Novel inhibitors of the v-raf murine sarcoma viral oncogene homologue B1 (BRAF) based on a 2,6-disubstituted pyrazine scaffold

Article abstract of DOI:10.1021/jm070776b

BRAF, a serine/threonine kinase, plays a key role in the development of certain types of cancer, particularly melanoma. 2-(3,4,5-Trimethoxyphenylamino)- 6-(3-acetamidophenyl)-pyrazine, 1, was identified as a low micromolar (IC ₅₀ = 3.5 μM) BRAF inhibitor from a high-throughput screen of a library of 23000 compounds. This compound was chosen as the starting point of a program aimed at developing inhibitors of mutant ^V600EBRAF. We have already reported on the optimization of the trimethoxyphenylamino moiety of 1. In this paper, we describe the synthesis of a series of compounds derived from 1 with the purpose of optimization of the pyrazine central core and the phenylacetamido moiety in order to increase the potency against ^V600EBRAF compared to CRAF. The biological activity of the new inhibitors was assessed against mutant ^V600EBRAF in vitro several compounds were identified with IC₅₀s of 300-500 nM for ^V600EBRAF, and all compounds that were assessed showed selectivity for ^V600EBRAF compared to CRAF by 5->86-fold.

Full text of DOI:10.1021/jm070776b

J. Med. Chem. 2008, 51, 3261–3274
3261
Novel Inhibitors of the v-raf Murine Sarcoma Viral Oncogene Homologue B1 (BRAF) Based on
a 2,6-Disubstituted Pyrazine Scaffold
Ion Niculescu-Duvaz,† Esteban Roman,† Steven R. Whittaker,‡ Frank Friedlos,† Ruth Kirk,‡ Ian J. Scanlon,†
Lawrence C. Davies,† Dan Niculescu-Duvaz,† Richard Marais,‡ and Caroline J. Springer*
The Institute of Cancer Research, Cancer Research UK Centre for Cancer Therapeutics, 15 Cotswold Road, Sutton,
Surrey SM2 5NG, United Kingdom
ReceiVed June 29, 2007
BRAF, a serine/threonine kinase, plays a key role in the development of certain types of cancer, particularly
melanoma. 2-(3,4,5-Trimethoxyphenylamino)-6-(3-acetamidophenyl)-pyrazine, 1, was identified as a low
micromolar (IC50 ) 3.5 µM) BRAF inhibitor from a high-throughput screen of a library of 23000 compounds.
This compound was chosen as the starting point of a program aimed at developing inhibitors of mutant
V600EBRAF. We have already reported on the optimization of the trimethoxyphenylamino moiety of 1. In
this paper, we describe the synthesis of a series of compounds derived from 1 with the purpose of optimization
of the pyrazine central core and the phenylacetamido moiety in order to increase the potency against
V600EBRAF compared to CRAF. The biological activity of the new inhibitors was assessed against mutant
V600EBRAF in vitro. Several compounds were identified with IC50s of 300-500 nM for V600EBRAF, and all
compounds that were assessed showed selectivity for V600EBRAF compared to CRAF by 5->86-fold.
Introduction
The RAF-MEK-ERK signal transduction cascade is a
conserved protein pathway that regulates cell growth, dif-
ferentiation, and proliferation in response to external stimuli
(growth factors, cytokines, or hormones). Phosphorylation of
RAF stimulates its serine/threonine activity, triggering sequential
phosphorylation of MEK and ERK. ERK further phosphorylates
transcription factors such as ELK-1, regulating gene expression
and controlling the response of the cell to external factors.1
BRAFa is mutated in approximately 7% of human cancers, with
particularly high frequency in melanoma (50-70%), ovarian
(35%), thyroid (30%), and colorectal (10%) cancers.2,3 The most
common mutation (90%) is valine substitution by glutamic acid
at position 600 (V600E). Mutated BRAF shows a 500-fold
elevated kinase activity, providing cancer cells with both
proliferation and survival signals.4 Therefore, mutant BRAF is
an important drug target for the treatment of human cancers.
