcnesis, reduced number of macrophages and monocytes in the waund, and a
reduced amount of disorganized collagen fiber deposition in the scar tissue.
TGF-β1 maybe a factor in the progressive thickening of the arterial wall which
5 results from the proliferation of smooth muscle cells and deposition of extracellular
matrix in the artery after balloon angioplasty. The diameter of the restenosed artery may
be reduced 90% by this thickening, and since most of the reduction in diameter is due to
extracellular matrix rather than smooth muscle cell bodies, it may be possible to open
these vessels to 50% simply by reducing extensive extracellular matrix deposition. In
10 uninjured pig arteries transfected in vivo wittiaTGF-β1 gene,TGF-β1 gene expression
was associated with both extracellular matrix synthesis and hyperplasia (Nabel, et al.
(1993) Proc. Natl. Acad. Sci. USA 90:10759-10763). The TGF-β1 induced hypraplasia
was not as extensive as that induced with PDGF-BB, but the extracellular matrix was
15 associated with FGF-1 (a secreted form of FGF) induced hyperplasia in this gene transfer
pig model (Nabol (1993) Nature 362:844-S46).
There are several types of cancer where TGF-βl produced by the tumor may be
deleterious. MATLyLu rat prostate cancer cells (Steiner and Barrack (1992) Mol.
Endocrinol 6.15-25) and MCF-7 human breast cancer cells (Arteaga, et al. (1993) Cell
20 Growth and Differ. 4:193-201) became more tumorigenic and metaslatic after
transfection with a vector expressing the mouse TGF-β1. TGF-βl has been associated
with angiogenesis, metastasis and poor prognosis in human prostate and advanced gastric
cancer (Wikstrom, P., et al. (1998) Prostate 37: 19-29; Saito, H. et al. (1999) Cancer 86-
1455-1462). In breast cancer, poor prognosis is associated with elevated TGF-β
25 (Dickson, et al. (1987) Proc. Natl. Acad. Sci. USA 84:837-841; Kasid, et al. (1987)
Cancer Res. 47:5733-5738; Daly, et al.(1990) J. Cell Biochem. 43:199-211; Barrett-Lee,
et al. (1990) Br. J Cancer 61:612-617; King, et al. (1989) J. Steroid Biochem. 34:133-138;
Welch, et al. (1990) Proc. Natl. Acad. Sci. USA 87:7678-76S2; Walker, et al. (1992) Eur.
J- Cancer 238:641-644) and induction of TGF-β1 by tamoxifen treatment (Butta, et al.
30 (1992) Cancer Res. 52:4261-4264) has been associated with failure of tamoxifen
treatment for breast cancer (Thompson, et al. (1991) Br. J. Cancer 63:609-614). Anti

WO 2004/048382 PCT/US2003/032747
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TGf-βl antibodies inhibit the growth of MDA-231 human breast cancer cells in athymic
mice (Arteaga, et al. (1993) J. Clin. Invest. 92:2569-2576), a treatment which is
correlated with an increase in spleen natural killer cell activity. CHO cells transfected
with latent TGF-β1 also showed decreased NK activity and increased tumor growth in
5 nude mice (Wallick, et al. (1990) J. Exp. Med. 172:1777-1784). Thus,TGF-β secreted by
breast tumors may cause an endocrine immune suppression. High plasma concentration
of TGF-βl have been shown to indicate poor prognosis for advanced breast cancer
patients (Anscher, et al. (1993) N. Engl. J. Med. 328:1592-1598). Patients with high
circulating TGF-β before high dose- chemotherapy and autologous bone marrow
10 transplantation are al high risk for hepatic veno-occlusive disease (15-50% of all patients
with a mortality rale up to 50%) and idiopathic interstitial pneumonitis (40-60% of all
patients). The implication af these findings is l)that elevated plasma levels of TGF-β1
can be used to identify at risk patients and 2) that reduction of TGF-β1 could decrease the
morbidity and mortality of these common treatments for breast cancer patients.
15 Many malignant cells secrete transforming growth factor-β (TGF-β), a potent
immunosuppressant, suggesting that TGF-β production may represent a significant tumor
escape mechanism from host immunosurveiliance. Establishment of a leukocyte sub-
population with disrupted TGF-β signaling in the tumor-boating host offers a potential
means for immumnotherapy of cancer. A transgentic animal model with disrupted TGF-β
20 signaiing in T cells is capable of eradicating a normally lethal TGF-β over Expressing
lymphoma tumor, EL4 (Gorelik and Flavell, (2001) Nature Medicine 7(10): 1118-1122).
Down regulation of TGF-β secretion in tumor cells results in restoration of
immunogenicity in the host, while T-cell insensitivicy to TGF-β results in accelerated
differentiation and autoimmunity, elements of which may be required in order to combat
25 self-antigen-expressing tumors in a telerized host. The immunosuppressive effects of
TGF-β have also been implicated in a subpopulation of HIV patients with lower than
predicted immune response based on their CD4/CD8 T cell counts (Garba, et al. J.
Immunology (2002) 168: 2247-2254). A TGF-β neutralizing antibody was capable of
reversing the effect in culture, indicating that TGF-β signaling inhibitors may have utility
30 in reversing the immune suppression present in this subset of HIV patients.

