4-(pyrazol-4-yl)-pyrimidines as selective inhibitors of cyclin-dependent kinase 4/6

Article abstract of DOI:10.1021/jm100571n

Identification and structure-guided optimization of a series of 4-(pyrazol-4-yl)-pyrimidines as selective CDK4/6 inhibitors is reported herein. Several potency and selectivity determinants were established based on the X-ray crystallographic analysis of representative compounds bound to monomeric CDK6. Significant selectivity for CDK4/6 over CDK1 and CDK2 was demonstrated with several compounds in both enzymatic and cellular assays.

Full text of DOI:10.1021/jm100571n

7938 J. Med. Chem. 2010, 53, 7938–7957
DOI: 10.1021/jm100571n
4-(Pyrazol-4-yl)-pyrimidines as Selective Inhibitors of Cyclin-Dependent Kinase 4/6
Young Shin Cho,*,† Maria Borland,† Christopher Brain,† Christine H.-T. Chen,† Hong Cheng,† Rajiv Chopra,† Kristy Chung,†
James Groarke,† Guo He,† Ying Hou,† Sunkyu Kim,† Steven Kovats,† Yipin Lu,† Marc O’Reilly,‡ Junqing Shen,† Troy Smith,†
†
Gary Trakshel, Markus Vogtle, Mei Xu, Ming Xu, and Moo Je Sung*
†
†
†
,†
€
†Novartis Institutes for Biomedical Research, 250 Massachusetts Avenue, Cambridge, Massachusetts 02139, United States, and
Astex Therapeutics, 436 Cambridge Science Park, Milton Road, Cambridge, CB4 0QA, United Kingdom
‡
Received May 11, 2010
Identification and structure-guided optimization of a series of 4-(pyrazol-4-yl)-pyrimidines as selective
CDK4/6 inhibitors is reported herein. Several potency and selectivity determinants were established
based on the X-ray crystallographic analysis of representative compounds bound to monomeric CDK6.
Significant selectivity for CDK4/6 over CDK1 and CDK2 was demonstrated with several compounds in
both enzymatic and cellular assays.
Introduction
a functional pRb protein but display genetic alterations
elsewhere).2a Some examples of the latter include: translocation
of D-cyclins in mantle-cell lymphoma and multiple myeloma,
amplification of cyclin D1 in squamous cell esophageal cancer
and breast cancer, amplifications of CDK4 in liposarcoma, and
suppression of p16 in melanoma, nonsmall-cell lung cancer, and
pancreatic cancer.4 In addition to these direct genetic defects,
CDK4/6 kinase activity can also be hyperactivated by mitogen
pathways with activating mutations of their own.5 These muta-
tions are expected to increase cell cycle progression via increas-
ing D-cyclin expression. Taken together, many different cancers
spanning various tissue origins appear to contain activating
aberrations of the CDK4/6 pathway.
Cyclin-dependent kinases (CDKsa) 4 and 6 are highly homol-
ogous proline-dependent serine/threonine kinases that belong
to the CDK enzyme family.1 CDK family members such as
CDK1, 2, and 4/6 function as regulators of cell cycle progres-
sion and check points, whereas CDK8 and 9 are involved in
transcriptional regulation.2 Each CDK enzyme is activated by
binding to a specific cyclin, whose expression is often tied to a
particular phase of the cell cycle. The CDK4/6|D-cyclin enzyme
complex is responsible for the deactivation of retinoblastoma
protein (pRb) via phosphorylation of pRb. This in turn leads to
the release of the E2F transcription factor and activation of
genes required for G1 phase to S phase transition.3
On the basis of clear evidence that numerous cancers hyper-
activate CDK4/6 kinase activity to achieve unchecked prolif-
eration, suppression of CDK4/6 kinase activity has been pro-
posed as an effective way to treat human neoplasia. Indeed,
inhibition of CDK4/6 kinase activity stops the progression of
tumors in various in vivo and in vitro models.6 Furthermore,
genetic knockout experiments involving CDK4/6 have demon-
strated that the inhibition of these kinases in fibroblast cells was
well tolerated due to compensation by CDK1, while CDK1
inhibition led to lethality in all cell systems investigated.7 This
indicates that selective CDK4/6 inhibitors may have a larger
therapeutic window as compared to pan-CDK inhibitors.
While there are several pan-CDK inhibitors in clinical trials,8
a variety of CDK4/6-selective inhibitors are emerging in the
literature.2d,9 Among those, 1 (PD-0332991, Figure 1)10 is note-
worthy, with selectivity achieved over a broad panel of protein
kinases including CDK1 and 2, oral efficacy in several xeno-
graft models, and advancement into clinical trials.11 Recently,
we embarked on efforts toward identification of CDK4/6-selec-
tive and potent inhibitors. Herein, we describe our structure-
based optimization of 4-(pyrazol-4-yl)-pyrimidines (A),12 a hit
series identified via high-throughput screening.
Alteration of the pRb pathway to favor cell proliferation is
one of the hallmarks of a transformed cell phenotype and is
universally observed in cancers. Loss of pRb function that
results in unchecked E2F activity can be achieved either by loss
of pRb itself or by aberrations in other parts of the pRb pathway
that lead to deactivation of the protein (80% of cancers maintain
*To whom correspondence should be addressed. For Y.S.C.: phone,
617-871-3495; fax, 617-871-4081; E-mail, youngshin.cho@novartis. For
M.J.S.: phone, 617-871-7522; fax, 617-871-4081; E-mail, mooje.sung@
novartis.
a Abbreviations: CDK, cyclin-dependent kinase; pRb, retinoblastoma
protein; SNAr, nucleophilic aromatic substitution; dba, dibenzylideneace-
tone;BINAP, 2,20-bis(diphenylphosphino)-1,10-binaphthyl; XANTPHOS,
4,5-bis(diphenylphosphino)-9,90-dimethylzanthene; SEM, 2-(trimethylsilyl)-
ethoxymethyl; LDA, lithium diisopropylamide; LHMDS, lithium
bis(trimethylsilyl)amide; mCPBA, meta-chloroperbenzoic acid; NCS,
N-chlorosuccinimide; DMF, dimethylformamide; NBS, N-bromosuccini-
mide; DDQ, 2,3-dichloro-5,6-dicyanobenzoquinone; TBAI, tetrabutylam-
monium iodide; Boc, t-butoxycarbonyl; THF, tetrahydrofuran; TFAA,
trifluoroacetic anhydride; AIBN, 2,20-azo bisisobutyronitrile; ATP, ade-
nosine-50-triphosphate; PSA, polar surface area; SAR, structure-activity
relationship; ELISA, enzyme-linked immunosorbent assay; FACS, fluor-
escence-activated cell sorting; ALK, anaplastic lymphoma kinase; ERK2,
extracellular signal-related kinase 2; FGFR-4, fibroblast growth factor
receptor-4; GSK 3β, glycogen synthase kinase 3β; JAK1, janus kinase 1;
MAPK2, mitogen-activated protein kinase 2; MAPK 5, mitogen-activated
protein kinase 5; PDGFRR, platelet-derived growth factor receptor R;
PDK1, 3-phosphoinositide dependent kinase 1; PKA, protein kinase A;
PKB R, protein kinase B R; SYK, spleen tyrosine kinase; cMET, mesenchymal-
epithelial transition factor.
Chemistry
The compounds described in this report were prepared via
two different coupling reactions: (1) displacement of a sulfone
r
pubs.acs.org/jmc
Published on Web 11/01/2010
2010 American Chemical Society

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7939
at the C2 position of the pyrimidine core with an amine
side chain,(2) palladium-mediated amination reaction of the
2-chloro-pyrimidine (Scheme 1). These reactions were followed
by removal of protecting groups where necessary.
an ammonia synthon,21 and subsequently deprotected to yield
31 and 32 (Scheme 8).
Scheme 2a
The general synthetic routes to B and C are described in
Scheme 2.13 Acylation of 4-methyl-2-(methylthio)pyrimidine 2
with various esters afforded the ketones 3a-f. The ketones were
then treated with dimethylformamide dimethyl acetal, and the
resulting product was condensed with hydrazine to provide the
pyrazoles 4a-f. Subsequent oxidation of 4a-f gave the sulfones
5a-f. 5b was further converted to 2-chloro-pyrimidine 6 upon
treatment with sulfuryl chloride in acetic acid at 60 °C. Protec-
tion of the pyrazole 6 with a 2-(trimethylsilyl)ethoxymethyl
(SEM) group followed by chlorination with N-chlorosuccinimide
afforded bischloro 4-(pyrazol-4-yl)-pyrimidine 7.
Exploration of other substitutions on the pyrazole is sum-
marized in Schemes 3 and 4. Regioselective bromination of 4b
and 5b to yield 8and 9, respectively, was disclosed previously.13
Oxidation of 8 with m-chloroperbenzoic acid provided 10,
which was converted to 11 via subsequent Stille reaction. The
same Stille reaction conditions were applied to 9 to obtain 12 in
69% yield. 3-Trifluoromethylpyrazole 1314 was protected with
a N,N-dimethylsulfamoyl group and installed on the pyrimi-
dine ring following the procedure developed by Strekowski
et al.15 to give 14 (Scheme 4).
a Reagents and conditions: (a) LDA or LHMDS, THF, 0 °C;
R1CO2Me, 0 °C to room temperature; (b) (i) dimethylformamide
dimethyl acetal, (ii) H2NNH2 H2O, MeOH; (c) mCPBA, CH2Cl2 or
3
oxone, DMF, 90 °C; (d) SO2Cl2, AcOH, 60 °C, 43%; (e) (i) Cs2CO3,
SEMCl, DMF, 73%, (ii) NCS, DMF, 40 °C, 62%.
Scheme 3
Scheme 5 describes a general synthetic route to pyridin-2-
ylamines.10a,16 Displacement of bromine with various piper-
azine derivatives or 4-(dimethylamino)piperidine was followed
by reduction of the nitro group to afford 16-22. Insertion of
a methylene between the pyridine and the piperazine was
achieved by following the reaction steps described in Scheme 6.
Conversion of 5-methyl-pyridin-2-ylamine 23 to its 2,2,2-
trifluoroacetamide and subsequent bromination of the methyl
group17 provided N-(5-bromomethyl-pyridin-2-yl)-2,2,2-
trifluoro-acetamide, which was then reacted with substituted
piperazine derivatives. Methanolysis of the resulting products
gave 24 and 25. A Boc-protected piperidine was installed via
Suzuki reaction of 26 and 27,18 followed by hydrogenation to
give 28 in 42% yield over two steps (Scheme 7). 6-Chloro-
pyridazine 2919 and 5-bromo-pyrazine 3020 were subjected to
palladium-catalyzed cross-coupling with benzophenone imine,
Scheme 4
Figure 1. Selective CDK4/6 inhibitor 1 and general structure of
4-(pyrazol-4-yl)-pyrimidines (A).
Scheme 1a
a Reagents and conditions: (I) R1NH2, heating; (II) R1NH2, Pd2(dba)3, BINAP or XANTPHOS, NaOtBu or Cs2CO3, dioxane, heating.

7940 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
Scheme 5
Table 1. IC50 Values for Compounds in the CDK Enzyme Assaysa,b
Scheme 6
Scheme 7
a See Experimental Section for detailed descriptions of each assay.
b Results are expressed as the mean ( standard deviation of one to two
IC50 determinations. For each determination, concentration-inhibition
curves were obtained in duplicate and then averaged to afford a single
IC50 curve with a g95% confidence interval.
Scheme 8
pyrazole with an isopropyl group (35) led to an improvement
in the overall potency against CDKs as compared to methyl
derivative 34. Subsequent incorporation of a basic amine into
the cyclohexyl ring at the C2 side chain, as exemplified by
compounds 36 and 37, resulted in the enhancement in selec-
tivity for CDK4 over CDK1 and CDK2. In addition to slight
improvement in CDK4 binding affinity, N-methylation of the
piperidine also improved the cellular potency (vide infra) of 37
(non-normalized IC50 0.300 μM, normalized IC50 0.640 μM)
relative to 36 (non-normalized IC50 0.944 μM, normalized IC50
1.615 μM).22
Results and Discussion
With a potent and modestly selective compound37 in hand,
we next probed the binding mode of 4-(pyrazol-4-yl)-pyrimi-
dines to the enzyme. To this end, an X-ray crystal structure
of 37 bound to inactive monomeric CDK6 was obtained
(Figure 2).23 CDK6 shares approximately 70% homology
with CDK4 and demonstrates a comparable SAR profile with
reduced potency (Table 2). The X-ray structure reveals that
37 binds to the CDK6 ATP binding pocket in a compact con-
formation in which the pyrimidine andthe pyrazole are coplanar
and the piperazine moiety packs against the isopropyl group of
the pyrazole. Two hydrogen-bonding interactions are indicated
in the CDK6 hinge region, one between the pyrimidine N1 and
the backbone NH of Val101 and the other between the 2-amino
NH and the Val101 backbone carbonyl. The pyrazole N and NH
form additional polar interactions with the side chains of Lys43
and Asp163, respectively. An edge-to-face aromatic-aromatic
contact is observed between the pyrazole and the kinase gate-
keeper residue, Phe80. The isopropyl substituent has comple-
mentary packing and hydrophobic interactions with Val27,
High-throughput screening of the Novartis compound
collection against the CDK4 enzyme identified a pyrazolyl-
pyrimidine derivative 33 as a promising hit for optimization
(Table 1). Competition binding experiments confirmed that 33
binds competitively and reversibly to the ATP binding site of
CDK4 (data not shown). This compound also demonstrated
selectivity against other non-CDK kinases despite the fact that
it contains an aminopyrimidine moiety, a common element
in many ATP-competitive kinase inhibitors. However, similar
activity against CDK1 and 2 alerted us a potential hurdle of
achieving selectivity within the CDK family. The low molec-
ular weight (MW 243) and calculated properties (cLogP 2.36,
polar surface area (PSA) 66) made compound 33 attractive as a
starting point for a medicinal chemistry campaign.
During the early hit-to-lead optimization, the rapid synthesis
of the core allowed us to quickly explore the structure-activity
relationship (SAR) at the C2 position of the pyrimidine as well
as substitution on the pyrazole moiety. Substitution on the

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7941
Figure 2. X-ray structure of compound 37 bound to CDK6.
Table 2. IC50 Values for Compounds in the CDK Enzyme Assaysa,b
IC50 (μM)
compd
R1
R2
CDK4/cyclin D1
CDK6/cyclin D3
NAc
CDK1/cyclin B
CDK2/cyclin A
38
39
40
41
Me
H
H
0.130 ( 0.020
1.979 ( 0.549
0.540 ( 0.018
0.076 ( 0.009
3.028 ( 0.035
>15
4.187 ( 0.251
>15
Me
H
7.769 ( 0.910
0.737 ( 0.058
0.231 ( 0.018
Br
H
2.507 ( 0.364
1.722 ( 0.214
1.965 ( 0.135
1.037 ( 0.124
Br
a See Experimental Section for detailed descriptions of each assay. b Results are expressed as the mean ( standard deviation of one to two IC50
determinations. For each determination, concentration-inhibition curves were obtained in duplicate and then averaged to afford a single IC50 curve
with a g95% confidence interval. c NA: not available
Gly20, Leu152, Ala162, and the hydrophobic residue of the
Asn150 side chain. The piperidinyl group sits in a solvent
exposed cleft which is mainly hydrophobic but lined with
several polar residues at the edge (Thr107, Asp104, Asp102,
Gln/Glu149).
solvent exposed region of CDK4/6, composed of residues
Asp96(CDK4)/Asp104(CDK6) and Thr99(CDK4)/Thr107-
(CDK6). As previously noted,24 this “selectivity” pocket
has a divergent charge profile due, in part, to the residues
Thr99/107 in CDK4/6 and Lys89 in CDK1/2. A basic sub-
stituent approaching this space could introduce unfavorable
electrostatic repulsion in CDK1/2, thereby boosting selec-
tivity over these closely related kinases.
Compound 42 indeed demonstrated improved selectivity
profile over 37, although CDK4 enzyme potency remained
similar (Table 3). The aniline counterpart of 42 proved to be a
less selective CDK inhibitor, as shown with compound 43.10a
Removal of the piperazine (44) resulted in reduction in
potency, selectivity, and solubility (42 0.9105 mM at pH 6.8,
44 0.0851 mM at pH 6.8). The piperazine moiety proved to be
not only a solubilizing group but also an important selectivity
determinant by reason of deleterious interactions unique to
CDK1/2. A crystal structure of CDK6 in complex with com-
pound 42 confirmed that the key hydrogen-bonding interac-
tions with the hinge region are maintained, and the core of the
pyrazole-pyrimidinescaffold is essentially identical in itsinter-
actions (data not shown).
The overlay of X-ray structures of compounds 37 and 1
bound to CDK6 suggests space available for substitution on
the pyrimidine and the pyrazole for a tighter binding in the
gatekeeper area defined by Phe98 (data not shown). Indeed,
exploiting substitutions at C5 and C6 of the pyridopyrimidine
series resulted in a potent and selective compound (1).9f,10a To
probe this region with the 4-(pyrazol-4-yl)-pyrimidines, we
prepared compounds which contain methyl or bromine at the
C5 of the pyrimidine or the C3 of the pyrazole. This exercise
resulted in reduced potency compared to 37 (Table 2).
Next, we turned our attention to the C2 side chain of the
pyrimidine to further improve enzyme potency and selectivity.
Analysis of reported SAR around19f,10 and X-ray structure of
compound 37 bound to CDK6, as well as molecular modeling
studies, suggested that additional selectivity could be obtained
by replacing the piperidinylamine with a piperazine-substituted
pyridinylamine. Introduction of the pyridine could enhance
selectivity over other kinases via interactions with the side-
chain of hinge residue His 100. An analysis of the sequence
alignment of over 400 kinases (data not shown) indicates that
this His residue is conserved in only a few kinases. The distal
amine of the piperazine could reach into a more polar and
To explore pyrazole substituents, we prepared a small set of
closely related analogues (Table 4). Although the selectivity
for CDK4 appeared to be retained, any change in size of the
substituent R1 (R2=H) diminished the potency for CDK4.
This pattern of substitution was also observed in a series of