Figure 1
The RAF family proteins (A, B, and C) are serine/threonine
High-throughput screening, of a Biofocus library of 23000
kinases.5 A number of inhibitors, designed to target CRAF, are
compounds, provided us with a lead compound, 1, that showed
known. These include benzylidene oxindoles,6 ZM 336,372,7
and sorafenib, 2.8–10 Sorafenib inhibits V600EBRAF (IC50 ) 43
nM) and, recently, isoquinolines11,12 and triaryl imidazoles have
been reported to have good activity against BRAF.13,14 The most
intensively studied RAF inhibitor, sorafenib (launched for renal
cell carcinoma [RCC]15) failed to show therapeutic activity in
the treatment of malignant melanoma despite positive results
in RCC. This failure can be attributed to its inability to reach a
concentration in melanoma cells sufficient to inhibit V600EBRAF.
Therefore, more potent V600EBRAF inhibitors are required for
melanoma patients.
low micromolar potency against isolated V600EBRAF enzyme
(IC50 ) 3.5 µM) and a tumor cell line driven by mutant BRAF
(SRB, G I50 ) 3.6 µM). See Figure 1.
Three main structural domains were chosen for optimization
of hit 1: the trimethoxyphenyl moiety (ring A), the central
pyrazine core (ring B), and the acetamidophenyl group (ring
C). We have described previously the modifications of ring A.16
Those efforts provided a few compounds that exhibited an
improved biological profile with respect to the hit 1. Herein we
focus on the optimization of rings B and C with the aim of
improving the inhibitory IC50 of the series for V600EBRAF and
examining some key V600EBRAF inhibitors thus found for their
selectivity compared to CRAF.
Rationale of the Design. The pyrazine hit 1 has a chemical
structure significantly different from sorafenib and the other
BRAF inhibitors. The design of new analogues was guided by
medicinal chemistry considerations. The rationale for the
synthesis of these compounds was to make inhibitors that are
more selective for mutant BRAF compared to CRAF. The most
* To whom correspondence should be addressed. Phone: +44 20 8722
4214. Fax: +44 20 8722 4046. E-mail: caroline.springer@icr.ac.uk
†
Cancer Research UK Centre for Cancer Therapeutics at the Institute
of Cancer Research.
‡
Cancer Research UK Centre for Cell and Molecular Biology at the
Institute of Cancer Research, 237 Fulham Road, London SW3 6JB, United
Kingdom.
a Abbreviations: BRAF, v-raf murine sarcoma viral oncogene homologue
B1; CRAF, v-raf murine sarcoma viral oncogene homologue C1.
10.1021/jm070776b CCC: $40.75
2008 American Chemical Society
Published on Web 05/13/2008

3262 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
Scheme 3. Route for Central-Core Modified Derivativesa
Scheme 1a
a Reagents and conditions: (a) 3,5-dibromopyridine, Pd(0)2dba3, BINAP,
NaOtBu, toluene, 90 °C; (b) PdCl2 ·dppf:DCM 1:1,DME, Na2CO3 1M, 90 °C.
a Reagents and conditions: (a) Pd(0)2dba3, BINAP, NaOtBu, toluene,
90 °C; (b) PdCl2 ·dppf:DCM 1:1, DME, Na2CO3 1M, 90 °C.
Scheme 4. Synthesis of the Diamino Analoguesa.
Scheme 2. Route for the Synthesis of Compound 16a
a Reagents and conditions: (a) aminoaryl compounds, Pd(0)2dba3, 1,3-
bis-(2,6-di-i-propylphenyl)-4,5-dihydroimidazolium tetrafluoroborate, KOt-
Bu, dioxane, 100 °C, microwave.
the NOESY spectrum of 15, which shows a crossed peak
between the singlet of the pyrazine central core and two of the
peaks of the phenyl moiety, fully consistent with the expected
isomer. Additionally, the 3-acetamido moiety behaved as an
isolated system exhibiting no crossed peaks with the rest of the
molecule. The other 6-(3-acetamido) group, on the other hand,
interacted with the phenyl moiety, giving the expected crossed
peaks. Following the NOESY crossed peaks, this last acetamido
unit can also be linked to the pyrazine singlet. All these
observations confirmed that the compound obtained was the
expected isomer, the Suzuki coupling displacing the least
hindered bromine.