WO 2004/048382 PCT/US2003/032747
-16-
During the earliest stages of carcinogenesis, TGF-β1 can act as a potent tumor
suppressor and may mediate the actions of some chemopreventive agents. However, at
some point during the development and progression of malignant neoplasms, tumor cells
appear to escape from TGF-β-dependent growth inhibition in parallel with the appearance
5 of bioaactive TGF-β in the microenvirionment. The dual tumor suppression/tumor
promotion roles of TGF-β have been most clearly elucidated in a transgenic system over
expressing TGF-β in keratinocytes. While the transgenics were more resistant to
formation of benign skin lesions, the rate of metastatic conversion in the transgenics was
dramatically increased (Cui, et al (1996) Cell 86(4):531-42). The production of TGF-β1
10 by malignant cells in primary tumors appears to increase with advancing stages of tumor
progression. Studies in many of the major epithelial cancers suggest that the increased
production of TGF-β by human cancers occurs as a relatively late event during tumor
progression. Further, this tumor-associated TGF-β provides the tumor cells with a
selective advantage and promotes tumor progression. The effects of TGF-β on cell/cell
15 and cell/storma interaction result in a greater propensity for invasion and metastasis
Tumor-associated TGF-β may allow tumor cells to escape from immune surveillance
since it is a potent inhibitor of the clonal expansion of activated lymphocytes. TGF-β has
also been shown to inhibit the production of angiostarin. Cancer therapeutic modalities
such as radiation therapy and chemotherapy induce the production of activated TGF-β in
20 the tumor, thereby selecting outgrowth of malignant cells that are resistant to TGF-β
the development of tumors with enhanced growth and invasiveness. In this situation,
agents targeting TGF-β-mediated signal transduction might be a very effective
therapeutic strategy. The resistance of tumor cells to TGF-β has been shown to negate
25 much of the cytotoxic effects of radiation therapy and chemotherapy and the treatment-
dependent activation of TGF-β in the strom a may even be detrimental as it can make the
micraenvironment more conducive to tumor progression and contributes to tissue damage
leading to fibrosis. The development of a TGF-β signal transduction inhibitors is likely
to benefit the treatment of progressed cancer alone and in combination with other
30 therapies.

WO 2004/048382 PCT/US2003/032747
-17-
The compounds are useful for the treatment of cancer and other disease states
influenced by TGF-β by inhibiting TGF-β in apatient in need thereof by administering
said compound(s) lo said patient. TGF-β would also be useful against atherosclerosis
(T.A. McCaffrey: TGF-βs and TGF-β Receptors in Atherosclerosis: Cytokine and Growth
5 Factor Reviews 2000,11,103-114) and Alzheimer's (Masliah, E.; Ho, G.; Wyss-Coray,
T.: Functional Role of TGF-β in Alzheimer's Disease Microvascular Injury: Lessons from
Transgmic Mice: Neurochemistry International 2001,39, 393-400) diseases.
PHARMACEUTICAL COMPOSITIONS
10
The compositions of the present invention are therapeutically effective amounts of
Ehe TGF-β antagonists, noted above. The composition may be formulated with common
excipients, diluents or carriers, and compressed into tablets, or formulated elixirs or
15 routes. The compounds can be administered transdermally and maybe formulated as
sustained release dosage forms and the like.
The method of treating a human patient according to the present invention
includes administration of the TGF-β antagonists. The TGF-β antagonists are formulated
into formulations which may be administered by the oral and rectal routes, topically,
20 parenterally, e.g., by injection and by continuous or discontinuous intra-arterial infusion,
in the form of, for example, tablets, lozenges, sublingual tablets, sachets, cachets, elixirs,
gels, suspensions, aerosols, ointments, for example, containing from 1 to 10% by weight
of the active compound in a suitable base, soft and hard gelatin capsules, suppositories,
injectable solutions and suspensions in physiologically acceptable media, and sterile
25 packaged powders adsorbed onto a support material for making injectable solutions.
Advantageously for [his purpose, compositions may be provided in dosage unit form,
preferably each dosage unit containing from about 5 to about 500 mg (front about 5 to 50
mg in the case of parenteral or inhalation administration, and from about 25 to 500 mg in
the case of oral or rectal administration) the compounds. Dosages from about 0.5 to about
30 300 mg/kg per day, preferably 0.5 to 20 mg/kg, of active ingredient may be administered
although it will, of course, readily be understood that the amount of the compound
actually to be administered will be determined by a physician, in the light of all the
rcievant circumstances including the condition to be treated, the choice of compound to

X-16038
19
WE CLAIM:
1. A compound according to formula II

5
Formula II
and the pharmaceutically acceptable salts thereof.
10
2. The compound which is 2-(6-methyl-pyridin-2-y1)-3-[6-amido-quinolin-4-
yl)-5,6-dihydro 4H-pyrrolo[ 1,2-b]pyrazole and the pharmaceutically acceptable salts
thereof.
3. A phdrmaceutical formulation comprising a compound according to Claim
15 1 or the pharmaceutically acceptable salt, estes or prodnig thereof togather with a
pharmaceutically acceptable diluent, excipient, or carrier.
4. A compound of claim 1 or a pharmaceutical salt thereof for use in a
method of treatment in the human or animal body,
5. Use of a compound of claim 1 or a pharmaceutical salt thereof for the
20. treatment of cancer.

A compund according to formula II and the plumma
ceutically acceptable salt thereof and the method of treating cancer in a
patient in need thereof by administration of salt compound.