7942 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Table 3. IC50 Values for Compounds in the CDK Enzyme Assaysa,b
Cho et al.
Table 4. IC50 Values for Compounds in the CDK Enzyme Assaysa,b
IC50 (μM)
CDK4/
cyclin D1
CDK1/
cyclin B
CDK2/
cyclin A
compd
R1
Me
R2
45
46
47
48
49
50
51
H
H
H
H
H
0.532 ( 0.001
0.037 ( 0.008 5.422 ( 0.175 10.065 ( 1.134
0.089 ( 0.003 15.031 ( 3.621 15.634 ( 4.455
0.187 ( 0.012
0.742 ( 0.019
>15
>15
CF3
cPr
tBu
>15
>15
>15
>15
p-F-Ph
iPr
iPr
Cl 0.011 ( 0.000 6.325 ( 0.549 1.435 ( 0.036
Me 0.274 ( 0.034 >15 >15
a See Experimental Section for detailed descriptions of each assay.
b Results are expressed as the mean ( standard deviation of one to two
IC50 determinations. For each determination, concentration-inhibition
curves were obtained in duplicate and then averaged to afford a single
IC50 curve with a g95% confidence interval.
a See Experimental Section for detailed descriptions of each assay.
bResults are expressed as the mean ( standard deviation of one to two
IC50 determinations. For each determination, concentration-inhibition
curves were obtained in duplicate and then averaged to afford a single
IC50 curve with a g95% confidence interval.
imidazole pyrimidine amides where isopropyl was found to
be optimal for hydrophobic interactions with CDKs.25 Intro-
duction of chloride atom at R2 (50) led to a modest improve-
ment in the CDK4 inhibitory activity. A crystal structure of
compound 50 bound to CDK6 (Figure 3) revealed that the
planes of the pyrimidine and the pyrazole are tilted relative
to each other, with an interplanar angle of ca 25°. The chlorine
substituent of the pyrazole is projected toward the aromatic
plane of Phe80, possibly forming a favorable Cl-π interac-
tion.26 Chlorination of the pyrazole also resulted in improved
metabolic stability in rat liver microsomes (hepatic extraction
ratio 67% for 42 and 36% for 50). As seen earlier, 3-isopropyl-
5-methyl-pyrazole of 51 led to a reduction in potency in CDK4,
suggesting that replacement of a hydrogen with a methyl may
interrupt a favorable edge-to-face aromatic-aromatic interac-
tion observed in the compound 42.
With the pyrazole substituents fixed as R1 = isopropyl and
R2 = chloride, different heteroaromatic rings and various basic
solubilizing groups were applied to the pyrimidine C2 side
chain in an attempt to further improve selectivity over CDK1
and 2 (Table 5). Pyridazine (52) and pyrazine (53) were not
equivalent to pyridine in terms of CDK4 potency and selec-
tivity. Replacement of a piperazine with a piperidine (54) and
substitution on the terminal amine of the piperazine (55 and
56) was well tolerated; however, no impact on selectivity was
seen. Compounds in which steric bulk was introduced around
the terminal amine (57-60) demonstrated only a slight loss in
CDK1/2 activity. When the piperazine moiety was further
extended into the solvent-exposed region, overall loss of CDK
activities was observed, as shown with compounds 61 and 62.
The best potency and selectivity profile was demonstrated
with compound 63, wherein the 4-(dimethylamino)piperidine
moiety is speculated to make a favorable polar interaction
with Thr107 of CDK4/6 and an unfavorable electrostatic
repulsion with Lys89 in CDK1/2. Finally, we tested whether
a trifluoromethyl group could act as a surrogate for Cl on the
Figure 3. Overlay of compounds 50 (carbon atoms, cyan; chlorine,
green) and 37 (carbon atoms, purple) bound to the active site of
CDK6 (carbon atoms, yellow).
pyrazole ring. Compound 64 showed an 8-fold decrease of
CDK4 potency as compared to 63; however, this decrease was
less dramatic than that observed in the case of compounds 51
vs 50, which suggests that the CF3-substituted pyrazole holds
some advantage relative to the methyl-substituted in terms of
its interaction with CDK4.
Five representive compounds with a range of potency and
selectivity for CDK4/6, as well as structural diversity, were
chosen for evaluation in a cell-based assay measuring phosphor-
ylation levels of pRb at the Ser780 site using an enzyme-linked
immunosorbent assay (ELISA) method (Table 6). Jeko-1, a
mantle-cell lymphoma cell line, was selected for use in this
assay due to its known translocation and subsequent over-
expression of cyclin D1.4a Percent inhibition was calculated
at each compound concentration by comparing phosphoryla-
tion levels against the level seen in the vehicle control, and
these inhibitions were used to determine non-normalized
IC50s. A separate ELISA assay was conducted to quantitate

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7943
Table 5. IC50 Values for Compounds in the CDK Enzyme Assaysa,b
a See Experimental Section for detailed descriptions of each assay. b Results are expressed as the mean ( standard deviation of one to two IC50
determinations. For each determination, concentration-inhibition curves were obtained in duplicate and then averaged to afford a single IC50 curve
with a g95% confidence interval.
total pRb levels at each compound concentration and to
compensate for the reduced signal due to the loss of total
pRb protein. The numbers incorporating the results from this
second ELISA are reported as normalized IC50s.

7944 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
Table 6. IC50 Values for Compounds in the CDK4 Cellular Assaysa,b
IC50 (μM)
compd
non-normalized
normalized
42
50
54
63
64
0.451 ( 0.052
0.223 ( 0.016
0.134 ( 0.024
0.324 ( 0.061
0.468 ( 0.107
0.591 ( 0.068
0.467 ( 0.033
0.383 ( 0.069
0.365 ( 0.068
0.557 ( 0.127
a See Experimental Section for detailed descriptions of each assay.
b Results are expressed as the mean ( standard deviation of single IC50
determinations. For each determination, concentration-inhibition
curves were obtained in triplicate and then averaged to afford a single
IC50 curve with a g95% confidence interval.
The same set of compounds was then tested for their ability
to affect individual phases of the cell cycle. To this end, a flow
cytometry analysis was performed, where it was expected that
selective CDK4 inhibitors would afford an exclusive G1 arrest
by inhibition of pRb phosphorylation and subsequent block-
ing of S phase transition (Figure 4). Jeko-1 cells were treated
with compound and then sorted based on their DNA content
using fluorescence-activated cell sorting (FACS). In eachcase,
a dose-related increase in G1 arrest was observed with in-
creasing concentrations of compound. Compound 42 demon-
strated a clean G1 block at 1 μM. However, at 3.3 μM, we
started to see blocks in non-G1 phases of the cell cycle,
suggesting that the ca 200-fold CDK4 vs CDK1/2 selectivity
seen in enzymatic assays with 42 is insufficient for achieving a
comparable cellular selectivity to that of compound 1.10 Com-
pounds 50, 54, and63 imposedaG1blockoncellsat 0.37μM, a
lower concentration than achieved with 42, thereby corrobor-
ating the data from the pRb phosphorylation studies, which
showed lower IC50s for these compounds. Exclusive G1 arrest
was maintained over a wider range of concentrations with these
compounds as compared to 42, which we attribute to their
improved enzyme selectivity for CDK4. A shift of the clean G1
arrest concentration window to higher concentrations with
compound 64 was in agreement with its less potent enzymatic
and cellular IC50s. Compounds 63 and 64 exhibited the most
sustained G1 arrest among five compounds and were tested
further against a selection of serine-threonine and tyrosine
kinases and proved to be selective for CDK4/6 over 35 other
kinases. Several representative data are shown in Table 7.
Conclusions
A series of substituted 4-(pyrazol-4-yl)-pyrimidines were
identified as selective inhibitors of CDK4/6. X-ray crystal-
lographic studies demonstrated a general binding mode of the
compounds bound to CDK6 and guided further optimization
efforts in the series. Our SAR agreed with that of previously
reported CDK inhibitors,10,25 wherein a substituted pyridiny-
lamine side-chainoff of the pyrimidineC2 position introduced
both potency and selectivity and an isopropyl-substituted pyr-
azole at C3 afforded further improvements in potency. Addi-
tional substitution of the pyrazole with chlorine was found to
enhance potency and selectivity in both enzymatic and cellular
settings. A robust G1 cell cycle block of Jeko-1 cells was demon-
strated with several compounds, suggesting that the enzymatic
selectivity of CDK4 over CDK1/2 could be translated to a
specific cellular phenotype that mirrored the genetic inhibition
of CDK4/6. Selective compounds in the cell cycle flow cytometry
analyses, 63 and 64, were evaluated against a panel of protein
kinases and exhibited broad selectivity. Further studies on these
compounds will be reported in due course.
Figure 4. Cell cycle flow cytometry analyses.

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7945
Table 7. Selected Inhibitory Activity of Compounds 63 and 64 against a
Panel of Kinasesa
were run for 2 h at 22 °C. Quenching and detection was carried
out following the protocols for the peptide substrate provided
by the vendor.
IC50 (μM)
CDK4 Cellular Assays. The pRb expressing Jeko-1 mantle cell
lymphoma cell line was grown in complete media consisting of
RPMI1640 (Gibco catalog no. 22400-071), 20% FBS (Gibco
catalog no. 10082-131), 2 mM L-glutamine (Gibco catalog no.
25030-081), and 1% penicillin/streptomycin (Gibco catalog no.
15140-133). Jeko-1 cells were seeded in Biocoat Cell Environ-
ment poly-D-lysine 96-well tissue culture plates (Becton Dick-
inson catalog no. 356461) at 20000 cells/well in 100 μL final
volume of complete media. Cells were allowed to adhere over-
night. Compounds were prepared as 10 mM stock solution in
DMSO and diluted to a concentration of 110 μM in complete
media in a 96-well tissue culture plate and then serially diluted
4-fold, allowing a titration curve of 7 points with a final con-
centration of 26 nM. Ten μL of the dilution were then transferred
to the cell culture plate, resulting in a final concentration range of
10 μM to 2 nM. The incubation was carried out at 37 °C with 5%
CO2. All compounds were tested in triplicates at each concentra-
tion. Following compound incubation, the media was removed
and the cells were lysed in 35 μL of lysis buffer, consisting of
50 mM Tris.Cl, pH 7.2, 120 mM NaCl, 1 mM EDTA, 6 mM
EGTA, 1% NP-40, complete protease inhibitor cocktail (Roche,
catalog no. 11836170001) and a protease inhibitor cocktail from
Calbiochem (catalog no. 524525). The plates were placed at 4 °C
with vigorous shaking for 5 min to lyse the cells. The resulting
lysates contained approximately 1 μg/μL of protein.
The 4H1 total pRb antibody from Cell Signaling Technology
(catalog no. 9309) was added to clear MaxiSorp plates (Nunc
catalog no. 442404) at a concentration of 50 ng per well in 50 μL
of Dulbecco’s phosphate buffered saline (DPBS) (Gibco catalog
no. 14190-144). Plates were incubated overnight at 4 °C with
rocking. After a 250 μL wash with TBST (Teknova catalog no.
T9501) and blot-drying, 250 μL Superblock (Pierce catalog no.
37535) was added to each well. After shaking for 10 min, the
Superblock solution was replaced with fresh Superblock and
plates were incubated on a shaker for an additional 50 min.
After blocking, 30 μL of Jeko-1 cell lysate, containing approxi-
mately 10 μg of total protein, were added to wells in triplicate.
Twenty μL of PBS (Gibco catalog no. 10010-023) containing
10% Superblock (Pierce catalog no. 37535) were added to each
well for a final reaction volume of 50 μL. Plates were then sealed
with Uniseal plate sealers (Whatman catalog no. 7704-0007) and
incubated for 2 h at room temperature on a shaker. Plates were
washed with 3 ꢀ 250 μL TBST. Fifty μL of a 1:1000 dilution of
antiphospho Rb Ser780 from Cell Signaling (catalog no. 9307) in
PBS/10% Superblock were added, and the plate was incubated
on a shaker for 1 h at room temperature. For all incubation
steps, plates were covered with Uniseal plate sealers. Following
incubation, plates were washed with 3 ꢀ 250 μL TBST. Next,
50 μL of a 1:2500 dilution of donkey-antirabbit HRP (Promega
catalog no. W401B) in PBS/10% Superblock were added, and
plates were incubated for 30 min at room temperature on a shaker.
Plates were again washed as described above. Finally, 50 μL of
Ultra TMB ELISA (Pierce catalog no. 34028) were added and
plates incubated, unsealed, 5-15 min in the dark, until blue color
developed. After incubation, 50 μL of 2 M sulfuric acid were
added to plates to top the reaction, and absorbance was deter-
mined on a SpectraMax (Molecular Devices, Sunnydale, CA)
within 15 min at 450 nm. All washes were performed using a Bio-
Tek plate washer.
protein kinase
63
64
anaplastic lymphoma kinase (ALK)
extracellular signal-related kinase 2 (ERK2)
fibroblast growth factor receptor-4 (FGFR-4)
glycogen synthase kinase (GSK3β)
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
9.6
janus kinase 1 (JAK1)
>10
>10
>10
>10
>10
mitogen-activated protein kinase 2 (MAPK2)
mitogen-activated protein kinase 5 (MAPK5)
platelet-derived growth factor receptor R (PDGFRR)
3-phosphoinositide dependent kinase 1 (PDK1)
protein kinase A (PKA)
6.8
protein kinase B (PKBR)
spleen tyrosine kinase (SYK)
mesenchymal-epithelial transition factor (cMET)
a Results are from single IC50 determinations. For each determina-
tion, concentration-inhibition curves were obtained in triplicate and
then averaged to afford a single IC50 curve with a g95% confidence
interval.
>10
>10
>10
Experimental Section
CDK Enzyme Assays. Human CDK4/cyclin D1 was expressed
in Sf21 cells via baculovirus infection. An assay for monitoring
CDK4/cyclin D1-catalyzed phosphorylation of pRb at the
Ser780 site was performed using TR-FRET in a 384-well format
and was used for IC50 determination and kinetic analysis. The
reaction was carried out in a 30 μL volume containing 0.25 nM
CDK4/cyclin D1, 150 nM biotin-pRb (773-924), 3 μM ATP,
and 1.3% DMSO (or compound in DMSO) in the assay buffer
(50 mM HEPES-Na, pH 7.5; 5 mM MgCl2, 1 mM DTT, 0.02%
Tween-20, and 0.05% BSA). Three μM ATP was added last to
initiate the reaction. The reaction was quenched with 10 μL of
240 mM EDTA-Na (pH 8.0) after 60 min incubation at 22 °C.
The signal was developed by the addition of 40 μL of detection
solution containing 40 nM SA-APC, 143 ng/mL antiphospho-
pRb (S780) antibody, and 2 nM Eu-W1024 antirabbit IgG anti-
body in the detection buffer (50 mM HEPES-Na, pH 7.5, 60 mM
EDTA-Na, pH 8.0, 0.05% BSA, and 0.1% Triton X-100). After
60 min incubation in the dark, the plate was read on Envision
(Perkin-Elmer 2102-0010).
Human CDK6/cyclin D3 was purchased from Carna Bio-
sciences Inc. (catalog no. 04-107). An assay for monitoring CDK6/
cyclin D3-catalyzed phosphorylation of pRb at the Ser780 site
was performed using TR-FRET in a 384-well format and was
used for IC50 determination and kinetic analysis. The reaction
was carried out in a 30 μL volume containing 0.20 nM CDK6/
cyclin D3, 200 nM biotin-pRb (773-924), 3 μM ATP, and 1.3%
DMSO (or compound in DMSO) in the assay buffer (50 mM
HEPES-Na, pH 7.5; 5 mM MgCl2, 1 mM DTT, 0.02% Tween-20,
and 0.05% BSA). Three μM ATP was added last to initiate the
reaction. The reaction was quenched with 10 μL of 120 mM
EDTA-Na (pH 8.0) after 120 min incubation at 22 °C. The signal
was developed by the addition of 40 μL of detection solution
containing 40 nM SA-APC, 143 ng/mL antiphospho-pRb (S780)
antibody, and 2 nM Eu-W1024 antirabbit IgG antibody in the
detection buffer (50 mM HEPES-Na, pH 7.5, 60 mM EDTA-Na,
pH 8.0, 0.05% BSA, and 0.1% Triton X-100). After 60 min
incubation in the dark, the plate was read on Envision (Perkin-
Elmer 2102-0010).
The inhibition of human CDK1/cyclin B and human CDK2/
cyclin A (catalog no. 14-450 and 14-448, respectively, Millipore)
was monitored using IMAP-FP assays (Molecular Devices, CA)
by following the phosphorylation of Tamra-Histone H1-derived
peptide (catalog no. R7385, Molecular Devices). The final reac-
tion volume was 20 μL and contained 0.25 nM CDK1/cyclin B
or 0.3 nM CDK2/cyclin A, 100 nM Tamra-H1, and 20 μM ATP
in 1ꢀ reaction buffer (R8139, Molecular Devices). The reactions
The Total Rb ELISA kit (Invitrogen catalog no. KHO0011)
was used to determine the levels of total pRb. This kit uses wells
precoated with a proprietary total pRb antibody for capture. All
reagents listed, with the exception of cell lysate, were included in
the kit. The nature of the antibodies used for capture and detec-
tion was labeled as proprietary and not disclosed. Ten μg of cell
lysate was loaded into the wells and volume adjusted to 50 μL
with standard dilution buffer. Plates were sealed with film