Several modifications to the pyrazine central core were
attempted by introduction of heterocycles other than pyrazine
(see Scheme 3). Reaction of 3,4,5-trimethoxyaniline with 3,5-
dibromopyridine yielded the corresponding intermediate 17. For
amination, we used Xantphos (for the dibromo heterocycles),
BINAP, dicyclohexylphosphino-2′-(N,N-dimethylaminobiphe-
nyl), or imidazolium ligands with Pd catalysts and conventional
or microwave heating. The resulting intermediates were reacted
with 3-acetamido-boronic acid under Suzuki conditions to obtain
the desired compound 18.
We attempted the introduction of a linker (O, N, or S) in
between rings B and C to ascertain whether it would be
beneficial in increasing the potency of these molecules. There-
fore, the synthesis of the diamino analogues (see Scheme 4)
was carried out from intermediate 3, which was converted under
harsher amination conditions with microwave or conventional
heating to the desired compounds 19-25. Higher loadings of
ligands and catalysts were used to compensate for the loss of
activity of the pyrazine system upon introduction of the first
amino group.
Synthesis of the Analogues with an Oxygen Linker. For
the synthesis of the O-linker analogues, two strategies were
followed. First, the common intermediate 3 (see Scheme 5) was
reacted with the corresponding phenol under microwave heating
conditions in the presence of potassium tert-butoxide. The same
chemistry was used to synthesize compound 30 with a -S-
linker. However, using this route, the overall yields are low
a Reagents and conditions: (a) Ac2O, Na2CO3, reflux; (b) 3-acetami-
dophenyl boronic acid, PdCl2 ·dppf:DCM 1:1, DME, Na2CO3 1M, 90 °C,
microwave; (c) 3,4,5-trimethoxyaniline, Pd2(0)dba3, Xantphos, NaOtBu,
toluene, 110 °C.
well-known RAF inhibitor, sorafenib, is often described as a
BRAF inhibitor, but we find here that it has no selectivity for
BRAF. Herein, we describe potent new inhibitors that are much
more selective for mutant BRAF compared to CRAF.
Chemistry. Scheme 1 outlines the synthesis of 1 and related
pyrazine analogues (compounds 4-13). The amination of 2,6-
dichloropyrazine with 3,4,5-trimethoxyaniline furnished the
common intermediate 3,19,20 which was reacted with several
boronic acids or esters, via Suzuki coupling,21–23 to furnish the
final targets. Several procedures were used for Suzuki couplings,
with good results, including ligands such as 1,4-bis-(diphe-
nylphosphino)-butane (dppb), 2,2′-bis-(diphenylphosphino)-1,1′-
binaphtalene (BINAP), 4,5-bis-(diphenylphosphino)-9,9-dim-
ethylxanthene (Xantphos), dicyclohexylphosphino-2′-(N,N-dim-
ethylaminobiphenyl) or imidazolium salts, and palladium catalyst
such as tris-(dibenzylideneacetone)-dipalladium (0) (Pd(0)2dba3),
[1,1′-bis-(diphenyl phosphino)-ferrocene]dichloropalladium (II)
complex with DCM 1:1, (PdCl2 ·dppf:DCM 1:1). The use of
phase transfer agents such as tetrabutyl ammonium fluoride
proved advantageous in some cases. Both conventional and
microwave heating were employed.
General Strategy for the Synthesis of 1 and Related Com-
pounds. The synthesis of acetamidopyrazine 16 (see Scheme
2) involved an additional acetylation step. 3-Amino-2,6-dibro-
mopyrazine was converted with acetic anhydride to the diamido
pyrazine 14, which was reacted with 3-acetamidophenylboronic
acid via Suzuki coupling. During this process, one amido group
was cleaved, giving intermediate 15, which in turn was subjected
to amination with 3,4,5-trimethoxoxyphenylamine to furnish
acetamidopyrazine 16. It was found that the Suzuki coupling
yielded the 2-bromo-3-acetamido-6-(3-acetamidophenyl)-pyra-
zine isomer and not the 2-(3-acetamidophenyl)-3-acetamido-6-
bromopyrazine isomer. This was ascertained by the analysis of

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3263
Table 2. Substitution of the Phenyl Ring C
Scheme 5a
a Reagents and conditions: (a) KOtBu, DMF, 90 °C; (b) Pd(0)2dba3,
dicyclohexylphosphino-2′-(N,N-dimethylamino-biphenyl), NaOtBu, toluene,
90 °C.
Table 1. Replacement of the Ring C with Unsubstituted Aryl and
Heteroaryl Systems
and the purification of the final compounds difficult. Therefore,
we took the alternative approach, which involved the reaction
of 2,6-dichloropyrazine with the corresponding potassium
phenolate and the subsequent coupling of the resulting inter-
mediates (33, 33a-51) with 3,4,5-trimethoxyaniline to yield the
desired products (52-69).
Table 3. Substitution of the Pyrazine Central Core with Other
Heterocycles
Several acetamidonaphthyl (4-, 5-, and 6-acetamido isomers)
were synthesized in one step from the corresponding commercial
amines by treatment with acetic anhydride. The resulting
intermediates were reacted with 2,6-dichloropyrazine to produce
the corresponding chloropyrazines (50, 51), which were sub-
sequently treated with a series of amines to give the desired
final targets (70, 77-79). The 2(3,4,5-trimethoxyphenylamino)-
6-(4-acetamidonaphthyloxy)-pyrazine, 76, was obtained by
coupling of the intermediate 3 with the corresponding 4-aceta-
mido-napthol. The oximes (71-75) were obtained by treating
the corresponding ketones (58, 59, 61, 68, 69) with hydroxy-
lamine in EtOH.
Structure-Activity Relationships. The biological activities
of the compounds were determined using the determination of
the potency against the V600EBRAF enzyme in vitro (IC50,
BRAF). The data are summarized in Tables 1-6.
whereas replacement of ring C with a pyridine restores the
activity (4-pyridyl-, 4).
We have shown previously16 that replacement of the methoxy
group from ring A with an imidazolyl moiety increased the
potency against BRAF to IC50 < 1µM. In addition, not all
3-methoxy groups are required for activity; equipotent com-
pounds with only two methoxy groups have been synthesized.
Modification of Ring C. There is a complete loss of activity
when ring C is a phenyl or naphthyl system (data not shown),
Replacement of the Acetamidophenyl Moiety (Ring C)
by Simple Scaffolds. Introduction of other substituents in
positions 3 and 4 of the phenyl ring C (see Table 2) resulted in
some interesting observations.
Substitution in Position 3. Introduction of an extra methylene
group on the 3-acetamido function results in a total loss of

3264 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
Table 4. Introduction of Substituents an -N- Linker between Rings B
and C
The most promising systems turned out to be fused rings,
especially the 1-naphthyloxy substituent (compound 26), which
is more potent on BRAF than hit 1. By contrast, the 2-naph-
thyloxy derivative 26a was devoid of activity. The most
inhibitory BRAF compounds are the 4- and 5-acetamido-
naphthyl-1-oxy derivatives 76 and 70 with IC50 of 410-560
nM (see Table 6).
A number of oximes were also synthesized (compounds
71-75). They are more active than the precursor ketones, but
none of them yield an inhibition of BRAF below 1 µM. Finally,
we synthesized a number of compounds combining the most
effective structural feature resulting from the foregoing SAR
studies on rings A, B, and C (compounds 77-79). The most
effective compound (79) shows an IC50 of 310 nM.