7946 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
included in the kit and incubated for 2 h at room temperature
on a shaker. Plates are then manually washed three times with
250 μL wash buffer. Fifty μL of proprietary primary antibody
(preconjugated to biotin) was added to wells and incubated for
1 h at room temperature on a shaker. Then plates were again
washed as noted above. The secondary antibody (HRP precon-
jugated to Streptavidin) was diluted 1:100 in streptavidin-HRP
diluent buffer, and 50 μL was added to each well. Plates were
then incubated for 30 min. Afterward, plates were washed four
times with buffer as outlined above. Finally, 50 μL of stabilized
Chromogen was added per well and plates were incubated for
15 min, at which point, 50 μL of stop solution was added. Plates
were then read on a Spectramax at 450 nm.
Upon quantitation of the pRb phosphorylation (p-pRb) levels,
% inhibition values were derived for each concentration tested
and used to determine 50% inhibitory concentrations (IC50) for a
particular compound (non-normalized). The total pRb levels
werethenusedtoadjust the p-pRb % inhibitionvaluestoaccount
for any loss of signal due to the absence of the pRb protein itself,
and the IC50 values obtained from the adjusted % inhibitions
represent normalized cellular p-pRb IC50.
Fluorescence Activated Cell Sorting (FACS). Analysis of
cellular DNA content by propidium iodide (PI) staining was used
to determine cells that were in G0/G1, S, or G2/M phase. The media
containing Jeko-1 cells was transferred to a fresh V-bottom 96-well
polypropylene plate (Nunc catalog no. 249944). Next, cells were
spun at 1000g for 10 min and the media was gently removed using
pipet. Then 100 μL of a hypotonic lysis buffer (0.1% sodium citrate
(Sigma catalog no. S-4641), 0.1% Triton X-100 (MP Biomedicals
catalog no. 807423), 25 μg/mL PI (MP Biomedicals catalog no.
195458), and 10 μg/mL RNase (Roche catalog no. 1 579 681) were
added to the cells, and the whole cocktail was incubated at room
temperature in the dark for 1 h. If necessary, cells were stored at this
step overnight at 4 °C. Finally, the cells were subjected to DNA
content analysis using the BD LSR II System and FACSDiva,
version 5.0.1 (BD biosciences, Franklin Lakes, NJ, USA). The
data was analyzed using the ModFit LT 3.1.
ammonium formate with a 2 min run, 3 μL injection through an
Inertisil C8 3 cm ꢀ 5 mm ꢀ 3 μm.
All tested compounds were g95% purity as determined by an
Agilent 1100 HPLC system and one of the following methods,
unless explicitly noted. Method 1 (at both 214 and 254 nm): used
an Inertisil ODS3 3 μm 3.0 mm ꢀ 100 mm C18 column at the
flow rate of 1.0 mL/min, with a gradient of 5-95% acetonitrile/
water 5 mM ammonia formate over 7.75 min. Method 2 (at both
214 and 254 nm): used an Inertisil ODS3 3 μm 3.0 mm ꢀ 100 mm
C18 column at the flow rate of 1.0 mL/min, with a gradient of
5-95% acetonitrile/water with 0.1% TFA over 7.75 min.
Method 3 (at 214 nm): used an Inertsil ODS3 3 μm 3.0 mmm ꢀ
100 mm C18 column at the flow rate of 1.5 mL/min, with
a gradient of 10-95% acetonitrile/water with 0.1% TFA over
15 min. Method 4 (at both 215 and 254 nm): used a Nova-Pak
4 μm 3.9 mm ꢀ 150 mm C18 column at the flow rate of 2.0 mL/min,
with a gradient of 10-90% acetonitrile/water with 0.1% TFA
over 5.0 min. Method 5 (at both 214 and 254 nm): used an
Inertisil ODS3 100 mm ꢀ 3 mm C18 column at the flow rate of
1.0 mL/min, with a gradient of 5-95% acetonitrile/water with
0.1% formic acid over 7.75 min. LC/ESI-MS data were recorded
using a Waters LCT Premier mass spectrometer with dual
electrospray ionization source and Agilent 1100 liquid chroma-
tograph. The resolution of the MS system was approximately
12000 (fwhm definition). HPLC separation was performed at
1.0 mL/min flow rate with the gradient from 10% to 95% in
2.5 min. Ammonia formate (10 mM) was used as the modifier
additive in the Aqueous phase. Sulfadimethoxine (Sigma; pro-
tonated molecule m/z 311.0814) was used as a reference and
acquired through the LockSpray channel every third scan.
3-Methyl-1-(2-methylsulfanyl-pyrimidin-4-yl)-butan-2-one (3b).
To a cooled (-20 °C) solution of diisopropylamine (1746 g,
2.0 equiv) in THF (10 L) was added a solution of nBuLi in THF
(1.6 M, 8L, 1.5equiv) dropwiseover3 h. The reactionmixture was
warmed to 0 °C and treated with a solution of 4-methyl-2-meth-
ylsulfanyl-pyrimidine (2, 1211 g, 8.63 mol) in THF (1 L) dropwise.
The resulting solution was stirred for 15 min and treated with a
solution of isobutyric acid methyl ester (1038 mL, 1.05 equiv) in
DMF-THF (5 L, 4:1 = v/v) dropwise. The resulting mixture was
warmed to room temperature and stirred overnight. The reaction
mixture was treated with 1 N aqueous solution of HCl (17 L) and
extracted with tert-butyl methyl ether (TBME) (3 ꢀ 10 L). The
combined organics were washed with water (3 ꢀ 5 L), brine (5 L),
dried over MgSO4, filtered, and concentrated. Column chroma-
tography (18 kg silica) with hexane/ethylacetate 12:1 gave 993 g
(4.66 mol, 54%) of 3-methyl-1-(2-methylsulfanyl-pyrimidin-4-yl)-
butan-2-one (3b). 1H NMR (600 MHz, DMSO-d6, a 4:1 mixture
of keto- and enol form) δ ppm 13.84 (br s, 0.2 H), 8.54 (d, J = 5.06
Hz, 1.6 H), 8.38 (d, J = 5.10 Hz, 0.2 H), 7.08 (d, J = 5.06 Hz, 1.6
H), 6.85 (d, J = 5.10 Hz, 0.2 H), 5.49 (s, 0.2 H), 3.99 (s, 1.6 H),
2.58-2.80 (m, 0.8 H), 2.31-2.56 (m, 3.2 H), 1.13 (d, J = 7.00 Hz,
1.6 H), 1.05 (d, J = 7.00 Hz, 4.8 H). MS m/z 211.3 (M þ H)þ.
1-Cyclopropyl-2-(2-methylsulfanyl-pyrimidin-4-yl)-ethanone
(3c). To a cooled (0 °C) solution of lithium hexamethyldisila-
zane (1 M, 14.2 mL, 2 equiv) was added neat 4-methyl-2-methyl-
sulfanyl-pyrimidine (2, 1 mL, 7.1 mmol), and the resulting mixture
wasstirredfor5minandthentreatedwithcyclopropylcarboxylate
methyl ester (0.72 mL, 3 equiv). The reaction mixture was stirred
at room temperature for 30 min, treated with water (25 mL) and
extracted into CH2Cl2 (3 ꢀ 50 mL). Combined organics were
washed with brine, dried over Na2SO4, filtered, and concentrated.
Column chromatography (EtOAc/heptanes 10 to 60%) gave
1-cyclopropyl-2-(2-methylsulfanyl-pyrimidin-4-yl)-ethanone (3c,
1.47 g) in quantitative yield. 1H NMR (400 MHz, DMSO-d6) δ
8.56 (d, J = 5.05 Hz, 1 H), 7.13 (d, J = 5.05 Hz, 1 H), 4.05 (s, 2 H),
2.49 (s, 3 H), 2.43 (m, 1 H), 0.92 (m, 4 H). MS m/z 209.3(M þ H)þ.
1,1,1-Trifluoro-3-(2-methylsulfanyl-pyrimidin-4-yl)-propan-2-
one (3d). Prepared from 4-methyl-2-methylsulfanyl-pyrimidine (2)
and methyltrifluoroacetate following the procedure described for 3c
in 60% yield. MS m/z 237.1 (M þ H)þ.
Crystallography. An equimolar complex of a compound and
CDK6 at 5 mg/mL in buffer (25 mM Tris, 300 mM NaCl, 1 mM
TCEP, pH 7.5, 20% glycerol) was crystallized using the hanging-
drop vapor diffusion method. The well contained 100 mM MES
pH 6.0, 25-50 mM NH4NO3, and 6-16% PEG3350. Crystals
were frozen using glycerol as a cryoprotectant, and data were
collected at beamline 17ID of the Argonne Photo Source (Chicago,
IL) using an ADSC-Q210 detector. Data were processed using the
HKL200 package.27 Using CDK2 as the start model, structures
were solved employing the software suites from CCP428 and
CNX29 and deposited to the PDB (PDB ID: 3NUP, 3NUX).
Chemistry. All solvents employed were commercially avail-
able “anhydrous” grade, and reagents were used as received
unless otherwise noted. A Biotage Initiator Sixty system was
used for microwave heating. Flash column chromatography was
performed on either an Analogix Intelliflash 280 using Si 50
˚
columns (32-63 μm, 230-400 mesh, 60 A) or on a Biotage
˚
SP1 system (32-63 μm particle size, KP-Sil, 60 A pore size).
Preparative high pressure liquid chromatography (HPLC) was
performed using a Waters 2525 pump with 2487 dual wave-
length detector and 2767 Sample manager. Columns were
Waters C18 OBD 5 μm, either 50 mm ꢀ 100 mm Xbridge or
30 mm ꢀ 100 mm Sunfire. Systems were run with either a
5-95% or 10-90% ACN/H2O gradient with either a 0.1%
TFA or 0.1% NH4OH modifier. NMR spectra were recorded
on Bruker AV400 (Avance 400 MHz) or AV500 (Avance
500 MHz) instruments. Analytical LC-MS was conducted
using an Agilent 1100 series with UV detection at 214 and
254 nm and an electrospray mode (ESI) coupled with a Waters
ZQ single quad mass detector. One of two methods was used:
method A, 5-95% ACN/H2O with 5 mM ammonium formate
with a 2 min run, 3 μL injection through an Inertisil C8 3 cm ꢀ
5 mm ꢀ 3 μm; method B, 20-95% ACN/H2O with 10 mM

Article
3,3-Dimethyl-1-(2-methylsulfanyl-pyrimidin-4-yl)-butan-2-one
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7947
2-Methylsulfanyl-4-(3-trifluoromethyl-1H-pyrazol-4-yl)-pyri-
midine (4d). A mixture of 1,1,1-trifluoro-3-(2-methylsulfanyl-
pyrimidin-4-yl)-propan-2-one (3d, 1.13 g, 4.8 mmol), dimethyl-
formamide dimethylacetal (2.3 mL, 2 equiv), and acetic acid
(600 μL, 2 equiv) in THF (15 mL) was stirred at room tempera-
ture for 18 h and treated with 55% hydrazine hydrate (40 mL).
After stirring additional 4 h, the reaction mixture was concen-
trated in vacuo. The residue was dissolved in CH2Cl2 (50 mL),
washed with saturated aqueous sodium bicarbonate (2 ꢀ 20 mL),
water (2 ꢀ 20 mL), and brine (30 mL). The organic phase was
dried over Na2SO4, filtered, and concentrated. Column chroma-
tography (EtOAc/heptanes 10-60%) gave 2-methylsulfanyl-
4-(3-trifluoromethyl-1H-pyrazol-4-yl)-pyrimidine (4d, 450 mg)
(3e). Prepared from 4-methyl-2-methylsulfanyl-pyrimidine (2)
and trimethylacetylchloride following the procedure described
for 3c in quantitative yield. 1H NMR (400 MHz, DMSO-d6) δ
8.54 (d, J = 5.05 Hz, 1 H), 7.07 (d, J = 5.05 Hz, 1 H), 4.05 (s,
2H), 2.48 (s, 3H), 1.16 (s, 9H). MS m/z 225.3 (M þ H)þ.
1-(4-Fluoro-phenyl)-2-(2-methylsulfanyl-pyrimidin-4-yl)-etha-
none (3f). Prepared from 4-methyl-2-methylsulfanyl-pyrimidine
(2) and methyl 4-fluorobenzoate following the procedure de-
scribed for 3c and used as a crude mixture for the next reaction.
MS m/z 263.2 (M þ H)þ.
4-(3-Methyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine (4a).
To a cooled (0 °C) solution of lithium hexamethyldisilazane (1 M,
14.2 mL, 2 euiv) was added neat 4-methyl-2-methylsulfanyl-pyr-
imidine (1 mL, 7.1 mmol), and the resulting mixture was stirred
for 5 min and then treated with anhydrous ethyl acetate (10 mL).
The reaction mixture was stirred for 30 min at room temperature
and concentrated in vacuo. The residue was dissolved in metha-
nol (2 mL) and treated with dimethylformamide dimethylacetal
(10 mL). The resulting mixture was heated at 100 °C for 1 h and
cooled to room temperature. The mixture was then treated with
55% hydrazine hydrate (2 mL) and stirred for 18 h. The reaction
mixture was concentrated in vacuo, and the residue was redis-
solved in CH2Cl2 (50 mL), washed with saturated aqueous
sodium bicarbonate (2 ꢀ 20 mL), water (2 ꢀ 20 mL), and brine
(30 mL). The organic phase was dried over Na2SO4, filtered, and
concentrated. Column chromatography (EtOAc/heptanes 10 to
60%) gave 4-(3-methyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyri-
midine (4a, 126 mg) in 19% yield over three steps. 1H NMR (400
MHz, DMSO-d6) δ 13 (broad doublet tautomers, 1 H), 8.47 (d,
J = 5.43 Hz, 1 H), 7.39 (d, J = 5.43 Hz, 1 H), 3.31 (s, 3 H), 2.53 (s,
3 H). MS m/z 207.3 (M þ H)þ.
1
in 40% yield. H NMR (400 MHz, DMSO-d6) δ 8.78 (s, 1 H),
8.61 (d, J = 5.18 Hz, 1 H), 7.47 (d, J = 5.18Hz, 1 H), 2.53 (s, 3H).
MS m/z 261.2 (M þ H)þ.
4-(3-tert-Butyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine
(4e). Prepared from 3,3-dimethyl-1-(2-methylsulfanyl-pyrimidin-
4-yl)-butan-2-one (3e) following the procedure described for 4c in
42% yield. MS m/z 249.3 (M þ H)þ.
4-[3-(4-Fluoro-phenyl)-1H-pyrazol-4-yl]-2-methylsulfanyl-pyri-
midine (4f). Prepared from 1-(4-fluoro-phenyl)-2-(2-methylsulfanyl-
pyrimidin-4-yl)-ethanone (3f) following the procedure described for
4c in 56% yield over three steps. 1H NMR (400 MHz, DMSO-d6)
δ 13.47 (br d tautomers, 1 H), 8.5 (br s tautomer 1, 0.5 H), 8.45
(d, J = 5.31 Hz, 1 H), 8.19 (br s tautomer 2, 0.5 H), 7.56 (br s, 2 H),
7.32 (m, 1 H), 7.25 (m, 1 H), 7.14 (m, 1 H), 2.23 (s, 3 H). MS m/z
287.3 (M þ H)þ.
2-Methanesulfonyl-4-(3-methyl-1H-pyrazol-4-yl)-pyrimidine (5a).
To a solution of 4-(3-methyl-1H-pyrazol-4-yl)-2-methylsulfanyl-
pyrimidine (4a) (110 mg, 0.5 mmol) in DMF (6 mL) was added
solid oxone (770 mg, 2.5 equiv), and the resulting mixture was
heated at 90 °C for 1 h. After cooling to room temperature, the
reaction mixture was treated with water (30 mL) and extracted with
CH2Cl2 (3 ꢀ 30 mL). Combined organics were washed with brine,
dried over Na2SO4, filtered, concentrated, and used for the next
reaction without purification. MS m/z 239.3 (M þ H)þ.
4-(3-Isopropyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine
(4b). A mixture of dimethylformamide dimethylacetal (1.15 L,
2.0 equiv) and 3-methyl-1-(2-methylsulfanyl-pyrimidin-4-yl)-
butan-2-one (3b, 890 g, 4.23 mol) in DMF (1 L) was heated at 90 °Cfor
2 h. The reaction mixture was then concentrated in vacuo and
dried under high vacuum overnight. The residue was triturated
with 100 mL of hexane. The solid was collected and dried under
high-vacuum to provide 1-dimethylamino-4-methyl-2-(2-methyl-
sulfanyl-pyrimidin-4-yl)-pent-1-en-3-one (583 g) in 52% yield as
red crystals. 1H NMR (600 MHz, DMSO-d6) δ ppm 8.38 (d, J =
5.23 Hz, 1 H), 7.66 (s, 1 H), 6.99 (d, J = 5.23 Hz, 1 H), 2.92-3.12
(sept, J = 6.80 Hz 1 H), 2.55-2.95 (br s, 6H), 2.42 (s, 3 H), 0.95
(d, J = 6.80 Hz, 6 H). MS m/z 266.2 (M þ H)þ.
4-(3-Isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine
(5b). To a cooled (0 °C) solution of 4-(3-isopropyl-1H-pyrazol-4-
yl)-2-methylsulfanyl-pyrimidine (4b, 530 g, 2.26 mol) in CH2Cl2
(5 L) was addedmCPBA (70%, 1.4kg, 2.5equiv) in100 g portions
to keep the internal temperature below 10 °C. The suspension was
then slowly warmed to room temperature and stirred for another
2 h. The reaction mixture was treated with saturated aqueous
NaHCO3 solution (1.1 L), and the phases were separated. The
aqueous phase was extracted CH2Cl2 (3 ꢀ 2 L). The combined
organics were washed with water (1 L), brine (0.5 L), dried over
MgSO4, filtered, and concentrated. Column chromatography
(1.5 kg silica) with hexane/ethylacetate 2:3 gave 375 g (1.4 mol,
62%) of 4-(3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyri-
To 1-dimethylamino-4-methyl-2-(2-methylsulfanyl-pyrimidin-
4-yl)-pent-1-en-3-one (580 g, 2.18 mol) in a 10 L reactor was added
aqueous hydrazine hydrate (24%, 2.5 L, 5.5 equiv) at 0 °C. The
resulting yellow suspension was slowly warmed to room tempera-
tureand stirred for 1 h. The reaction mixturewas extracted EtOAc
(3 ꢀ 2.5 L). Organics were washed with water (2 ꢀ 1 L), brine
(0.5 L), dried over MgSO4, filtered, and concentrated to give 4-(3-
isopropyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine (4b, 511 g)
in quantitative yield as red crystals. 1HNMR(600MHz, DMSO-d6)
δ ppm 13.03 (br s, 1H), 8.44 (d, J = 5.47 Hz, 1 H), 7.38 (d, J = 5.47
Hz, 1H), 3.95(brs, 1H), 2.48(s, 3H), 1.25(brs, 6H). MSm/z235.3
(M þ H)þ.
1
midine (5b) as beige crystals. H NMR (600 MHz, DMSO-d6) δ
13.23 (br s, 1H), 8.86 (d, J = 5.41 Hz, 1 H), 7.99 (d, J = 5.41 Hz,
1 H), 3.89 (br s, 1 H), 3.40 (s, 3 H), 1.29 (br s, 6 H). MS m/z 267.1
(M þ H)þ.
4-(3-Cyclopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine
(5c). Prepared from 4-(3-cyclopropyl-1H-pyrazol-4-yl)-2-methyl-
sulfanyl-pyrimidine (4c) following the procedure described for 5a:
MS m/z 265.3 (M þ H)þ.
4-(3-Cyclopropyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine
(4c). A mixture of 1-cyclopropyl-2-(2-methylsulfanyl-pyrimidin-
4-yl)-ethanone (3c, 1.47 g, 7.1 mmol) and dimethylformamide
dimethylacetal (10 mL) in methanol (2 mL) was heated at 100 °C
for 1.5 h. After cooling to room temperature, the reation mixture
was treated with 55% hydrazine hydrate (3 mL) and stirred for
18 h. The mixture was concentrated, and the residue was dissolved
in CH2Cl2 (50 mL), washed with saturated aqueous sodium bicar-
bonate (2 ꢀ 20 mL), water (2 ꢀ 20 mL), and brine (30 mL). The
organic phase was dried over Na2SO4, filtered, and concentrated.
Column chromatography (EtOAc/heptanes 10-60%) provided
4-(3-cyclopropyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine
(4c, 1.2 g) in 73% yield. MS m/z 233.3 (M þ H)þ.
2-Methanesulfonyl-4-(3-trifluoromethyl-1H-pyrazol-4-yl)-pyrimi-
dine (5d). Prepared from 2-methylsulfanyl-4-(3-trifluoromethyl-
1H-pyrazol-4-yl)-pyrimidine (4d) following the procedure de-
scribed for 5a in 71% yield: MS m/z 293.1 (M þ H)þ.
4-(3-tert-Butyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine
(5e). Prepared from 4-(3-tert-butyl-1H-pyrazol-4-yl)-2-methyl-
sulfanyl-pyrimidine (4e) following the procedure described for
5a: MS m/z 281.3 (M þ H)þ.
4-[3-(4-Fluoro-phenyl)-1H-pyrazol-4-yl]-2-methanesulfonyl-
pyrimidine (5f). Prepared from 4-[3-(4-fluoro-phenyl)-1H-pyrazol-
4-yl]-2-methylsulfanyl-pyrimidine (4f) following the procedure
described for 5a: MS m/z 319.2 (M þ H)þ.