Sorafenib was originally identified as an inhibitor of CRAF
and subsequently found to inhibit mutant BRAF among other
kinases.17 Sorafenib is reportedly 6-fold more potent against
CRAF than it is against V600EBRAF. Therefore, we sought to
generate inhibitors that were capable of preferentially inhibiting
V600EBRAF compared to CRAF in order specifically to target
oncogenic BRAF. Compounds were tested for activity in a
CRAF kinase assay and a selectivity index for inhibition of
oncogenic BRAF over CRAF was calculated (see Table 7). In
contrast to sorafenib, which in our hands displayed equal
potency against both BRAF and CRAF, all pyrazine compounds
tested proved to be 5->86-fold more selective for BRAF
compared to CRAF.
Conclusions
A 2,6-disubstituted pyrazine scaffold selected from a high-
throughput screen led to a series of new inhibitors of
V600EBRAF. Pyrazine inhibitors with potency <500 nM and
5-86-fold more selective for mutant BRAF compared to CRAF
have now been obtained.
activity. None of the substituents replacing the 3-acetamido
group is beneficial.
Substituents in Position 4. Attaching the acetamido moiety
in position 4 results in a loss of activity. The only modification
that is fully tolerated in this position is the carboxamide function
(compound 13), which results in a compound active on BRAF.
Modifications of Ring B. To assess the role of the pyrazine
ring in binding, the pyrazine central core was replaced by
pyridine (see Table 3). This modification was partially tolerated,
resulting compound 18 being less active against the enzyme.
Replacement of the pyrazine ring by other heterocycles such
as pyrimidine or pyridazine renders the compounds inactive.
Electronic effects might explain the decrease in activity shown
by the pyridine compound 18. Many kinase inhibitors have a
hinge binding motif consisting of a hydrogen donor-hydrogen
acceptor pair. The 3-acetamidopyrazine compound 16 proved
to be inactive, possibly due to the steric hindrance of the hinge
binding.
Linkers between Rings B and C. Three different linkers,
-N-, -O-, and -S-, were explored between the central core
(pyrazine ring B) and the phenyl ring C (see Tables 4 and 5).
Introduction of an -NH- group as linker between rings B and
C (compound 22, see Table 4) offered no improvement over
hit 1, the equivalent with an -N- linker of the hit, was less
active.
Experimental Section
All starting materials, reagents, and solvents for reactions were
reagent grade and used as purchased. Chromatography solvents were
HPLC grade and were used without further purification. Reactions
were monitored by thin layer chromatography (TLC) analysis using
Merck Silica Gel 60 F-254 thin layer plates. Flash column
chromatography was carried out on Merck silica gel 60 (0.015-0.040
mm) or in disposable Isolute Flash Si and Si II silica gel columns.
Preparative TLC was performed on either Macherey-Nagel (809
023) precoated TLC plates SIL G-25 UV254 or Analtech (2015)
precoated preparative TLC plates, 2000 µm with UV254. LC-MS
analyses were performed on a Micromass LCT/Waters Alliance
2795 HPLC system with a Discovery 5 µm, C18, 50 mm × 4.6
mm i.d. column from Supelco at a temperature of 22 °C using the
following solvent system: solvent A, methanol; solvent B, 0.1%
formic acid in water, at a flow rate of 1 mL/min. Gradient starting
with 10% A/90% B from 0-0.5 min then 10% A/90% B to 90%
A/10% B from 0.5 to 6.5 min and continuing at 90% A/10% B up
to 10 min. From 10 to 10.5 min, the gradient reverted back to 10%
A/ 90% B and was held until 12 min. UV detection was at 254 nm
and ionization was positive or negative ion electrospray. The
molecular weight scan range was 50-1000. Samples were supplied
as 1 mg/mL in DMSO or methanol with 3 µL injected on a partial
loop fill. NMR spectra were recorded in DMSO-d6 on a Bruker
DPX 250 operating at 250.13 MHz or on a Bruker Avance 500
operating at 500.26 MHz. The signal of the deuterated solvent was
used as internal reference; the chemical shifts are expressed in ppm
(δ). Preparative HPLC purifications were performed on an Axial
Chromatospac Prep 10 (Jobin-Yvon) using Merck Kieselgel 60
(0.015-0.040). Microwave reactions were run on a CM discovery
unit operating at 200 W with simultaneous stirring. The parallel
An oxygen atom as linker was also investigated (see Table
5) by using substituted phenyl rings, fused rings (1- and
2-naphthyl, quinolines, indanones, tetralones, etc.), and biphenyl
systems as ring C. Regarding phenyl substitution, hydrophobic
atoms such as F and Cl resulted in a decrease of potency with
respect to the hit. Introduction of groups able to establish
hydrogen bond interactions did not produce any improvement.

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3265
Table 5. Introduction of an -O- Linker between Rings B and C
synthesis in liquid phase were run on a Carousel (12 place reaction
station) from Radley Discovery Technology.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-pyridyl)-pyrazine (4)
(Method A). 2-(3,4,5-trimethoxyphenylamino)-6-chloropyrazine 3
(200 mg, 0.68 mmol), PdCl2.dppf:DCM 1:1 (55 mg, 0.068 mmol)
and 4-pyridyl boronic acid (100 mg, 0.82 mmol) were dissolved in
DME (15 mL). To this solution, an aqueous solution of Na2CO3
(1.4 mL, 1M) was added under stirring and the reaction mixture
was heated at 90 °C for 24 h. After cooling, Et2O (25 mL) was
added and the organic solution washed with brine (2 × 30 mL),
dried (MgSO4), and evaporated to dryness. The brown solid
obtained was recrystallized from AcOEt, giving the title compound

3266 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Table 6. Combination of the Core with the Best Rings B and C
Niculescu-DuVaz et al.
min; m/z: 354.5 [(M + H)+, 100] calcd for (C19H20N3O4). HRMS:
(M + H)+ calcd for C19H20N3O4, 354.1454; found, 354.1425.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-hydroxyphenyl)-pyra-
zine (7). Using method B (reaction time: 48 h) with 4-(4,4,5,5-
tetramethyl-1,3,2-dioxoborolan-2-yl)phenol (242 mg, 1.10 mmol),
1
the title compound was obtained (83 mg). Yield: 60%. H NMR
(250 MHz, DMSO-d6) δ: 3.63 (s, 3H), 3.82 (s, 6H), 6.88 (d, 2H,
J ) 8.5 Hz), 7.24 (s, 2H), 7.97 (d, 1H, J ) 8.7), 8.05 (s, 1H), 8.40
(s, 1H), 9.46 (s, 1H, OH), 9.82 (s, 1H, NH). 13C NMR (62.9 MHz,
DMSO-d6) δ: 55.59, 60.11, 95.65, 96.22, 115.60, 127.37, 127.88,
129.02, 131.82, 132.06, 132.20, 133.27, 134.89, 136.72, 140.98,
147.89, 151.32, 152.37, 152.78, 158.90. LC-MS: tR ) 6.90 min;
m/z: 354.4 [(M + H)+, 100] calcd for C19H20N3O4. HRMS: (M +
H)+ calcd for C19H20N3O4, 354.1454; found, 354.1444.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-hydroxymethylphenyl)pyra-
zine (8). Using method B with 4-hydroxy-methylphenyl boronic
acid (131 mg, 0.86 mmol), the title compound was obtained (22
1
mg). Yield: 8%. H NMR (250 MHz, DMSO-d6) δ: 3.69 (s, 3H),
3.90 (s, 6H), 4.57 (d, 2H, J ) 5.7 Hz), 5.28 (t, 1H, J ) 5.7 Hz),
7.26 (s, 2H), 7.45 (s, 2H, J ) 8.2 Hz), 8.10 (d, 1H), 8.14 (s, 1H),
8.51 (s, 1H), 9.56 (s, 1H, NH). LC-MS: tR ) 6.84 min; m/z: 368.1
[(M + H)+, 100] calcd for C20H21N3O4. HRMS: (M + H)+ calcd
for C20H21N3O4, 368.1610; found, 368.1606.