7948 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
2-Chloro-4-(3-isopropyl-1H-pyrazol-4-yl)-pyrimidine (6). To a
suspension of 4-(3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-
pyrimidine (5b, 15.2 g, 56.9 mmol) in acetic acid (15 mL) was
added sulfuryl chloride (20 mL, 4.2 equiv), and the resulting
mixture was heated at 60 °C for 7 h. After cooling to room tem-
perature, the reaction mixture was concentrated to remove acetic
acid. The residue was diluted with EtOAc (300 mL) and washed
with water (100 mL), 20% aqueous Na2S2O3 (100 mL), and satu-
rated aqueous NaHCO3 (2 ꢀ 100 mL). The organic layer was
dried over MgSO4, filtered, and concentrated in vacuo. Column
chromatography (EtOAc/heptanes 10-60%) gave2-chloro-4-(3-
isopropyl-1H-pyrazol-4-yl)-pyrimidine (6, 5.5 g) in 43% yield.
1H NMR (400 MHz, CD3OD) δ 8.36 (d, J = 6.0 Hz, 1 H), 8.04
(s, 1 H), 7.48 (d, J = 6.0 Hz, 1 H), 3.82 (q, J = 7.0 Hz, 1 H), 1.24
(d, J = 7.0 Hz, 6 H). MS m/z 223.3 (M þ H)þ.
4-(5-Bromo-3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-
pyrimidine (9). To a solution of 4-(3-isopropyl-1H-pyrazol-4-yl)-2-
methanesulfonyl-pyrimidine (5b, 3.0 g, 11 mmol) in DMF (90 mL)
was added N-bromosuccinimide (2.0 g, 1 equiv) at 0 °C. The mix-
ture was stirred at room temperature for 2 d. The mixture was
treated with water and extracted with EtOAc. The organic extracts
were washed with water, dried over Na2SO4, filtered, and con-
centrated under reduced pressure. Thecrudeproduct was purified
by column chromatography (32% EtOAc/pet. ether) to give 4-(5-
bromo-3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimi-
dine (9, 2.4 g) as a white solid in 61% yield. 1H NMR (400 MHz,
CDCl3) δ 8.81 (d, J = 5.5 Hz, 1 H), 8.09 (d, J = 5.5 Hz, 1 H), 3.81
(m, 1 H), 3.31 (s, 3 H), 1.31 (d, J = 7.0 Hz, 6 H). MS m/z 347.3
(M þ H)þ.
5-Bromo-4-(3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-
pyrimidine (10). To a solution of 5-bromo-4-(3-isopropyl-1H-
pyrazol-4-yl)-2-methylsulfanyl-pyrimidine (8, 157 mg, 0.5 mmol)
in CH2Cl2 (2 mL) was added mCPBA (336 mg, 77%, 0.54 mmol)
0 °C. The mixture was warmed to room temperature and stirred
for 2 h. The reaction mixture was quenched with 20% Na2S2O3
aqueous solution. The mixturewas stirred for several minutesand
then basified with saturated NaHCO3 aqueous solution. The two
phases were separated, and the aqueous layer was extracted with
CH2Cl2. The combined organic extracts were dried over Na2SO4,
filtered, andconcentrated under reduced pressure. The crude pro-
duct was purified by column chromatography (EtOAc:heptane =
10:90 to 100:0) to give 5-bromo-4-(3-isopropyl-1H-pyrazol-4-yl)-
2-methanesulfonyl-pyrimidine (10, 160mg) as a light-yellow solid
in 93% yield. 1H NMR (400 MHz, CDCl3) δ 8.89 (s, 1 H), 8.55
(s, 1 H), 3.86 (ddd, J = 13.68, 7.03, 6.90 Hz, 1 H), 3.37 (s, 3 H),
1.42 (d, J = 7.03 Hz, 6 H). MS m/z 346.8 (M þ H)þ.
A Mixture of 2-Chloro-4-[5-chloro-3-isopropyl-1-(2-trimethyl-
silanyl-ethoxymethyl)-1H-pyrazol-4-yl]-pyrimidine and 2-Chloro-
4-[3-chloro-5-isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-
pyrazol-4-yl]-pyrimidine (7). To a solution of 2-chloro-4-(3-iso-
propyl-1H-pyrazol-4-yl)-pyrimidine (6, 4.3 g, 19 mmol) in DMF
(28 mL) was added Cs2CO3 (13.99 g, 2.2 equiv), and the resulting
mixture was stirred for 45 min at room temperature. The reaction
mixture was treated with SEMCl (8.0 mL, 2.3 equiv) and stirred
at room temperature for 90 min. The reaction mixture was diluted
with EtOAc (150 mL), washed with 4% aqueous NaCl (150 mL),
dried over MgSO4, filtered, and concentrated in vacuo. Column
chromatography (EtOAc/heptanes 5-30%) gave a mixture of
2-chloro-4-[3-isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-
pyrazol-4-yl]-pyrimidine and 2-chloro-4-[5-isopropyl-1-(2-trimeth-
ylsilanyl-ethoxymethyl)-1H-pyrazol-4-yl]-pyrimidine (5.0 g) in
73% yield. 1H NMR (400 MHz, CDCl3, ∼9:1 mixture of tauto-
mers, only major peaks analyzed) δ 8.51 (d, J = 5.3 Hz, 1 H), 8.13
(s, 1 H), 7.31 (d, J = 5.3Hz, 1 H), 5.42 (s, 2 H), 3.64 (t, J = 8.3 Hz,
2 H), 3.61 (m, 1H), 1.37 (d, J = 6.8 Hz, 6 H), 0.94 (t, J = 8.2 Hz, 2
H). 0.00 (s, 9 H). MS m/z 353.3 (M þ H)þ.
4-(3-Isopropyl-1H-pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyri-
midine (11). To a degassed solution of 5-bromo-4-(3-isopropyl-1H-
pyrazol-4-yl)-2-methylsulfanyl-pyrimidine (10, 100 mg, 0.29 mmol)
and Pd(PPh3)4 (66.9 mg, 0.058 mmol) in DMF (2 mL) was added
tetramethyltin (155 mg, 0.87 mmol) by syringe at room tempera-
ture. The mixture was heated at 100 °C for 16 h. After cooling to
room temperature, the mixture was filtered through a pad of Celite
and rinsed with CH2Cl2. The filtrate was washed with water, dried
over Na2SO4, filtered, and concentrated under reduced pressure.
The crude product was purified by silica gel column chromatog-
raphy (MeOH:CH2Cl2 = 1:99 to 8:92) to give 4-(3-isopropyl-1H-
pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyrimidine (11, 70 mg)
as a white solid in 86% yield. 1H NMR (400 MHz, CDCl3) δ 8.66
(s, 1 H), 7.94 (s,1 H), 3.81 (q, J =7.07, 1 H), 3.33 (s, 3H), 2.54 (s, 3
H), 1.37 (d, J = 7.07 Hz, 6 H). MS m/z 281.3 (M þ H)þ.
4-(3-Isopropyl-1H-pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyri-
midine (12). To a degassed solution of 5-bromo-4-(3-isopropyl-1H-
pyrazol-4-yl)-2-methylsulfonyl-pyrimidine (9, 100 mg, 0.29 mmol)
and Pd(PPh3)4 (66.9 mg, 0.058 mmol) in DMF (2 mL) was added
tetramethyltin (0.11 mL, 0.78 mmol) by syringe at room tem-
perature. The mixture was heated at 100°C for 16h. After cooling
to room temperature, the mixture was filtered through a pad of
Celiteand rinsed withEtOAc. The filtratewas washedwithwater,
dried over Na2SO4, filtered, and concentrated under reduced
pressure. The crude product was purified by column chromatog-
raphy (EtOAc:heptane = 10:90 to 100:0) to give 4-(3-isopropyl-
1H-pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyrimidine as a light-
yellow solid (12, 56 mg) in 69% yield. 1H NMR (400MHz, CDCl3)
δ 8.63 (d, J = 5.05 Hz, 1 H), 7.55 (d, J = 5.05 Hz, 1 H), 3.69 (ddd,
J = 13.77, 7.07, 6.96 Hz 1 H), 3.39 (s, 3 H), 2.58 (s, 3 H), 1.39 (d,
J = 7.07 Hz, 6 H). MS m/z 281.1 (M þ H)þ.
To a solution of a mixture of 2-chloro-4-[3-isopropyl-1-(2-
trimethylsilanyl-ethoxymethyl)-1H-pyrazol-4-yl]-pyrimidine and
2-chloro-4-[5-isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-
pyrazol-4-yl]-pyrimidine (3.1 g, 8.7 mmol) in DMF (15 mL) was
added N-chlorosuccinicimide (3.53 g, 3.0 equiv), and the resulting
mixture was stirred at 40 °C for 3 h. The reaction mixture was
cooled to room temperature, diluted with EtOAc (400 mL), washed
with 20% aqueous Na2S2O3 (80 mL), saturated aqueous NaHCO3
(80 mL), and 4% aqueous NaCl (100 mL), dried over MgSO4,
filtered, and concentrated in vacuo. Column chromatography
(EtOAc/heptanes 5-30%) gave a mixture of 2-chloro-4-[5-
chloro-3-isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-
pyrazol-4-yl]-pyrimidine and 2-chloro-4-[3-chloro-5-isopropyl-
1-(2-trimethylsilanyl-ethoxymethyl)-1H-pyrazol-4-yl]-pyrimidine
(7, 2.1 g) in 62% yield. 1H NMR (400 MHz, CDCl3, ∼19:1 mixture
of tautomers, only major peaks analyzed) δ 8.61 (d, J = 5.3 Hz,
1 H), 7.62 (d, J = 5.3 Hz, 1 H), 5.50 (s, 2 H), 3.68 (m, 2 H), 3.64
(m, 1 H), 1.30 (d, J = 6.9 Hz, 6 H), 0.94 (m, 2 H), 0.01 (s, 9 H). MS
m/z 387.4 (M þ H)þ.
5-Bromo-4-(3-isopropyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyr-
imidine (8). To a solution of 4-(3-isopropyl-1H-pyrazol-4-yl)-2-
methylsulfanyl-pyrimidine (4b, 0.50 g, 2.1 mmol) in DMF (5 mL)
was added N-bromosuccinicimide (0.37 g, 1.0 equiv) at 0 °C. The
mixture was stirred at room temperature for 2 h. The reaction
mixture was quenched with 20% Na2S2O3 aqueous solution. The
mixture was stirred for several min and then basified with satu-
rated NaHCO3 aqueous solution. The two phases were separated,
and the aqueous layer was extracted with CH2Cl2. The organic
extracts were dried over Na2SO4, filtered, and concentrated under
reduced pressure. The crude product was purified by column
chromatography (EtOAc/pet. ether) to give 5-bromo-4-(3-isopro-
pyl-1H-pyrazol-4-yl)-2-methylsulfanyl-pyrimidine (8, 0.28 g) as
a yellow solid in 42% yield. 1H NMR (400 MHz, CDCl3) δ 8.53 (s,
1 H), 8.24 (s, 1 H), 3.59-3.75 (m, 1 H), 2.49 (s, 3 H), 1.29 (d, J =
7.03 Hz, 6 H). MS m/z 315.3 (M þ H)þ.
4-(2-Chloro-pyrimidin-4-yl)-5-isopropyl-3-trifluoromethyl-pyra-
zole-1-sulfonic Acid Dimethylamide (14). To a cooled (0 °C) solu-
tion of 4-bromo-5-isopropyl-3-trifluoromethyl-1H-pyrazole (13,
1.52 g, 5.91 mmol) inTHF (40mL) was added NaH (60%, 0.478 g,
4.0 equiv) and the resulting mixture was stirred at room tempera-
ture for 1 h. The reaction mixture was treated with N,N-dimethyl-
sulfamoyl chloride (1.2 mL, 3.9 equiv) and stirred at room tem-
perature for 20 h. The reaction mixture was diluted with EtOAc

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7949
(150 mL), washed with saturated aqueous NaCl (2 ꢀ 50 mL),
dried over MgSO4, filtered, and concentrated in vacuo. Column
chromatography (EtOAc/heptanes 0 to 8%) gave a mixture of
4-bromo-5-isopropyl-3-trifluoromethyl-pyrazole-1-sulfonic acid
dimethylamide and 4-bromo-3-isopropyl-5-trifluoromethyl-pyra-
zole-1-sulfonic acid dimethylamide (1.04 g) in 48% yield: MS m/z
366.0 (M þ H)þ.
(R)-4-(6-Amino-pyridin-3-yl)-2-methyl-piperazine-1-carboxylic
Acid tert-Butyl Ester (18). To a solution of 5-bromo-2-nitropyr-
idine (15, 4.0 g, 19.7 mmol) in DMSO (40 mL) were added R-(-)-
2-methyl-piperazine (3.0 g, 29.8 mmol), tetrabutylammonium
iodide (0.42 g, 1.2 mmol), and potassium carbonate (4.1 g, 29.7
mmol). The reaction mixture was heated at 80 °C for 16 h. The
reaction mixture was poured into ice-water (150 mL) and then
extracted with chloroform. The extracts were washed with water,
dried over Na2SO4, and concentrated under reduced pressure. Puri-
fication by column chromatography (1:9 methanol/chloroform)
gave (R)-3-methyl-1-(6-nitro-pyridin-3-yl)-piperazine (4.3 g, 98%)
as a solid. 1H NMR (400 MHz, CDCl3) δ 8.18 (d, J = 9.2 Hz, 1 H),
8.13 (d, J = 3.1 Hz, 1 H), 7.20 (dd, J = 9.2, 3.1 Hz, 1 H), 3.78-3.74
(m, 2 H), 3.15 (m, 1 H), 3.04-3.00 (m, 3 H), 2.68 (m, 1 H), 1.19 (d,
J = 6.3 Hz, 3 H). MS m/z 222.8 (M þ H)þ.
A solution of a mixture of 4-bromo-5-isopropyl-3-trifluoro-
methyl-pyrazole-1-sulfonic acid dimethylamide and 4-bromo-3-
isopropyl-5-trifluoromethyl-pyrazole-1-sulfonic acid dimethy-
lamide (1.04 g, 2.84 mmol) in THF (10 mL) was added to a
cooled (-78 °C) solution of nBuLi (2.5 M, 1.5 mL, 1.3 equiv) in
THF (5 mL) over 5 min. The resulting mixture was treated with a
suspension of 2-chloropyrimidine (0.348 g, 1.07 equiv) in THF
(2 mL) quickly and stirred at -30 °C for 40 min and 0 °C for
20 min. The reaction mixture was quenched with a solution of
acetic acid (0.17 mL, 1.04 equiv) and water (0.02 mL, 0.39 equiv)
in THF (0.5 mL) and treated with DDQ (0.681 g, 1.05 equiv).
The resulting mixture was stirred at room temperature for 5 min,
cooled to 0 °C, treated with a cold aqueous solution of NaOH
(1 N, 3.2 mL, 1.1 equiv), and stirred at 0 °C for 5 min. The reac-
tion mixture was diluted with EtOAc (150 mL), washed with
saturated aqueous NaHCO3 (70mL) andbrine(70mL), driedover
MgSO4, filtered, and concentrated in vacuo. Column chromatography
(EtOAc/heptanes 3 to 20%) gave 4-(2-chloro-pyrimidin-4-yl)-
5-isopropyl-3-trifluoromethyl-pyrazole-1-sulfonic acid dimethyl-
amide (14, 0.686 g) in 57% yield and 4-(2-chloro-pyrimidin-
4-yl)-3-isopropyl-5-trifluoromethyl-pyrazole-1-sulfonic acid
To a solution of (R)-3-methyl-1-(6-nitro-pyridin-3-yl)-piper-
azine (2.4 g, 10.8 mmol) in CH2Cl2 (30 mL) were added di-tert-
butyl dicarbonate (2.8 g, 12.9 mmol) and triethylamine (3.0 mL,
21.6 mmol). The reaction mixture was heated to reflux for 4 h
and concentrated under reduced pressure. The residue was dis-
solved in EtOAc (50 mL). The solution was washed with water
and brine. The organic layer was dried over Na2SO4 and con-
centrated in vacuo. Purification by column chromatography (1:9
methanol/chloroform) gave (R)-2-methyl-4-(6-nitro-pyridin-3-
yl)-piperazine-1-carboxylic acid tert-butyl ester (2.5 g, 71.8%)
as a solid. 1H NMR (400 MHz, CDCl3) δ 8.19 (d, J = 12.4 Hz,
1 H), 8.10 (d, J = 4.0 Hz, 1 H), 7.16 (dd, J = 12.4 Hz, 4.0 Hz,
1 H), 4.40 (br s, 1 H), 3.99 (m, 1 H), 3.75 (m, 1 H), 3.63 (m, 1 H),
3.43-3.34 (m, 2 H), 3.25 (m, 1 H), 1.49 (s, 9 H), 1.25 (d, J = 4.0
Hz, 3 H). MS m/z 323.0 (M þ H)þ.
1
dimethylamide (0.093 mg) in 8% yield. H NMR (400 MHz,
CD3OD, 14 only) δ 8.78 (d, J = 5.0 Hz, 1 H), 7.50 (d, J = 5.0 Hz,
1 H), 3.78 (m, 1 H), 3.19 (s, 6 H), 1.24(d, J = 7.2 Hz, 6 H). MSm/z
398.1 (M þ H)þ. The structurewas determinedby2DNMRanal-
ysis including HSQC and HMBC.
A solution of (R)-2-methyl-4-(6-nitro-pyridin-3-yl)-piperazine-
1-carboxylic acid tert-butyl ester (2.5 g, 7.7 mmol) in methanol
(100 mL) was hydrogenated in the presence of 10% Pd/C (0.2 g)
using an H2 balloon. After 16 h, the reaction mixture was filtered
through a pad of Celite and rinsed with methanol (2ꢀ15 mL).
The filtrate was concentrated and purified by column chromatog-
raphy (1:9 methanol/chloroform) to afford the title compound
1
(18, 1.86 g, 82%) as a solid. H NMR (400 MHz, DMSO-d6) δ
7.59 (d, J = 2.8 Hz, 1 H), 7.14 (dd, J = 8.8, 2.9 Hz, 1 H), 6.39 (d,
J = 8.8 Hz, 1 H), 4.16 (br s, 1 H), 3.76 (m, 1 H), 3.23 (m, 1 H),
3.13-3.06 (m, 2 H), 2.60 (m, 1 H), 2.42 (m, 1 H); 1.40 (s, 9 H) 1.21
(d, J = 6.7 Hz, 3 H). MS m/z 293.5(M þ H)þ.
(S)-4-(6-Amino-pyridin-3-yl)-2-methyl-piperazine-1-carboxylic
Acid tert-Butyl Ester (19). Prepared from S-(þ)-2-methyl-piper-
1
azine following the procedure for 18 in 50% yield. H NMR
(400 MHz, DMSO-d6) δ 7.59 (d, J = 2.8 Hz, 1 H), 7.14 (dd, J =
8.8, 2.9 Hz, 1 H), 6.39 (d, J = 8.8 Hz, 1 H), 4.16 (br s, 1 H), 3.76
(m, 1 H), 3.23 (m, 1 H), 3.13-3.06 (m, 2 H), 2.60 (m, 1 H), 2.42
(m, 1 H); 1.40 (s, 9 H) 1.21 (d, J = 6.7 Hz, 3 H). MS m/z 293.0
(M þ H)þ.
5-(4-Methyl-piperazin-1-yl)-pyridin-2-ylamine (17). To a solu-
tion of 5-bromo-2-nitro-pyridine (15, 15.0 g, 73.9 mmol) in DMSO
(150 mL) were added 1-methyl-piperazine (12.7 g, 126.1 mmol),
tetrabutylammonium iodide (1.6 g, 4.3 mmol), and potassium
carbonate (15.3 g, 110.7 mmol). The reaction mixture was heated
at 80 °C for 5 h. The reaction mixture was poured into ice-water
and then extracted with EtOAc. The combined extracts were washed
with water and brine. The organic layer was dried over Na2SO4,
filtered, and concentrated under reduced pressure. Purification by
column chromatography (1:9 methanol/chloroform) gave 1-methyl-
4-(6-Amino-pyridin-3-yl)-2,2-dimethyl-piperazine-1-carboxylic
Acid tert-Butyl Ester (20). Prepared from 2,2-dimethyl-pipera-
zine following the procedure for 18 in 51% yield. 1H NMR (400
MHz, DMSO-d6) δ 7.54 (d, J = 3.0 Hz, 1 H), 7.09 (dd, J = 8.8,
3.0 Hz, 1 H), 6.39 (d, J = 8.8 Hz, 1 H), 5.32 (br s, 2 H), 3.52-3.49
(m,2H),3.01-2.98(m,2H),2.89(s,2H),1.40(s,9H),1.34(s,6H).
MS m/z 307.0 (M þ H)þ.
1
4-(6-nitro-pyridin-3-yl)-piperazine (16.2 g, 98.6%) as a solid. H
NMR (400 MHz, DMSO-d6) δ 8.25 (d, J=4.0 Hz, 1 H), 8.14 (d, J=
8.0 Hz, 1 H), 7.47 (dd, J = 8.0, 4.0 Hz, 1 H), 3.48 (t, J = 4.0 Hz, 4
H), 2.43 (t, J = 4.0 Hz, 4 H), 2.20 (s, 3 H). MS m/z 222.9 (M þ H)þ.
A solution of 1-methyl-4-(6-nitro-pyridin-3-yl)-piperazine (2.0 g,
9.0 mmol) in methanol (100 mL) was hydrogenated in the presence
of 10% Pd/C (0.2 g) using an H2 balloon. After 16 h, the reaction
mixture was filtered through a pad of Celite and rinsed with
methanol (2 ꢀ 15 mL). The filtrate was concentrated and purified
by column chromatography (1:9 methanol/chloroform) to afford
the title compound (17, 1.5 g, 87%) as a solid. 1H NMR (400 MHz,
D2O) δ 7.64 (dd, J = 9.5, 2.8 Hz, 1 H), 7.40 (d, J = 2.8 Hz, 1 H),
6.83 (d, J = 9.5 Hz, 1 H), 3.50 (br s, 4 H), 3.11-3.05 (m, 4 H), 2.82
(s, 3 H). MS m/z 192.9 (M þ H)þ.
4-(6-Amino-pyridin-3-yl)-2,6-dimethyl-piperazine-1-carboxylic
Acid tert-Butyl Ester (21). Prepared from 2,6-dimethyl-pipera-
zine following the procedure for 18 in 54% yield. 1H NMR (400
MHz, CDCl3) δ 7.78 (d, J = 2.7 Hz, 1 H), 7.17 (dd, J = 8.7, 2.7
Hz, 1 H), 6.50 (d, J = 8.7 Hz, 1 H), 4.24-4.21 (m, 4 H), 3.10 (d,
J = 11.6 Hz, 2 H), 2.81 (dd, J = 11.6, 4.2 Hz, 2 H), 1.50 (s, 9 H),
1.37 (d, J = 6.8 Hz, 6 H). MS m/z 307.0 (M þ H)þ.
N*4*,N*4*-Dimethyl-3,4,5,6-tetrahydro-2H-[1,30]bipyridinyl-
4,60-diamine (22). To a solution of 5-bromo-2-nitropyridine (15,
5.10 g, 24.9 mmol) in MeCN (60 mL) were added 4-dimethyla-
mino piperidine (3.64 g, 1.14 equiv) and iPr2NEt (4.75 mL, 1.09
equiv), andthe resulting mixturewas heatedtorefluxfor 20h. The
reaction mixture was cooled to room temparature and concentrated.