Table 7. Comparison of Compound Activity vs V600EBRAF and CRAFa
2-(3,4,5-Trimethoxyphenylamino)-6-(ethyl,3-benzoate)-pyra-
zine (9). Using method B (reaction time: 3 h) with ethyl-3-(4,4,5,5-
tetramethyl-1,3,2-dioxoborolan-2-yl)-benzoate (450 mg, 1.63 mmol),
compound
number
V600EBRAF
IC50 (µM)
CRAF
IC50 (µM)
selectivity
index
1
the title compound was obtained (391 mg). Yield: 70%. H NMR
sorafenib
0.043
1.16
1.12
0.56
0.65
0.31
0.052
>100
38.15
20.26
6.96
1
>86
34
36
11
(250 MHz, DMSO-d6) δ: 1.34 (t, 3H, CH3CH2, J ) 7.1 Hz), 3.64
(s, 3H), 3.81 (s, 6H), 4.36 (q, 2H, CH3CH2), 7.22 (s, 2H), 7.67 (t,
1H, J ) 7.8 Hz), 8.05 (d, 1H, J ) 7.2 Hz), 8.20 (s, 1H), 8.36 (d,
1H, J ) 7.8 Hz), 8.57 (s, 1H), 8.68 (s, 1H), 9.63 (s, 1H, NH).
LC-MS: tR ) 8.65 min; m/z: 410.1 [(M + H)+, 100] calcd for
C22H24N3O5. HRMS: (M + H)+ calcd for C22H23N3O5, 410, 1710;
found, 410,1720.
11
13
76
77
79
1.52
5
a A selectivity index was calculated (CRAF IC50/V600EBRAF IC50) to
indicate selectivity of compounds for oncogenic BRAF over CRAF.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-formylphenyl)-pyra-
zine (10). Using method A with 4-formylphenylboronic acid (123
mg, 0.82 mmol), the title compound was obtained (153 mg). Yield:
62%. 1H NMR (500 MHz, DMSO-d6) δ: 3.64 (s, 3H), 3.83 (s, 6H),
7.25 (s, 2H), 8.05 (d, 2H, J ) 8.2 Hz), 8.23 (s, 1H), 8.35 (d, 2H),
8.64 (s, 1H), 9.68 (s, 1H, NH), 10.08 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 55.62, 60.12, 96.00, 126.97, 129.93, 130.68, 132.25,
132.87, 136.53, 136.64, 142.10, 146.30, 151.50, 152.84, 192.70.
LC-MS: tR ) 7.34 min; m/z: 366.1 [(M + H)+, 100] calcd for
C20H20N3O4. HRMS: (M + H)+ calcd for C20H20N3O4, 366.1454;
found, 366.1458.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-phenylamino)-pyrazine
(11a). Using method A (reaction time: 18 h) with 4-(4,4,5,5-
tetramethyl-1,3,2-borolan-2-yl)-aniline (500 mg, 1.69 mmol), the
title compound was obtained (350 mg). Yield: 59%. 1H NMR (500
MHz, DMSO-d6) δ: 3.63 (s, 3H), 3.82 (s, 6H), 5.53 (s, 2H, NH2),
6.64 (d, 2H, J ) 8.6 Hz), 7.24 (s, 2H), 7.84 (d, 2H), 7.95 (s, 1H),
8.34 (s, 1H), 9.38 (s, 1H, NH). LC-MS: tR ) 4.08 min; m/z: 352.9
[(M + H)+, 100] calcd for C19H20N4O3.