7950 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
Dimethyl-(60-nitro-3,4,5,6-tetrahydro-2H-[1,30]bipyridinyl-4-yl)-
amine (4.89 g) was obtained by trituration in MeCN in 79% yield.
1H NMR (400 MHz, CD3OD) δ 8.23 (d, J = 6.0 Hz, 1 H), 8.21
(s, 1 H), 7.56 (dd, J = 9.3, 3.0 Hz, 1 H), 4.29 (dd, J = 16, 2.5 Hz,
2 H), 3.51 (m, 1 H), 3.12 (m, 2 H), 2.91 (s, 6 H), 2.13-2.37 (m, 2 H),
1.82 (dd, J = 12, 4.1 Hz, 2 H). MS m/z 251.2 (M þ H)þ.
MeOH/chloroform) to give (R)-4-(6-amino-pyridin-3-ylmethyl)-2-
methyl-piperazine-1-carboxylic acid tert-butyl ester (25, 0.72 g,
56%) as a solid. 1H NMR (400 MHz, DMSO-d6) δ 7.75 (s, 1 H),
7.27(d,J= 8.4 Hz, 1 H), 6.39 (d, J= 8.4 Hz, 1 H), 5.80 (s, 2 H), 4.04
(br s, 1 H), 3.65 (m, 1 H), 3.29 (d, J = 13.0 Hz, 1 H), 3.18 (d, J =
13.0 Hz, 1 H), 2.93 (m, 1 H), 2.71 (m, 1 H), 2.51 (m, 1 H), 1.94 (m,
1 H), 1.81 (m, 1 H), 1.38 (s, 9 H), 1.17 (d, J = 7.3 Hz, 3 H). MS m/z
307.2 (M þ H)þ.
A mixture of dimethyl-(60-nitro-3,4,5,6-tetrahydro-2H-[1,30]-
bipyridinyl-4-yl)-amine (3.44 g, 13.6 mmol) and 10% Pd/C (333 mg)
in MeOH-H2O (80 mL, v/v = 1:1) was stirred under H2 (1atm) for
24 h. The reaction mixture was filtered through a pad of Celite
(rinsed with MeOH) and concentrated in vacuo. Trituration of
the residue in MeCN gave N*4*,N*4*-dimethyl-3,4,5,6-tetrahy-
dro-2H-[1,30]bipyridinyl-4,60-diamine (22, 2.554 g) in 85% yield.
1H NMR (400 MHz, CD3OD) δ 7.61 (d, J = 2.8 Hz, 1 H), 7.40
(dd, J = 9.1, 2.9 Hz, 1 H), 6.64 (d, J = 9.0 Hz, 1 H), 3.56 (dd, J =
10.3, 2.1 Hz, 2 H), 3.23 (m, 1 H), 2.87 (s, 6 H), 2.74 (m, 2 H), 2.17
(m, 2 H), 1.86 (m, 2 H). MS m/z 221.3 (M þ H)þ.
6-Amino-30,40,50,60-tetrahydro-20H-[3,40]bipyridinyl-10-carboxylic
Acid tert-Butyl Ester (28). To a mixture of 5-bromo-2-amino-
pyridine (26, 0.67 g, 3.9 mmol) and 4-(4,4,5,5-tetramethyl-[1,3,2]-
dioxaborolan-2-yl)-3,6-dihydro-2H-pyridine-1-carboxylic acid tert-
butyl ester (27, 1.20 g, 3.9 mmol) were added Pd(PPh3)4 (0.45 g, 0.39
mmol) and KF/Al2O3 (3.6 g). The mixture was degassed for 0.5 h
and then heated at 100 °C for 2 h. The reaction mixture was diluted
with EtOAc (100 mL), filtered through a pad of Celite, and the filter
cake was rinsed with EtOAc (2 ꢀ 50 mL). The combined filterates
were dried over Na2SO4, filtered, and concentrated in vacuo. Puri-
fication by column chromatography (1:2 EtOAc/petroleum ether)
provided 6-nitro-30,60-dihydro-20H-[3,40]bipyridinyl-10-carboxylic
(S)-4-(6-Amino-pyridin-3-ylmethyl)-2-methyl-piperazine-1-car-
boxylic Acid tert-Butyl Ester (24). Prepared following the procedure
described for 25 (see below), substituting (R)-2-methyl-piperazine-
1-carboxylic acid tert-butyl ester for (S)-2-methyl-piperazine-
1
acid tert-butyl ester (0.75 g, 70%) as a brownish solid. H NMR
1
(400MHz, DMSO-d6) δ7.97(d, J= 2.4 Hz, 1 H), 7.48 (dd, J=8.7,
2.4 Hz, 1 H), 6.41 (d, J = 8.7 Hz, 1 H), 5.98 (s, 1 H), 5.93 (br s, 2 H),
3.94 (br s, 2 H), 3.51-3.48 (m, 2 H), 2.37 (br s, 2 H), 1.41 (s, 9 H).
MS m/z 276.2 (M þ H)þ.
1-carboxylic acid tert-butyl ester in 28% yield. H NMR (400
MHz, DMSO) δ 7.75 (s, 1 H), 7.27 (d, J = 8.4 Hz, 1 H), 6.39 (d,
J = 8.4 Hz, 1 H), 5.80 (s, 2 H), 4.04 (br s, 1 H), 3.65 (m, 1 H), 3.29
(d, J = 13.0 Hz, 1 H), 3.18 (d, J = 13.0 Hz, 1 H), 2.93 (m, 1 H),
2.71 (m, 1 H), 2.51 (m, 1 H), 1.94 (m, 1 H), 1.81 (m, 1 H), 1.38 (s, 9
H), 1.17 (d, J = 7.3 Hz, 3 H). MS m/z 307.2 (M þ H)þ.
A solution of 6-nitro-30,60-dihydro-20H-[3,40]bipyridinyl-10-
carboxylic acid tert-butyl ester (0.75 g, 2.7 mmol) in EtOH
(20 mL) was hydrogenated in the presence of 10% Pd/C (0.2 g)
using an H2 balloon. After 16 h, the reaction mixture was filtered
through a pad of Celite and rinsed with EtOH (2 ꢀ 10 mL). The
filtrate was concentrated and purified by column chromatog-
raphy to give 6-amino-30,40,50,60-tetrahydro-20H-[3,40]bipyridinyl-
10-carboxylic acid tert-butyl ester (28, 0.45 g, 60%) as a solid. 1H
NMR (400 MHz, MeOD) δ 7.76 (d, J = 2.3 Hz, 1 H), 7.38 (dd,
J = 8.6, 2.3 Hz, 1 H), 6.56 (d, J = 8.6 Hz, 1 H), 4.21-4.18 (m,
2 H), 2.84 (br s, 2 H), 2.60 (m, 1 H), 1.78-1.75 (m, 2 H), 1.58-
1.57(m, 2 H), 1.48 (s, 9 H). MS m/z 278.2 (M þ H)þ.
(R)-4-(6-Amino-pyridin-3-ylmethyl)-2-methyl-piperazine-1-car-
boxylic Acid tert-Butyl Ester (25). To a cooled (0 °C) solution of
2-amino-5-methylpyridine (23, 25.0 g, 231.4 mmol) in CH2Cl2
(250 mL) were added pyridine (181.0 mL) and trifluoroacetic
anhydride (35.7 mL, 254.6 mmol). The reaction mixture was
stirred for 2 h, quenched with water, and extracted with CH2Cl2.
The extract was washed with citric acid solution, water, and brine.
The organic layer was dried over Na2SO4, filtered, and concentrated
under reduced pressure. Purification by column chromatography
(3:7 EtOAc/pet. ether) gave 2,2,2-trifluoro-N-(5-methyl-pyridin-2-
yl)-acetamide (37.0 g, 78%) as a solid. 1H NMR (400 MHz, CDCl3)
δ 9.32 (br s, 1 H), 8.17 (s, 1 H), 8.08 (d, J = 8.4 Hz, 1 H), 7.62 (dd,
J = 8.4 Hz, 1 H), 2.53 (s, 3 H). MS m/z 204.7 (M þ H)þ.
4-(6-Amino-pyridazin-3-yl)-piperazine-1-carboxylic Acid tert-
Butyl Ester (31). To a solution of 4-(6-chloro-pyridazin-3-yl)-
piperazine-1-carboxylic acid tert-butyl ester (29, 2.0 g, 6.7 mmol)
in toluene (30 mL) were added benzophenone imine (1.3 g, 7.3
mmol), Pd2(dba)3 (0.18 g, 0.2 mmol), BINAP (0.37 g, 0.6 mmol),
and sodium tert-butoxide (0.9 g, 0.9 mmol). The reaction mix-
ture was degassed for 0.5 h and then heated at 115 °C for 14 h.
The mixture was cooled to room temperature, diluted with water
(200 mL), and extracted with EtOAc (200 mL ꢀ 3). The extracts were
washed with brine, dried over Na2SO4, filtered, and concentrated
in vacuo. The residue was purified by column chromatography (3:7
EtOAc/petroleum ether) to afford 4-[6-(benzhydrylidene-amino)-
pyridazin-3-yl]-piperazine-1-carboxylic acid tert-butyl ester (1.5 g,
51%) as a pale-yellow solid. 1H NMR (400 MHz, CDCl3) δ
7.92-7.76 (m, 2 H), 7.36-7.18 (m, 8 H), 6.82 (d, J = 9.5 Hz,
1 H), 6.73(d, J =9.5Hz, 1H), 3.53(brs, 8H), 1.49(s, 9H). MSm/z
444.2 (M þ H)þ.
To a solution of 2,2,2-trifluoro-N-(5-methyl-pyridin-2-yl)-acet-
amide (37.0 g, 181.2 mmol) in carbon tetrachloride (500 mL) were
added N-bromosuccinicimide (31.9 g, 179.4 mmol) and AIBN
(3.27 g, 19.9 mmol). The reaction mixture was heated to reflux for
4 h. After cooling to room temperature, the mixture was filtered
through a pad of Celite and the filtrate was concentrated under
reducedpressure togive thecrudebromide (50.0 g, ca98%), which
was used for the next step without further purification.
To a solution of the crude bromide (2.0 g, 10 mmol) in dimethyl-
formamide (40 mL) were added (R)-2-methyl-piperazine-1-car-
boxylic acid tert-butyl ester (4.7 g, 16.7 mmol) and triethylamine
(1.4mL, 10mmol) at0°C. The reaction mixture was stirred for 36 h.
The mixture was diluted with water (150 mL) and extracted with
EtOAc (50 mL ꢀ 3). The combined extracts were washed with brine,
dried over Na2SO4, filtered, and concentrated in vacuo. The residue
was purified by column chromatography (1:10 EtOAc/petroleum
ether) to provide (R)-2-methyl-4-[6-(2,2,2-trifluoroacetylamino)-
pyridin-3-ylmethyl]-piperazine-1-carboxylic acid tert-butyl ester
(1.6 g, 40%) as a solid. 1H NMR (400 MHz, CDCl3) δ 8.80 (br s,
1 H), 8.29 (d, J = 1.8 Hz, 1 H), 8.16 (d, J = 8.5 Hz, 1 H), 7.81 (dd,
J = 8.5, 1.8 Hz, 1 H), 4.21 (br s, 1 H), 3.83 (m, 1 H), 3.55 (d, J =
13.6 Hz, 1 H), 3.42 (d, J = 13.6 Hz, 1 H), 3.11 (m, 1 H), 2.74 (d,
J = 10.6 Hz, 1 H), 2.55 (d, J = 11.1 Hz, 1 H), 2.18 (dd, J = 11.1,
3.9 Hz, 1 H), 2.06 (m, 1 H), 1.48 (s, 9 H), 1.19 (d, J = 8.0 Hz, 3 H).
MS m/z 403.2 (M þ H)þ.
To a solution of 4-[6-(benzhydrylidene-amino)-pyridazin-3-
yl]-piperazine-1-carboxylic acid tert-butyl ester (1.5 g, 3.4 mmol)
in MeOH (14 mL) were added sodium acetate (0.56 g, 6.8 mmol)
and hydroxyamine hydrochloride (0.43 g, 6.1 mmol). The reaction
mixture was stirred for 0.5 h and then concentrated in vacuo. The
residue was purified by column chromatography (1:9 MeOH/
chloroform) to give 4-(6-amino-pyridazin-3-yl)-piperazine-1-car-
boxylic acid tert-butyl ester (31, 0.87 g, 92%) as a yellow solid. 1H
NMR (400 MHz, MeOD) δ 7.30 (d, J = 9.7 Hz, 1 H), 6.97 (dd,
J = 9.7, 2.1 Hz, 1 H), 3.53-3.52 (m, 4 H), 3.37-3.35 (m, 4 H), 1.47
(s, 9 H). MS m/z 280.2 (M þ H)þ.
To a solution of ammonia in MeOH (ca 7 N, 150 mL) was added
(R)-2-methyl-4-[6-(2,2,2-trifluoroacetylamino)-pyridin-3-ylmethyl]-
piperazine-1-carboxylic acid tert-butyl ester (1.6 g, 4.0 mmol). The
mixture was stirred for 24 h and concentrated under reduced pres-
sure. The residue was purified by column chromatography (1:100
50-Amino-2,3,5,6-tetrahydro-[1,20]bipyrazinyl-4-carboxylic Acid
tert-Butyl Ester (32). Prepared from 50-bromo-2,3,5,6-tetrahydro-
[1,20]bipyrazinyl-4-carboxylic acid tert-butyl ester (30) following
the procedure described for 31 in 59% yield. 1H NMR (400 MHz,
DMSO-d6) δ 7.66 (d, J = 1.4 Hz 1 H), 7.57 (d, J = 1.4 Hz, 1 H),

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7951
5.61 (s, 2 H), 3.42-3.40 (m, 4 H), 3.19-3.16 (m, 4 H) 1.41 (s, 9 H).
MS m/z 280.2 (M þ H)þ.
and 1-methyl-piperidin-4-ylamine following the procedure de-
scribed for 37 in 53.2% yield. 1H NMR (400 MHz, CD3OD) δ
8.20 (d, J = 5.0 Hz, 1 H), 6.67 (d, J = 5.0 Hz, 1 H), 3.88 (dd, J =
14.6, 6.6 Hz, 1 H), 3.67 (s, 1 H), 2.90 (d, J = 11.6 Hz, 2 H), 2.42 (s,
3H), 2.31 (s, 3H), 2.19 (t, J = 11.1 Hz, 2 H), 2.04 (d, J = 11.1 Hz,
2 H), 1.64 (d, J = 9.6 Hz, 2 H), 1.31 (d, J = 6.6 Hz, 6 H). Anal.
RP-HPLC tR = 5.71 min (method 1, purity 98.2%/95.1%). HR-MS
m/z (M þ H)þ: measured 315.2285, calcd 315.2297.
Cyclopentyl-[4-(3-methyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-amine
(33). Synthesized from 4-(3-methyl-1H-pyrazol-4-yl)-2-methane-
sulfonyl-pyrimidine (5a) and cyclopentylamine following the pro-
cedure described for 37 (see below) in 46.7% yield. 1H NMR (400
MHz, DMSO-d6) δ12.95 (br s, 1 H), 8.16 (d, J=5.56Hz, 1H), 7.34
(s, 1H),6.88(d, J=5.56Hz, 1H), 4.21(brs, 1H), 2.57(s, 3H), 1.93
(d, J = 6.06 Hz, 2 H), 1.78-1.61 (m, 2 H), 1.61-1.44 (m, 4 H).
Anal. RP-HPLC tR = 6.20 min (method 2, purity 96.30%/90.40%).
HR-MS m/z (M þ H)þ: measured 244.1567, calcd 244.1562.
Cyclohexyl-[4-(3-methyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-amine
(34). Synthesized from 4-(3-methyl-1H-pyrazol-4-yl)-2-methane-
sulfonyl-pyrimidine (5a) and cyclohexylamine following the pro-
cedure described for 37 (see below) in 32.4% yield. 1H NMR (300
MHz, DMSO-d6) δ 12.84 (br s, 1 H), 8.15 (d, J = 6 Hz, 1 H), 7.96
(s, 1 H), 6.78 (d, J = 6 Hz, 1 H), 6.75 (br s, 1 H), 3.72 (m, 1 H), 2.58
(s, 3 H), 1.94 (m, 2 H), 1.71 (m, 2 H), 1.63 (m, 1 H), 1.30 (m, 5 H).
Anal. RP-HPLC tR = 7.25 min (method 2, purity 96.6%/100%).
HR-MS m/z (M þ H)þ: measured 258.1718, calcd 258.1719.
Cyclohexyl-[4-(3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
amine (35). Synthesized from 4-(3-isopropyl-1H-pyrazol-4-yl)-
2-methanesulfonyl-pyrimidine (5b) and cyclohexylamine follow-
ing the procedure described for 37 (see below) in 36.3% yield. 1H
NMR (400 MHz, CDCl3) δ 10.12 (b. s, 1 H), 8.10 (d, J = 5.05 Hz,
1 H), 7.85 (s, 1 H), 6.61 (d, J= 5.05 Hz, 1 H), 5.08 (br s, 1 H), 3.93 (br s,
1 H), 3.77 (ddd, J = 14.40, 10.36, 4.04 Hz, 1 H), 2.00 (dd, J = 12.38,
3.28 Hz, 2 H), 1.71 (ddd, J = 13.26, 3.66, 3.28 Hz, 2 H), 1.58 (dd, J =
9.09, 4.04 Hz, 1 H), 1.30 (d, J = 7.07 Hz, 8 H), 1.24-1.09 (m, 3 H).
Anal. RP-HPLC tR = 7.87 min (method 2, purity 100.00%/100.00%).
HR-MS m/z (M þ H)þ: measured 286.2039, calcd 286.2032.
[5-Bromo-4-(3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(1-
methyl-piperidin-4-yl)-amine (40). Synthesized from 5-bromo-
4-(3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine
(10) and 1-methyl-piperidin-4-ylamine following the procedure
described for 37 in 11.3% yield. 1H NMR (400 MHz, CD3OD) δ
8.22 (s, 1 H), 8.00 (s, 1 H), 3.70 (dd, J = 15.4, 6.3 Hz, 1 H), 3.61
(d, J = 6.6 Hz, 1 H), 2.79 (d, J = 11.6 Hz, 2 H), 2.19 (s, 3H), 2.06
(t, J = 12.1 Hz, 2 H), 1.90 (d, J = 11.6 Hz, 2 H), 1.52 (d, J = 9.1
Hz, 2 H), 1.21 (d, J = 7.1 Hz, 6 H). Anal. RP-HPLC tR = 5.71
min (method 1, purity 88.8%/93.3%). HR-MS m/z (M þ H)þ:
measured 379.1250, calcd 379.1246.
[4-(5-Bromo-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(1-
methyl-piperidin-4-yl)-amine (41). Synthesized from 4-(5-bromo-
3-isopropyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine (9)
and 1-methyl-piperidin-4-ylamine following the procedure de-
scribed for 37 in 66.7% yield. 1H NMR (400 MHz, CD3OD) δ
8.25 (d, J = 5.0 Hz, 1 H), 7.00 (d, J = 5.0 Hz, 1 H), 3.88 (dd, J =
15.2, 6.6 Hz, 1 H), 3.77 (dt, J = 14.2, 7.1 Hz, 1 H), 2.93 (d, J =
12.1 Hz, 2 H), 2.33 (s, 3H), 2.22 (t, J = 11.4 Hz, 2 H), 2.05 (d, J =
10.1 Hz, 2 H), 1.66 (d, J = 9.6 Hz, 2 H), 1.33 (d, J = 7.1 Hz, 6 H).
Anal. RP-HPLC tR = 5.79 min (Method 1, purity 98%/95.2%).
HR-MS m/z (M þ H)þ: measured 379.1245, calcd 379.1246.
4-(3-Isopropyl-1H-pyrazol-4-yl)-N-(5-(piperazin-1-yl)pyridin-
2-yl)pyrimidin-2-amine (42). To a solution of 2-chloro-4-[3-iso-
propyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-pyrazol-4-yl]-
pyrimidine and 2-chloro-4-[5-isopropyl-1-(2-trimethylsilanyl-ethoxy-
methyl)-1H-pyrazol-4-yl]-pyrimidine (a mixture of intermediates
from 6 to 7, 40 mg, 0.113 mmol) in 1,4-dioxane (3 mL) in a sealed
tube were added tert-butyl 4-(6-aminopyridin-3-yl)piperazine-1-
carboxylate (16, 32.2 mg, 1.02 equiv), BINAP (6.97 mg, 0.1 equiv),
NaOtBu (16.3 mg, 1.5 equiv), and Pd2(dba)2 (5.20 mg, 0.05 equiv).
Nitrogen was bubbled into the resulting mixture for 3 min to
degas. The mixture was sealed and heated at 110 °C for 3.5 h. After
cooling to room temperature, the reaction mixture was filtered
through a pad of Celite (rinsed with EtOAc). The filtrate was
washed with water (2 ꢀ 10 mL) and brine (10 mL). The organic
layer was dried over Na2SO4, filtered, and concentrated in vacuo.
Column chromatography (EtOAc/heptanes 0 to 50%) and sub-
sequent HPLC purification gave tert-butyl 4-(6-(4-(3-isopropyl-
1-((2-silylethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)-
pyridin-3-yl)piperazine-1-carboxylate (25 mg) in 37% yield.
1HNMR (400 MHz, CD3OD) δ 8.38 (d, J = 5.3 Hz, 1 H),
8.34 (s, 1 H), 8.11 (d, J = 9.1 Hz, 1 H), 8.01 (d, J = 2.9 Hz, 1 H),
7.51 (dd, J = 9.2, 3.0 Hz, 1 H), 7.07 (d, J = 5.3 Hz, 1 H), 5.44
(s, 2 H), 3.86 (q, J = 6.9 Hz, 1 H), 3.71-3.58 (m, 5 H), 3.52 (t,
J = 6.7 Hz, 1 H), 3.19-3.01 (m, 4 H), 1.51 (s, 9 H), 1.31 (d, J =
7.0 Hz, 6 H), 0.98-0.86 (m, 2 H), 0.0 (s, 9 H). MS m/z 595.6
(M þ H)þ.
[4-(3-Isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-piperidin-4-yl-
amine (36). Synthesized from 4-(3-isopropyl-1H-pyrazol-4-yl)-2-
methanesulfonyl-pyrimidine (5b) and piperidin-4-ylamine follow-
ing the procedure described for 37 (see below) in 38% yield. 1H
NMR (400 MHz, CD3OD) δ 8.06 (d, J = 5.0 Hz, 1 H), 7.91 (s,
1 H), 6.76, (d, J = 5.0 Hz, 1 H), 3.95 (m, 2 H), 3.29 (ddd, J = 12.8,
3.8, 3.7 Hz, 2 H), 2.93-2.87 (m, 2 H), 2.10 (dd, J= 13.9 Hz, 3.3 Hz,
2 H), 1.65 (q, J = 10.6 Hz, 2 H), 1.26 (d, J = 7.1 Hz, 6 H). Anal.
RP-HPLC tR = 9.58 min (method 1, purity 100.00%/100.00%).
HR-MS m/z (M þ H)þ: measured 287.1985, calcd 287.1984.
[4-(3-Isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(1-methyl-
piperidin-4-yl)-amine (37). The mixture of 4-(3-isopropyl-1H-
pyrazol-4-yl)-2-methanesulfonyl-pyrimidine (5b, 30 mg, 0.11 mmol)
and 1-methyl-piperidin-4-ylamine (63 mg, 0.55 mmol) in DMSO
(0.5 mL) was heated at 130 °C for 1 h. The crude product was
purified by prep-HPLC with a gradient of 5-90% acetonitrile/
water with 3% n-propanol to give [4-(3-isopropyl-1H-pyrazol-4-
yl)-pyrimidin-2-yl]-(1-methyl-piperidin-4-yl)-amine (37, 10.9 mg)
in 33% yield. 1H NMR (400 MHz, CD3OD) δ 8.13 (d, J = 5.6
Hz, 1 H), 8.01 (s, 1 H), 6.84 (d, J = 5.6Hz, 1 H), 4.11(s, 1 H), 3.88,
(dd, J = 14.6, 6.6 Hz, 1 H), 2.94 (d, J = 12.1 Hz, 2 H), 2.34 (s, 3
H), 2.22 (t, J = 11.4 Hz, 2 H), 2.06 (d, J = 10.6 Hz, 2 H), 1.66 (q,
J = 9.6 Hz, 2 H), 1.37 (d, J = 6.6 Hz, 6 H); Anal. RP-HPLC tR =
5.16 min (method 1, purity 97.4%/97.8%). HR-MS m/z (M þ H)þ:
measured 301.2142, calcd 301.2141.
[4-(3-Isopropyl-1H-pyrazol-4-yl)-5-methyl-pyrimidin-2-yl]-(1-
methyl-piperidin-4-yl)-amine (38). Synthesized from 4-(3-isopropyl-
1H-pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyrimidine (11) and
1-methyl-piperidin-4-ylamine following the procedure described
To a solution of tert-butyl 4-(6-(4-(3-isopropyl-1-((2-silylethoxy)-
methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)pipera-
zine-1-carboxylate (25 mg, 0.042 mmol) in 1,4-dioxane (1.0 mL) was
added 4 M HCl in dioxane (1.0 mL, 100 equiv). The mixture was
stirred at room temperature for 5 h. The yellow solid was collected
by filtration, washed with cold dioxane (5 mL) and then ether (10
mL), and dried under high vacuum overnight to give 4-(3-isopropyl-
1H-pyrazol-4-yl)-N-(5-(piperazin-1-yl)pyridin-2-yl)pyrimidin-2-
amine as a hydrochloride salt (23 mg) in 91% yield. 1HNMR (600
MHz, CD3OD) δ 8.49 (d, J = 6.0 Hz, 1 H), 8.28 (s, 1 H), 8.04-7.97
(m, 2 H), 7.48 (d, J = 6.0 Hz, 1 H), 7.45 (d, J = 9.9 Hz, 1 H), 4.16
(m, 1 H), 3.51-3.47 (m, 4 H), 3.43-3.40 (m, 4 H), 1.36 (d, J = 6.8
Hz, 6 H). Anal. RP-HPLC tR = 3.30 min (method 5, 100.00%/
100%). HR-MS m/z(Mþ H)þ: measured 364.2238, calcd 364.2250.
1
for 37 in 26.4% yield. H NMR (400 MHz, CD3OD) δ 8.08 (s,
1H), 7.77(brs, 1H),3.85(dd, J= 14.9, 6.3 Hz, 1 H), 3.66 (br s, 1 H),
2.95 (d, J = 12.1 Hz, 2 H), 2.36 (s, 3 H), 2.28 (t, J = 11.4 Hz, 2 H),
2.18 (s, 3 H), 2.03 (d, J = 10.6 Hz, 2 H), 1.63 (d, J = 10.6 Hz, 2 H),
1.29 (d, J = 7.1 Hz, 6 H). Anal. RP-HPLC tR = 5.72 min (method
1, purity 100.0%/100.0%). HR-MS m/z (M þ H)þ: measured
315.2287, calcd 315.2297.
4-(3-Isopropyl-5-methyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(1-
methyl-piperidin-4-yl)-amine (39). Synthesized from 4-(3-isopro-
pyl-1H-pyrazol-4-yl)-5-methyl-2-methylsulfonyl-pyrimidine (12)

7952 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
[4-(3-Isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(4-piperazin-
1-yl-phenyl)-amine (43). To a solution of 2-chloro-4-(3-isopro-
pyl-1H-pyrazol-4-yl)-pyrimidine (6, 55 mg, 0.247 mmol) in
1,4-dioxane (3 mL) in a microwave vial were added 4-(4-amino-
phenyl)piperazine-1-carboxylic acid t-butyl ester (75.4 mg,
1.1 equiv), XANTPHOS (14.3 mg, 0.1 equiv), Cs2CO3 (86.9 mg,
2.0 equiv), and Pd2(dba)2 (11.3 mg, 0.05 equiv). Nitrogen was
bubbled into the resulting mixture for 3 min to degas. The mixture
was sealed and heated in a microwave reactor at 150 °C for 40 min.
After cooling to room temperature, the reaction mixture was fil-
tered through a pad of Celite (rinsed with EtOAc) and washed
with water (2 ꢀ 10 mL) and brine (10 mL). The organic layer was
dried over Na2SO4, filtered, and concentrated in vacuo. Column
chromatography (EtOAc/heptanes 0-50%) and subsequent
HPLC purification gave tert-butyl 4-(4-(4-(3-isopropyl-1H-pyra-
zol-4-yl)pyrimidin-2-ylamino)phenyl)piperazine-1-carboxylate (13.2
mg) in 11% yield. 1H NMR (400 MHz, CD3OD) δ 8.21 (s, 1 H),
8.15 (d, J = 6.3 Hz, 1 H), 7.42 (d, J = 8.8 Hz, 2 H), 7.18 (d, J =
6.2 Hz, 1 H), 7.10 (d, J = 9.0 Hz, 2 H), 3.97 (m, 1 H), 3.63 (br s,
4 H), 3.13-3.26 (m, 4 H), 1.51 (s, 9 H), 1.24 (d, J = 7.0 Hz, 6H).
MS m/z 464.5 (M þ H)þ.
1.89 min (method 3, purity 94%/88%). HR-MS m/z (MþH)þ:
measured 337.1895, calcd 337.1889.
(5-Piperazin-1-yl-pyridin-2-yl)-[4-(3-trifluoromethyl-1H-pyra-
zol-4-yl)-pyrimidin-2-yl]-amine (46). Prepared from 2-methane-
sulfonyl-4-(3-trifluoromethyl-1H-pyrazol-4-yl)-pyrimidine (5d)
and 4-(6-amino-pyridin-3-yl)-piperazine-1-carboxylic acid tert-
butyl ester (16) following the procedure described for 45 in <1%
yield. 1H NMR (400 MHz, DMSO-d6) δ 8.90 (s, 1 H), 8.34 (d,
J = 5.31, 1 H), 8.24 (s, 1 H), 8.19 (d, J = 8.97 Hz, 1 H), 7.94 (d,
J = 2.78 Hz, 1 H), 7.39 (dd, J = 8.97, 2.78 Hz, 1 H), 6.92 (d, J =
5.31 Hz, 1 H), 3.01 (m, 4 H), 2.84 (m, 4 H). Anal. RP-HPLC tR =
3.36 min (method 4, 94%, 89%). HR-MS m/z (M þ H)þ:
measured 391.1618, calcd 391.1607.
[4-(3-Cyclopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(5-pipera-
zin-1-yl-pyridin-2-yl) amine (47). Prepared from 4-(3-cyclopro-
pyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine (5c) and
4-(6-amino-pyridin-3-yl)-piperazine-1-carboxylic acid tert-bu-
tyl ester (16) following the procedure described for 45, except
neutralization, in 3% yield as a trifluoroacetic acid salt. 1H
NMR (400 MHz, DMSO-d6) δ 10.89 (br s, 1 H), 8.93 (s, 2 H),
8.50 (d, J = 5.31 Hz, 1 H), 8.30 (s, 1 H), 8.02 (s, 1 H), 7.84 (d, J =
8.72 Hz, 1 H), 7.76 (d, J = 8.72 Hz, 1 H), 7.43 (d, J = 5.31 Hz,
1 H), 3.37 (m, 4 H), 3.29 (m, 4 H), 2.88 (m, 1H), 1.02 (m, 2 H), 0.91
(m, 2 H). Anal. RP-HPLC tR = 3.10 min (method 5, 87%,87%).
HR-MS m/z (M þ H)þ: measured 363.2060, calcd 363.2046.
[4-(3-tert-Butyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(5-piperazin-
1-yl-pyridin-2-yl)-amine (48). Prepared from 4-(3-tert-butyl-1H-
pyrazol-4-yl)-2-methanesulfonyl-pyrimidine (5e) and 4-(6-amino-
pyridin-3-yl)-piperazine-1-carboxylic acid tert-butyl ester (16)
following the procedure described for 45, except neutralization, as
a trifluoroacetic acid salt in 7% yield. 1H NMR (400 MHz, DMSO-
d6) δ 10.29 (s, 1 H), 8.81 (bs, 1 H), 8.48 (d, J = 5.56 Hz, 1 H), 8.07
(s, 1 H), 8.01 (s, 1 H), 7.85 (d, J = 9.22 Hz, 1 H), 7.78 (d, J = 9.22
Hz, 1 H), 7.24 (d, J = 5.43 Hz, 1 H), 3.36 (m, 4 H), 3.28 (m, 4 H),
1.42 (s, 9 H). Anal. RP-HPLC tR = 3.36 min (method 5, 91%/
90%). HR-MS m/z (M þ H)þ: measured 379.2365, calcd 379.2359.
{4-[3-(4-Fluoro-phenyl)-1H-pyrazol-4-yl]-pyrimidin-2-yl}-(5-pi-
perazin-1-yl-pyridin-2-yl)-amine (49). Prepared from 4-[3-(4-fluoro-
phenyl)-1H-pyrazol-4-yl]-2-methanesulfonyl-pyrimidine (5f) and
4-(6-amino-pyridin-3-yl)-piperazine-1-carboxylic acid tert-butyl
ester (16) following the procedure described for 45 in <1% yield.
1H NMR (400 MHz, DMSO-d6) δ 9.10 (s, 1 H), 8.34 (d, J = 5.18
Hz, 1 H), 8.24 (s, 1 H), 7.91 (d, J = 2.91 Hz, 1 H), 7.59 (dd, J =
8.84, 8.84 Hz, 2 H), 7.57 (d, J = 2.91 Hz, 1 H), 7.25 (dd, J = 8.84,
8.84 Hz, 2 H), 7.02 (dd, J = 8.84, 2.91 Hz, 1 H), 6.81 (d, J = 5.18
Hz, 1 H), 2.97 (m, 4 H); 2.84 (m, 4 H). Anal. RP-HPLC tR = 2.20
min (method 4, purity 97%/93%. HR-MS m/z (M þ H)þ: mea-
sured 417.1965, calcd 417.1951.
[4-(3-Isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(4-piperazin-
1-yl-phenyl)-amine (43) was prepared from tert-butyl 4-(4-(4-(3-
isopropyl-1H-pyrazol-4-yl)pyrimidin-2-ylamino)phenyl)piperazine-
1-carboxylate following the procedure descirbed for the deprotection
reaction of 42 as a hydrochloride salt in quantitative yield.
1H NMR (600 MHz, CD3OD) δ 8.39 (s, 1 H), 8.05 (br s., 1 H),
7.39-7.21 (m, 3 H), 7.13 (d, J = 8.7 Hz, 2 H), 3.79 (br s, 1 H),
3.50-3.41 (m, 4 H), 3.41-3.30 (m, 4 H), 2.58 (s, 6 H). Anal. RP-
HPLC tR = 3.30 min (method 5, 100%/100%). HR-MS m/z (M þ
H)þ: measured 364.2238, calcd 364.2250.
4-(3-Isopropyl-1H-pyrazol-4-yl)-N-(pyridin-2-yl)pyrimidin-2-
amine (44). To a solution of 2-chloro-4-(3-isopropyl-1H-pyra-
zol-4-yl)pyrimidine (6, 100 mg, 0.45 mmol) and pyridine-2-
amine (54.4 mg, 0.49 mmol) in 1,4-dioxane (5 mL) in a sealed
tube were added Xantphos (26 mg, 0.1 equiv), LHMDS 1 M
solution (670 uL, 1.5 equiv), and Pd2(dba)2 (21 mg, 0.05 equiv).
Nitrogen was bubbled into the resulting mixture for 3 min to
degas. The mixture was sealed and heated at 110 °C for 3.5 h.
After cooling to room temperature, the reaction mixture was
filtered through a pad of Celite (rinsed with EtOAc). The filtrate
was washed with water (2 ꢀ 10 mL) and brine (10 mL). The orga-
nic layer was dried over Na2SO4, filtered, and concentrated in
vacuo. Column chromatography (EtOAc/heptanes 0-50%) and
subsequent HPLC purification gave 4-(3-isopropyl-1H-pyrazol-
4-yl)-N-(pyridin-2-yl)pyrimidin-2-amine (44, 15 mg) in 12% yield.
1HNMR (400 MHz, CD3OD) δ 8.50 (d, J = 5.6 Hz, 1 H), 8.33 (d,
J = 5.4 Hz,1 H), 8.15 (br s,1 H), 7.99 (br m, 2 H), 7.32 (d, J = 5.4
Hz, 1 H), 7.19 (d, J = 2.4 Hz, 1 H), 1.69-1.45 (m, 2 H), 1.38 (d,
J = 7.0 Hz, 6 H). Anal. RP-HPLC tR = 2.70 min (method 5,
100%/100%). HR-MS m/z (M þ H)þ: measured 281.1511, calcd
281.1515.
4-{6-[4-(3-Methyl-1H-pyrazol-4-yl)-pyrimidin-2-ylamino]-pyr-
idin-3-yl}-piperazine-1-carboxylic Acid tert-Butyl Ester (45). A mix-
ture of 2-methanesulfonyl-4-(3-methyl-1H-pyrazol-4-yl)-pyrimidine
(5a, 120 mg, 0.5 mmol) and 4-(6-amino-pyridin-3-yl)-piperazine-1-
carboxylic acid tert-butyl ester (16, 211 mg, 1.5 equiv) in toluene
(6 mL) was heated at 120 °C for 16 h, allowing the solvent to boil off
to yield a melt. After cooling to room temperature, the residue was
dissolved in CH2Cl2 (10 mL) and treated with trifluoroacetic acid
(2 mL). After 1 h, the reaction mixture was concentrated and
purified by HPLC. A trifluoroacetic acid salt was then neutralized
using polymer-linked carbonate resin (Stratospheres SPE PL-
HCO3 MP 500 mg cartridges) to yield 5 mg of 4-{6-[4-(3-methyl-
1H-pyrazol-4-yl)-pyrimidin-2-ylamino]-pyridin-3-yl}-pipera-
zine-1-carboxylic acid tert-butyl ester (45) in 3% yield. 1H NMR
(400MHz, DMSO-d6) δ12.94 (br s, 1 H), 9.19 (s, 1 H), 8.37 (d, J =
5.31 Hz, 1 H), 8.08 (d, J = 8.97 Hz, 1 H), 7.96 (d, J = 3.03 Hz,
1 H), 7.41 (dd, J = 8.97, 3.03 Hz, 1 H), 7.06 (d, J = 5.31 Hz, 1 H),
3.01 (m, 4 H), 2.84 (m, 4 H), 2.56 (2.56, 3 H). Anal. RP-HPLC tR =
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(5-
piperazin-1-yl-pyridin-2-yl)-amine (50). tert-Butyl 4-(6-(4-(5-
chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-
4-yl)pyrimidin-2-ylamino)pyridin-3-yl)piperazine-1-carboxylate
was prepared from 7 and 4-(6-amino-pyridin-3-yl)-piperazine-
1-carboxylic acid tert-butyl ester (16) following the procedure des-
cribed for the coupling reaction of 42 in 10.4% yield. 1HNMR(400
MHz, CD3OD) δ 8.50 (d, J = 5.2 Hz, 1 H), 8.15 (d, J = 9.1 Hz,
1 H), 8.01 (d, J = 2.9 Hz, 1 H), 7.48 (dd, J = 9.2 Hz, 1 H), 7.13
(d, J = 5.2 Hz, 1 H), 5.52 (s, 2 H), 3.71 (m, 2 H), 3.19-3.07
(m, 5 H), 3.13(m, 4 H), 1.50 (s, 9 H), 1.25 (d, J = 6.9 Hz, 6 H), 0.93
(m, 2 H), 0.00 (s, 9 H). MS m/z 629.6 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
(5-piperazin-1-yl-pyridin-2-yl)-amine (50) was prepared from
tert-butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)pipera-
zine-1-carboxylate following the procedure described for the depro-
tection reaction of 42 as a hydrochloride salt in 65% yield. 1HNMR
(400 MHz, CD3OD) δ 8.59 (d, J = 5.7 Hz, 1 H), 8.07 (dd, J = 9.5,
3.0 Hz, 1 H), 7.86 (d, J = 2.6 Hz, 1 H), 7.56 (d, J = 5.7 Hz, 1 H),
7.44 (d, J = 9.5 Hz, 1 H), 3.97-3.73 (m, 1 H), 3.47-3.39 (m, 4 H),
3.38-3.30 (m, 4 H), 1.25 (d, J = 7.1 Hz, 6 H). Anal. RP-HPLC

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7953
tR = 3.45 min (method 5, purity 93.38%/95.94%). HR-MS m/z
(M þ H)þ: measured 339.1821, calcd 399.1812.
1H NMR(400 MHz, CD3OD) δ9.22 (d, J = 5.3 Hz, 1 H), 8.95 (d,
J = 8.6 Hz, 1 H), 8.84 (d, J = 2.3 Hz, 1 H), 8.34 (dd, J = 8.7, 2.4
Hz, 1 H), 7.85 (d, J = 5.2Hz, 1 H), 6.20 (s, 2 H), 4.91 (d, J = 13.01
Hz, 2 H), 4.46-4.25 (m, 3 H), 3.57 (br s, 2 H), 3.50-3.32 (m, 1 H),
2.51 (br s, 2 H), 2.37-2.21 (m, 2 H), 2.17 (s, 9 H),1.93 (d, J = 6.8
Hz, 6 H), 1.61 (m, 2 H), 0.67 (s, 9 H). MS m/z 629.6 (M þ H)þ.
4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-N-(5-(piperidin-4-yl)-
pyridin-2-yl)pyrimidin-2-amine (54) was prepared from tert-butyl
4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-
1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)piperidine-1-
carboxylate following the procedure described for the deprotec-
tion reaction of 42 as a hydrochloride salt in quantitative yield.
1H NMR(400 MHz, CD3OD) δ 8.72 (d, J = 5.5Hz, 1 H), 8.31 (s,
1 H), 8.21 (d, J = 8.5 Hz, 1 H), 7.76 (d, J = 9.03 Hz, 1 H), 7.65 (d,
J = 5.5 Hz, 1 H), 3.98 (dt, J = 14.05, 7.03 Hz, 1 H), 3.58 (d, J =
13.0 Hz, 2 H), 3.28-3.09 (m, 3 H), 2.19 (d, J = 14.0 Hz, 2 H),
2.06-1.89 (m, 2 H), 1.37 (d, J = 7.03 Hz, 6 H). Anal. RP-HPLC
[4-(5-Isopropyl-3-methyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(5-
piperazin-1-yl-pyridin-2-yl)-amine (51). Prepared from 4-(5-iso-
propyl-3-methyl-1H-pyrazol-4-yl)-2-methanesulfonyl-pyrimidine
(12) and 4-(6-amino-pyridin-3-yl)-piperazine-1-carboxylic acid
tert-butyl ester (16) following the procedure described for 45 in
3% yield. 1H NMR (400 MHz, DMSO-d6) δ 9.18 (s, 1 H), 8.40
(d, J = 5.18 Hz, 1 H), 8.02 (d, J = 9.22, 1 H), 7.96 (d, J = 2.78 Hz,
1 H), 7.37 (dd, J = 9.22, 2.78 Hz, 1 H), 6.84 (d, J = 5.18 Hz, 1 H),
3.59 (dd, J = 8.84, 8.84 Hz, 2 H), 7.02 (dd, J = 8.84, 2.91 Hz, 1 H),
6.84 (d, J = 5.18 Hz, 1H), 3.59 (m, 1 H), 3.01 (m, 4 H), 2.84 (m,
4 H), 1.20 (d, J = 6.95, 6 H). Anal. RP-HPLC tR = 3.27 min
(method 2, purity 98%/95%). HR-MS m/z (M þ H)þ: measured
379.2371, calcd 379.2359.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(6-
piperazin-1-yl-pyridazin-3-yl)-amine (52). tert-Butyl 4-(6-(4-(5-
chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyr-
azol-4-yl)pyrimidin-2-ylamino)pyridazin-3-yl)piperazine-1-carbox-
ylate was prepared from 7and 4-(6-amino-pyridazin-3-yl)-piperazine-
1-carboxylic acid tert-butyl ester (31) following the procedure
described for the coupling reaction of 42 in 16% yield. 1H NMR
(400 MHz, CD3OD) δ 8.52 (d, J = 5.0 Hz, 1 H), 8.35 (d, J = 9.5
Hz, 1 H), 7.39 (d, J = 10.0 Hz, 1 H), 7.18 (d, J = 5.0Hz, 1 H), 5.52
(s, 2H), 3.76-3.66 (m, 2 H), 3.66-3.53 (m, 10 H), 1.51 (s, 9 H),
1.24 (d, J = 7.0 Hz, 6 H), 0.93 (t, J = 8.0 Hz, 2H), 0.00 (s, 9 H).
MS m/z 630.5 (M þ H)þ.
tR = 3.63 min (method 5, purity 96.73%/98.27%). HR-MS m/z
(M þ H)þ: measured 398.1842, calcd 398.1860.
{4-[5-Chloro-3-isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-
1H-pyrazol-4-yl]-pyrimidin-2-yl}-[5-(4-methyl-piperazin-1-yl)-
pyridin-2-yl]-amine (55). To a solution of 7 (100 mg, 0.258
mmol) in 1,4-dioxane (4 mL) and water (100 μL) in a microwave
vial were added 5-(4-methyl-piperazin-1-yl)-1-pyridin-2-ylamine
(17, 49.6 mg, 1.0 equiv), BINAP (16.1 mg, 0.1 equiv), NaOtBu
(37.2 mg, 1.5 equiv), and Pd2(dba)2 (11.8 mg, 0.05 equiv).
Nitrogen was bubbled into the resulting mixture for 3 min to
degas. The mixture was sealed and heated at 140 °C for 6 h. After
cooling to room temperature, the reaction mixture was filtered
through a pad of Celite (rinsed with EtOAc). The filtrate was
washed with water (2 ꢀ 10 mL) and brine (10 mL). The organic
layer wasdried over Na2SO4, filtered, andconcentrated in vacuo.
Column chromatography (EtOAc/heptanes 0-100%) yielded a
solid, which was triturated with cold MeCN to give {4-[5-chloro-3-
isopropyl-1-(2-trimethylsilanyl-ethoxymethyl)-1H-pyrazol-4-yl]-
pyrimidin-2-yl}-[5-(4-methyl-piperazin-1-yl)-pyridin-2-yl]-amine
(33 mg) in 23.5% yield. 1H NMR (400 MHz, CDCl3) δ 8.48 (d,
J = 5.0 Hz, 1 H), 8.31 (d, J = 9.0 Hz, 1 H), 8.01 (d, J = 2.5 Hz, 1
H), 7.78 (s, 1 H), 7.32 (dd, J = 9.0, 3.0 Hz, 1 H), 7.05 (d, J = 5.5
Hz, 1 H), 5.50 (s, 2 H), 3.68 (m, 2 H), 3.66-3.55 (m, 1 H), 3.21 (br
s, 4 H), 2.66 (br s, 4 H), 2.41 (br s, 3 H), 1.28 (d, J = 7.0 Hz, 6 H),
0.94 (m, 2 H), 0.00 (s, 9 H). MS m/z 543.6 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(6-
piperazin-1-yl-pyridazin-3-yl)-amine (52) was prepared fromtert-
butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridazin-3-yl)-
piperazine-1-carboxylate following the procedure described for
the deprotection reaction of 42 as a hydrochloride salt in 63%
yield. 1HNMR (400 MHz, CD3OD) δ 8.79 (d, J = 5.6 Hz, 1 H),
8.14 (d, J = 10.1 Hz, 1 H), 7.90 (d, J = 10.0 Hz, 1 H), 7.69 (d, J =
5.4 Hz, 1 H), 4.02-3.80 (m, 5 H), 3.47-3.39 (m, 4 H), 1.38 (d, J =
7.1 Hz, 6 H). Anal. RP-HPLC tR = 3.33 min (method 5, purity
100.00%/100.00%). HR-MS m/z (M þ H)þ: measured 400.1771,
calcd 400.1765.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(3,4,5,
6-tetrahydro-2H-[1,20]bipyrazinyl-50-yl)-amine (53). tert-Butyl
4-(5-(4-(5-chloro-3-isopropyl-1-((2-(trimethyl-silyl)ethoxy)methyl)-1H-
pyrazol-4-yl)pyrimidin-2-ylamino)pyrazin-2-yl)piperazine-1-car-
boxylate was prepared from 7 and 50-amino-2,3,5,6-tetrahydro-
[1,20]bipyrazinyl-4-carboxylic acid tert-butyl ester (32) following
the procedure described for the coupling reaction of 55 (see below)
in 50% yield. 1H NMR (400 MHz, CDCl3) δ 9.22 (d, J = 1.4 Hz,
1 H), 8.47 (d, J = 5.2 Hz, 1 H), 7.86 (d, J = 1.5 Hz, 1 H), 7.58 (s,
1 H), 7.06 (d, J = 5.2 Hz, 1 H), 5.49 (s, 2 H), 3.75-3.64 (m, 2 H),
3.64-3.44 (m, 9 H), 1.50 (s, 9 H), 1.27 (d, J = 7.0 Hz, 6 H),
1.01-0.89 (m, 2 H), 0.00 (s, 9 H). MS m/z 630.5 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-(3,
4,5,6-tetrahydro-2H-[1,20]bipyrazinyl-50-yl)-amine (53) was pre-
pared from tert-butyl 4-(5-(4-(5-chloro-3-isopropyl-1-((2-(trimethy-
lsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyrazin-
2-yl)piperazine-1-carboxylate following the procedure described
for the deprotection reaction of 42 as a hydrochloride salt in 87%
yield. 1H NMR (400 MHz, CD3OD) δ 8.52 (d, J = 6.9 Hz, 1 H),
8.37 (d, J = 1.4 Hz, 1 H), 8.23 (d, J = 1.3 Hz, 1 H), 7.87 (d, J = 6.8
Hz, 1 H), 4.16 (dt, J= 14.0, 6.9 Hz, 1 H), 3.98-3.86 (m, 4 H), 3.47-
3.34 (m, 4 H), 1.40 (d, J = 7.1 Hz, 6 H). Anal. RP-HPLC tR = 3.54
min (method 5, purity 100.00%/100.00%). HR-MS m/z(M þ H)þ:
measured 400.1765, calcd 400.1765.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
[5-(4-methyl-piperazin-1-yl)-pyridin-2-yl]-amine (55) was pre-
pared from {4-[5-chloro-3-isopropyl-1-(2-trimethylsilanyl-ethoxy-
methyl)-1H-pyrazol-4-yl]-pyrimidin-2-yl}-[5-(4-methyl-piperazin-
1-yl)-pyridin-2-yl]-amine following the procedure described for the
deprotection reaction of 42 as a hydrochloride salt in 18.6% yield.
1H NMR (400 MHz, CD3OD) δ 8.59 (d, J = 6.0 Hz, 1 H), 8.08
(dd, J = 9.5, 3.0 Hz, 1 H), 7.86 (d, J = 3.0 Hz, 1 H), 7.57 (d, J =
5.5 Hz, 1 H), 7.44 (d, J = 9.5 Hz, 1 H), 3.97-3.75 (m, 3 H),
3.65-3.49 (m, 3 H), 3.30-3.06 (m, 3 H), 2.89 (s, 3 H), 1.25 (d, J =
7.0 Hz, 6 H). Anal. RP-HPLC tR = 4.04 min (method 5, purity
100.00%/100.00%). HR-MS m/z (M þ H)þ: measured 413.1960,
calcd 413.1969.
2-(4-{6-[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-
2-ylamino]-pyridin-3-yl}-piperazin-1-yl)-ethanol (56). To a solu-
tion of [4-(5-chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
(5-piperazin-1-yl-pyridin-2-yl)-amine hydrochloride (50, 50 mg,
0.125 mmol) in acetonitrile (5 mL) were added di-isopropylethy-
lamine (44 μL, 2.0 equiv) and 2-bromoethanol (9.3 μL, 1.05
equiv). The reaction mixture was stirred at 40 °C for 24 h. The
reaction mixture was diluted in EtOAc (10 mL) and washed with
water (2 ꢀ 20 mL) and then with brine (10 mL). The organic layer
was dried over Na2SO4, filtered, and concentrated in vacuo. The
residue was purified by HPLC to give 2-(4-{6-[4-(5-chloro-3-
isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-ylamino]-pyridin-3-yl}-
piperazin-1-yl)-ethanol (56, 9.5 mg) in 17% yield. 1H NMR (400
MHz, CDCl3) δ 8.50 (d, J = 5.0 Hz, 1 H), 8.32 (d, J = 9.0 Hz,
1 H), 8.20 (br s, 1 H), 8.08 (d, J= 2.5 Hz, 1 H), 7.34 (dd, J=9.5,3.0Hz,
4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-N-(5-(piperidin-4-yl)-
pyridin-2-yl)pyrimidin-2-amine (54). tert-Butyl 4-(6-(4-(5-chloro-
3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)-
pyrimidin-2-ylamino)pyridin-3-yl)piperidine-1-carboxylate was
prepared from 7 and 6-amino-30,40,50,60-tetrahydro-20H-[3,40]-
bipyridinyl-10-carboxylic acid tert-butyl ester (28) following the
procedure described for the coupling reaction of 42 in 8% yield.

7954 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22
Cho et al.
1 H), 7.22 (d, J = 5.0 Hz, 1 H), 3.97-3.80 (m, 1 H), 3.75 (t,
J = 5.3 Hz, 2 H), 3.64 (t, J = 6.8 Hz, 1 H), 3.25-3.13 (m, 4 H),
2.79 (br s, 4 H), 2.71 (t, J = 5.0 Hz, 1 H), 1.81-1.62 (m, 1 H), 1.34
(d, J = 7.0 Hz, 6 H), 0.96 (t, J = 7.3 Hz, 1 H). Anal. RP-HPLC
(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-
pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)-2,2-dimethylpi-
perazine-1-carboxylate was prepared from 7 and 4-(6-amino-
pyridin-3-yl)-2,2-dimethyl-piperazine-1-carboxylic acid tert-butyl
ester (20) following the procedure described for the coupling
reaction of 55 in 42% yield. 1H NMR (400 MHz, CDCl3) δ 8.46
(d, J = 5.2 Hz, 1 H), 8.26 (d, J = 9.1 Hz, 1 H), 7.84 (d, J = 2.8 Hz,
1 H), 7.69 (br s, 1 H), 7.13 (dd, J = 9.22, 2.8 Hz, 1 H), 7.03 (d, J =
5.2 Hz, 1 H), 5.49 (s, 2 H), 3.79 (t, J = 5.6 Hz, 2 H), 3.73-3.66 (m,
2 H), 3.61 (quin, J = 6.9 Hz, 1 H), 3.34 (t, J = 5.6 Hz, 2 H), 3.22 (s,
2 H), 1.56 (s, 6 H), 1.50 (s, 3 H), 1.45 (s, 6 H), 1.28 (d, J = 6.9 Hz,
6 H), 0.99-0.91 (m, 2 H), 0.00 (s, 9 H). MS m/z 657.4 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
(3,3-dimethyl-piperazin-1-yl)-pyridin-2-yl]-amine (59) was prepared
from tert-butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-
3-yl)-2,2-dimethylpiperazine-1-carboxylate following the procedure
described for the deprotection reaction of 42 as a hydrochloride
salt in 65% yield. 1H NMR (400 MHz, CD3OD) δ 8.70 (d, J =
5.5 Hz, 1 H), 8.18 (dd, J = 9.8, 2.8 Hz, 1 H), 7.96 (d, J = 3.0 Hz,
1 H), 7.67 (d, J = 5.5 Hz, 1 H), 7.55 (d, J = 9.5 Hz, 1 H), 3.98 (dt,
J = 14.0,7.0 Hz, 1 H), 3.49 (br s, 4 H), 3.35 (s, 2 H), 1.54 (s, 6 H),
1.35 (d, J = 7.0 Hz, 6 H). Anal. RP-HPLC tR = 3.88 min (method
5, purity 100.00%/100.00%). HR-MS m/z (M þ H)þ: measured
427.2122, calcd 427.2125.
t
R = 3.90 min (method 5, purity 94.44%/95.66%). HR-MS m/z
(M þ H)þ: measured 443.2079, calcd 443.2075.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((S)-3-methyl-piperazin-1-yl)-pyridin-2-yl]-amine (57). (S)-tert-
Butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)-2-
methylpiperazine-1-carboxylate was prepared from 7 and
(S)-4-(6-amino-pyridin-3-yl)-2-methyl-piperazine-1-carboxylic
acid tert-butyl ester (19) following the procedure described for
the coupling reaction of 55 in 51% yield. 1H NMR (400 MHz,
CDCl3) δ 8.90(d, J = 9.7 Hz, 1 H), 8.58(d, J = 5.3 Hz, 1 H), 7.76
(dd, J = 10.0, 3.0 Hz, 1 H), 7.54 (d, J = 2.8 Hz, 1 H), 7.32 (d, J =
5.3 Hz, 1 H), 5.50 (s, 2 H), 4.43 (br s, 1 H), 4.03 (d, J = 15.0 Hz,
1 H), 3.76-3.54 (m, 3 H), 3.43 (d, J = 12.4 Hz, 1 H), 3.33-3.22
(m, 2 H), 3.05 (d, J = 3.9 Hz, 1 H), 2.84 (br s, 1 H), 1.54-1.46 (m,
9 H), 1.38-1.24 (m, 9 H), 1.20 (d, J = 5.2 Hz, 1 H), 1.00-0.94
(m, 2 H), 0.00 (s, 9 H). MS m/z 643.6 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
[5-((S)-3-methyl-piperazin-1-yl)-pyridin-2-yl]-amine (57) was pre-
pared from (S)-tert-butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-
(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-yla-
mino)pyridin-3-yl)-2-methylpiperazine-1-carboxylate following
the procedure described for the deprotection reaction of 42 as a
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
(3,5-dimethyl-piperazin-1-yl)-pyridin-2-yl]-amine (60). tert-Butyl
4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-
1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)-2,6-dimethyl-
piperazine-1-carboxylate was prepared from 7 and 4-(6-amino-
pyridin-3-yl)-2,6-dimethyl-piperazine-1-carboxylic acid tert-butyl
ester (21) following the procedure described for the coupling
reaction of 55 in 82% yield. 1H NMR (400 MHz, CDCl3) δ 8.48
(d, J = 5.2 Hz, 1 H), 8.35 (d, J = 9.1 Hz, 1 H), 7.97 (d, J = 2.5 Hz,
1 H), 7.33 (dd, J = 9.1, 2.5 Hz, 1 H), 7.06 (d, J = 5.2 Hz, 1 H), 5.50
(s, 2H), 4.32-4.21 (m, 2H), 3.74-3.65 (m, 2 H), 3.61 (dt, J = 13.7,
6.8 Hz, 1 H), 3.26 (d, J = 11.7 Hz, 2 H), 2.9 (dd, J = 11.6, 4.2 Hz, 2
H), 1.50 (s, 10 H), 1.39 (d, J = 6.8 Hz, 6 H), 1.29 (d, J = 7.0 Hz, 6
H), 0.99-0.91 (m, 2 H), 0.00 (s, 9 H). MS m/z 657.4 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-
[5-(3,5-dimethyl-piperazin-1-yl)-pyridin-2-yl]-amine (60) was
prepared from tert-butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(tri-
methylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)-
pyridin-3-yl)-2,6-dimethylpiperazine-1-carboxylate following the
procedure described for the deprotection reaction of 42 as a
1
hydrochloride salt in quantitative yield. H NMR (400 MHz,
CD3OD) δ 8.68 (d, J = 5.8 Hz, 1 H), 8.17 (dd, J = 9.0, 2.8 Hz,
1 H), 7.96 (d, J = 2.8 Hz, 1 H), 7.65 (d, J = 5.8 Hz, 1 H), 7.53 (d,
J = 9.6 Hz, 1 H), 3.96 (quin, J = 7.0 Hz, 1 H), 3.86 (dd, J = 19.9,
12.9 Hz, 2 H), 3.61-3.50 (m, 2 H), 3.40-3.31 (m, 1 H), 3.24-3.13
(m, 1 H), 2.95 (dd, J = 13.1,10.5 Hz, 1 H), 1.43 (d, J = 6.6 Hz, 3
H), 1.34 (d, J = 7.0 Hz, 6 H). Anal. RP-HPLC tR = 3.75 min
(method 5, purity 100.00%/100.00%). HR-MS m/z (M þ H)þ:
measured 413.1970, calcd 413.1969.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((R)-3-methyl-piperazin-1-yl)-pyridin-2-yl]-amine (58). (R)-tert-
Butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-yl)-
2-methylpiperazine-1-carboxylate was prepared from 7 and
(R)-4-(6-amino-pyridin-3-yl)-2-methyl-piperazine-1-carboxylic
acid tert-butyl ester (18) following the procedure described for
the coupling reaction of 55 in 54% yield. 1H NMR (400 MHz,
CDCl3) δ 8.48(d, J = 5.2 Hz, 1 H), 8.31(d, J = 9.1 Hz, 1 H), 7.98
(d, J = 2.8 Hz, 1 H), 7.77 (s, 1 H), 7.29 (d, J = 3.0 Hz, 1 H), 7.05
(d, J = 5.2 Hz, 1 H), 5.49 (s, 2 H), 4.38 (br s, 1 H), 3.98 (d, J =
13.4 Hz, 1 H), 3.72-3.65 (m, 2 H), 3.61 (quin, J = 6.9 Hz, 1 H),
3.49 (q, J = 7.1 Hz, 1 H), 3.41 (d, J = 11.6 Hz, 1 H), 3.29-3.22
(m, 2 H), 2.93 (dd, J = 11.8, 3.8 Hz, 1 H), 2.75 (td, J = 11.7, 3.5
Hz, 1 H), 1.50 (s, 9 H), 1.34 (d, J = 6.8 Hz, 3 H), 1.28 (d, J = 7.0
Hz, 6 H), 1.22 (t, J = 7.0 Hz, 1 H), 0.91-0.98 (m, 2 H), 0.00 (s, 9
H). MS m/z 643.6 (M þ H)þ.
1
hydrochloride salt in quantitative yield. H NMR (400 MHz,
DMSO-d6) δ 8.70 (d, J = 5.4 Hz, 1 H), 8.10 (d, J = 9.2 Hz, 1 H),
8.03 (d, J = 2.8 Hz, 1 H), 7.81 (d, J = 9.5 Hz, 1 H), 7.44 (d, J =
5.4 Hz, 1 H), 3.91-3.83 (m, 3 H), 3.78 (dt, J = 14.0, 6.9 Hz, 2 H),
3.40(br s, 3 H), 2.88 (t, J = 12.1Hz, 2 H), 1.35 (s, (d, J = 6.6Hz, 6
H), 1.29 (d, J = 7.1 Hz, 6 H). Anal. RP-HPLC tR = 3.77 min
(method 5, purity 98.58%/100.00%). HR-MS m/z (M þ H)þ:
measured 427.2109, calcd 427.2125.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((R)-3-methyl-piperazin-1-yl)-pyridin-2-yl]-amine (58) was prepared
from (R)-tert-butyl 4-(6-(4-(5-chloro-3-isopropyl-1-((2-(trimethyl-
silyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-
3-yl)-2-methylpiperazine-1-carboxylate following the procedure
described for the deprotection reaction of 42 as a hydrochloride
salt in quantitative yield. 1H NMR (400 MHz, CD3OD) δ 8.60
(d, J = 5.5 Hz, 1 H), 8.10 (dd, J = 9.5, 3.0 Hz, 1 H), 7.87 (d, J =
3.0 Hz, 1 H), 7.56 (d, J = 5.5 Hz, 1 H), 7.45 (d, J = 9.5 Hz, 1H),
3.88 (dt, J = 14.0,7.0 Hz, 1 H), 3.84-3.70 (m, 2 H), 3.48 (d, J =
4.5 Hz, 2 H), 3.26 (td, J = 12.3, 3.0 Hz, 1 H), 3.15-3.04 (m, 1 H),
2.86 (dd, J = 13.0, 10.5 Hz, 1 H), 1.34 (d, J = 6.5 Hz, 3 H), 1.25
(d, J = 7.0 Hz, 6 H). Anal. RP-HPLC tR = 3.70 min (method 5,
purity 95.28%/97.72%). HR-MS m/z (M þ H)þ: measured
413.1956, calcd 413.1969.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((S)-3-methyl-piperazin-1-ylmethyl)-pyridin-2-yl]-amine (61).
(S)-tert-Butyl 4-((6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-
3-yl)methyl)-2-methylpiperazine-1-carboxylate was prepared from
7 and (S)-4-(6-amino-pyridin-3-ylmethyl)-2-methyl-piperazine-1-
carboxylic acid tert-butyl ester (24) following the procedure de-
scribed for the coupling reaction of 55 in 15% yield. 1H NMR (400
MHz, CDCl3) δ 8.54 (d., J = 5.0 Hz, 1 H), 8.42 (d., J = 8.5 Hz,
1 H), 8.23 (s,1 H), 8.20 (br s, 1 H), 7.68 (dd, J = 8.5, 2.0 Hz, 1 H),
7.10 (d., J = 5.0 Hz, 1 H), 5.50 (s, 2 H), 4.20 (br s, 1 H), 3.82 (d, J =
12.0 Hz, 1 H), 3.74-3.66 (m, 2 H), 3.66-3.57 (m, 1 H), 3.54-3.46
(m, 1H), 3.41-3.33 (m, 1 H), 3.10 (td, J = 12.5, 3.0 Hz, 1 H), 2.78
(d., J = 11.0 Hz, 1 H), 2.59 (d, J = 11.5 Hz, 1 H), 2.14 (dd, J =
11.3, 3.8 Hz, 1 H), 2.08-1.97(m, 1H), 1.46(s, 9H), 1.28(d, J=6.5
Hz, 6 H), 1.22 (d, J = 6.5 Hz, 3 H), 0.99-0.91 (m, 2 H), 0.00 (s, 9
H). MS m/z 657.3 (M þ H)þ.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-(3,
3-dimethyl-piperazin-1-yl)-pyridin-2-yl]-amine (59). tert-Butyl 4-

Article
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 22 7955
RP-HPLC tR = 3.84 min (method 5, purity 100.00%/100.00%).
HR-MS m/z (M þ H)þ: measured 441.2282, calcd 441.2282.
N*60*-[4-(5-Isopropyl-3-trifluoromethyl-1H-pyrazol-4-yl)-pyri-
midin-2-yl]-N*4*,N*4*-dimethyl-3,4,5,6-tetrahydro-2H-[1,30]-
bipyridinyl-4,60-diamine (64). 4-[2-(4-Dimethylamino-3,4,5,6-
tetrahydro-2H-[1,30]bipyridinyl-60-ylamino)-pyrimidin-4-yl]-5-
isopropyl-3-trifluoromethyl-pyrazole-1-sulfonic acid dimethyl-
amide was prepared from 4-(2-chloro-pyrimidin-4-yl)-5-iso-
propyl-3-trifluoromethyl-pyrazole-1-sulfonic acid dimethylamide
(14) and N*4*,N*4*-dimethyl-3,4,5,6-tetrahydro-2H-[1,30]bipyr-
idinyl-4,60-diamine (22) following the procedure described for
((S)-3-methyl-piperazin-1-ylmethyl)-pyridin-2-yl]-amine (61) was
prepared from (S)-tert-butyl 4-((6-(4-(5-chloro-3-isopropyl-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-
ylamino)pyridin-3-yl)methyl)-2-methylpiperazine-1-carboxylate
following the procedure described for the deprotection reaction
of 42 as a hydrochloride salt in 81% yield. 1H NMR (400 MHz,
CD3OD) δ 8.63 (d, J = 6.0 Hz, 1 H), 8.55 (s, 1 H), 8.35 (dd, J =
9.0, 2.0 Hz, 1 H), 7.69 (d, J = 6.0 Hz, 1 H), 7.50 (d, J = 9.0 Hz,
1 H), 4.35 (br s, 2 H), 3.93 (dt, J = 14.0, 7.0 Hz, 1 H), 3.71 (br s,
1 H), 3.28 (br s, 1 H), 3.07 (br s, 1 H), 1.31 (d, J = 7.0 Hz, 3 H),
1.26 (d, J= 7.0 Hz, 6 H). Anal. RP-HPLC tR = 3.84 min (method 5,
purity 81.28%/94.42%). HR-MS m/z (M þ H)þ: measured
427.2135, calcd 427.2125.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((R)-3-methyl-piperazin-1-ylmethyl)-pyridin-2-yl]-amine (62). (R)-
tert-Butyl 4-((6-(4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)-
ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-ylamino)pyridin-3-
yl)methyl)-2-methylpiperazine-1-carboxylate was prepared from
7 and (R)-4-(6-Amino-pyridin-3-ylmethyl)-2-methyl-piperazine-
1-carboxylic acid tert-butyl ester (25) following the procedure
described for the coupling reaction of 55 in 32% yield. 1H NMR
(400 MHz, CDCl3) δ 8.52 (d, J = 5.0 Hz, 1 H), 8.42 (d, J = 8.6
Hz, 1 H), 8.22 (d, J = 1.5Hz, 1 H), 7.97(br s, 1 H), 7.70 (br s, 1 H),
7.11 (d., J = 5.0 Hz, 1 H), 5.50 (d, J = 8.6 Hz, 1 H), 4.21 (br s, 1
H), 3.83 (d, J = 12.1 Hz, 1 H), 3.75-3.57 (m, 3 H), 3.52 (d, J =
11.1 Hz, 1 H), 3.38 (d, J = 14.1 Hz, 1 H), 3.12 (br s, 1 H), 2.79 (br s,
1 H), 2.78 (d, J = 11.0 Hz, 1 H), 2.61 (d, J = 10.6 Hz, 1 H), 2.16
(br s, 1 H), 2.05 (br s, 1 H), 1.46 (s, 9 H), 1.29 (d, J = 7.1 Hz, 6 H),
1.24 (d, J = 6.6 Hz, 3 H), 0.99-0.91 (m, 2 H), 0.00 (s, 9 H). MS m/z
657.6 (M þ H)þ.
1
the coupling reaction of 55 in 56% yield. H NMR (400 MHz,
DMSO-d6) δ 9.75 (s, 1 H), 8.58 (d, J = 5.0 Hz, 1 H), 8.00 (d, J =
3.0 Hz, 1 H), 7.96, (d, J = 9.0 Hz, 1 H), 7.40 (dd, J = 9.0, 3.0 Hz,
1 H), 6.97 (d, J = 5.0 Hz, 1 H), 3.71-3.57 (m, 3 H), 3.14 (s, 6 H),
2.74-2.56 (m, 2 H), 2.18 (s, 6 H), 1.82 (d, J = 12 Hz, 2 H), 1.45 (m,
2 H), 1.18 (d, J = 7.0 Hz, 6 H). MS m/z 582.4 (M þ H)þ.
To a suspension of 4-[2-(4-dimethylamino-3,4,5,6-tetrahydro-
2H-[1,30]bipyridinyl-60-ylamino)-pyrimidin-4-yl]-5-isopropyl-3-tri-
fluoromethyl-pyrazole-1-sulfonic acid dimethylamide (140 mg,
0.241 mmol) in MeOH (12 mL) was added a few drops of 37%
HCl and the resulting solution was stirred at 40 °C for 16 h. The
reaction mixture was concentrated and dissolved in MeOH (2 mL).
The solution was filtered through PL-HCO3 MP-Resin (Strato-
Spheres SPE, www.polymerlabs.com) and concentrated to give
N*60*-[4-(5-Isopropyl-3-trifluoromethyl-1H-pyrazol-4-yl)-pyri-
midin-2-yl]-N*4*,N*4*-dimethyl-3,4,5,6-tetrahydro-2H-[1,30]-
bipyridinyl-4,60-diamine (64, 84.7 mg) in 74% yield. 1H NMR
(400 MHz, DMSO-d6) δ 13.75 (br s, 1 H), 9.52 (s, 1 H), 8.51 (d,
J = 5.0 Hz, 1 H), 8.16-7.92 (m, 2 H), 7.37 (dd, J = 9.3, 2.8 Hz,
1 H), 6.84 (d, J = 5.0 Hz, 1 H), 3.63 (d, J = 12 Hz, 2 H), 3.41 (m,
1 H), 2.63 (t, J = 11 Hz, 2 H), 2.19 (s, 6 H), 1.83 (d, J = 12 Hz,
2H), 1.48(m, 2H), 1.24(d, J= 7.0 Hz, 6 H). Anal. RP-HPLC tR =
3.91 min (method 1, purity 100.00%/100.00%). HR-MS m/z
(M þ H)þ: measured 475.2543, calcd 475.2546.
[4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-pyrimidin-2-yl]-[5-
((R)-3-methyl-piperazin-1-ylmethyl)-pyridin-2-yl]-amine (62) was
prepared from (R)-tert-butyl 4-((6-(4-(5-chloro-3-isopropyl-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazol-4-yl)pyrimidin-2-
ylamino)pyridin-3-yl)methyl)-2-methylpiperazine-1-carboxylate
following the procedure described for the deprotection reaction
of 42 as a hydrochloride salt in quantitative yield. 1H NMR (400
MHz, CD3OD) δ 8.63 (d, J = 5.5 Hz, 1 H), 8.53 (br s, 1 H), 8.34
(d, J = 9.0 Hz, 1 H), 7.69 (d, J = 6.0 Hz, 1 H), 7.50 (d, J = 9.0Hz,
1 H), 4.29 (br s, 3 H), 3.93 (m, 1 H), 3.68 (br s, 1 H), 3.30-3.51 (m,
3 H), 3.02 (br s, 1 H), 1.30 (d, J = 6.5Hz, 3 H), 1.26 (d, J = 7.0Hz,
6 H). Anal. RP-HPLC tR = 3.85 min (method 5, purity 89.92%/
100.00%). HR-MS m/z (M þ H)þ: measured 427.2120, calcd
427.2125.
Acknowledgment. We thank Dr. Karen Miller-Moslin for
helpful suggestions in the preparation of the manuscript.
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4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-N-(5-(4-(dimethyl-
amino)piperidin-1-yl)pyridin-2-yl)pyrimidin-2-amine (63). 4-(5-
Chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-
pyrazol-4-yl)-N-(5-(4-(dimethylamino)piperidin-1-yl)pyridin-2-
yl)pyrimidin-2-amine was prepared from 7 and N*4*,N*4*-
dimethyl-3,4,5,6-tetrahydro-2H-[1,30]bipyridinyl-4,60-diamine (22)
following the procedure described for the coupling reaction of 55 in
25% yield. 1H NMR (400 MHz, CDCl3) δ 8.50 (d, J = 5.0 Hz, 1
H), 8.30 (d, J=9.0Hz, 1H), 8.14(s, 1H), 8.04(d, J= 3.0 Hz, 1 H),
7.33 (dd, J = 9.0, 3.0 Hz, 1 H), 7.04 (d, J = 5.0 Hz, 1H), 5.50 (s, 2
H), 3.76-3.55 (m, 5 H), 3.17 (br s, 1 H), 2.74 (td, J = 12.0, 2.0 Hz, 2
H), 2.37 (s, 6 H), 1.99 (d, J = 12.5 Hz, 2 H), 1.78-1.63 (m, 2 H),
1.28 (d, J = 6.5 Hz, 6 H), 0.99-0.90 (m, 2 H), 0.00 (s, 9 H). MS m/z
571.3 (M þ H)þ.
4-(5-Chloro-3-isopropyl-1H-pyrazol-4-yl)-N-(5-(4-(dimethyl-
amino)piperidin-1-yl)pyridin-2-yl)pyrimidin-2-amine (63) was prepared
from 4-(5-chloro-3-isopropyl-1-((2-(trimethylsilyl)ethoxy)-
methyl)-1H-pyrazol-4-yl)-N-(5-(4-(dimethylamino)piperidin-1-yl)-
pyridin-2-yl)pyrimidin-2-amine following the procedure described
for the deprotection reaction of 42 as a hydrochloride salt in 71.8%
yield. 1H NMR (400 MHz, CD3OD) δ 8.68 (d, J = 5.7 Hz, 1 H),
8.17 (dd, J = 9.7, 3.0 Hz, 1 H), 7.90 (d, J = 2.8 Hz, 1 H), 7.63 (d,
J = 5.7 Hz, 1 H), 7.51 (d, J = 9.7 Hz, 1 H), 3.84-4.05 (m, 3 H),
3.53-3.36 (m, 1 H), 2.98-2.95 (m, 2 H), 2.91 (s, 7 H), 2.25 (d, J =
13.0 Hz, 2 H), 1.91 (m, 2 H), 1.34 (d, J = 7.0 Hz, 6 H). Anal.
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