1
(40 mg). Yield: 17%. H NMR (250 MHz, DMSO-d6) δ: 3.64 (s,
3H), 3.83 (s, 6H), 7.22 (s, 2H), 8.07 (d, 2H, J ) 4.73 Hz), 8.26 (s,
2H), 8.65 (d, 2H,), 8.72 (d, 1H, HPy), 9.70 (s, 1H, NH). LC-MS: tR
) 6.20 min; m/z: 339.1 [M+ + H, 100] calcd for C18H19N4O3.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-pyridyl)-pyrazine (5). Us-
ing method A (reaction time: 3 h) with 3-pyridylboronic acid (100
mg, 0.82 mmol), the title compound was obtained (120 mg). Yield:
52%. 1H NMR (500 MHz, DMSO-d6) δ: 3.67 (s, 3H), 3.85 (s, 6H),
7.26 (s, 2H, H), 7.58-7.62 (m, 1H), 8.25 (s, 1H), 8.49 (d, 1H, J )
7.9 Hz), 8.63 (s, 1H), 8.69 (d, 1H, J ) 3.2 Hz), 9.51 (d, 1H, J )
2.2 Hz), 9.69 (s, 1H, NH). 13C NMR (62.9 MHz, DMSO-d6) δ
55.41, 60.20, 96.20, 123.79, 124.40, 129.80, 130.40, 132.50, 133.70,
134.59, 137.00, 145.49, 147.65, 150.20, 151.99, 152.90. LC-MS:
tR ) 6.19 min; m/z: 339.1 [M+ + H, 100] calcd for C18H19N4O3.
HRMS: (M + H)+ calcd for C18H19N4O3, 339.1457; found,
339.1445.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-hydroxyphenyl)-pyra-
zine (6) (Method B). In an oven-dried flask, the catalyst is prepared
by adding under stirring and Ar atmosphere dppb (40 mg, 0.09
mmol) and PdCl2(benzonitrile)2 (30 mg, 0.077 mmol) to dry toluene
(10 mL). After 30 min, 2 (112 mg, 0.38 mmol), 3-(4,4,5,5-
tetramethyl-1,3,2-dioxoborolan-2-yl)-phenol (100 mg, 0.45 mmol),
EtOH (1.60 mL), and an aqueous solution of Na2CO3 (1.5 mL,
1M) in toluene (10 mL) were added and the reaction mixture was
heated at 90 °C for 16 h. After cooling, AcOEt (50 mL) was added
to the reaction, and the mixture was washed (2 × 100 mL), dried
(MgSO4), and evaporated to dryness. The solid obtained was
purified by preparative HPLC (Kieselgel 60, 0.015-0.043; eluent:
AcOEt) to furnish the title compound (74 mg). Yield: 55%. 1H
NMR (250 MHz, DMSO-d6) δ: 3.63 (s, 3H), 3.82 (s, 6H), 6.87 (d,
1H, J ) 9.8 Hz), 7.25 (s, 2H), 7.29 (t, 1H, J ) 9.5 Hz), 7.52 (d,
1H, J ) 6.5 Hz), 7.53 (s, 1H), 8.14 (s, 1H), 8.43 (s, 1H), 9.54 (s,
1H, OH), 9.59 (s, 1H, NH). 13C NMR (62.9 MHz, DMSO-d6) δ:
55.61, 60.11, 95.70, 113.28, 116.46, 117.14, 129.80, 131.90, 133.44,
136.90, 138.04, 147.73, 151.38, 152.79, 157.87. LC-MS: tR ) 7.04
1-{4-[6-(3,4,5-Trimethoxyphenylamino)-pyrazin-2-yl]phenyl}-3-
phenyl urea (11). To 200 mg of 2-(3,4,5-trimethoxyphenylamino)-
6-(4-phenylamino)-pyrazine (11a) dissolved in dry THF 68 mL
(0.60 mmol) of phenyl isocyanate were added under stirring at 40
°C (block). After 20 h, the reaction mixture was evaporated under
vacuum and the residue was recrystallized from AcOEt, when 130
1
mg of a yellow solid were obtained. Yield: 49%. H NMR (500
MHz, DMSO-d6) δ: 3.65 (s, 3H), 3.85 (s, 6H), 7.00 (t, 1H), 7.26
(s, 2H), 7.32 (s, 2H, J ) 7.6 Hz), 7.48 (d, 2H), 7.60 (s, 2H, J )
8.7 Hz), 8.08 (d, 2H), 8.11 (s, 1H), 8.42 (s, 1H), 8.42 (s, 1H), 8.73
(s, 1H, NHurea), 8.92 (s, 1H, NHurea), 9.53 (s, 1H, NH). LC-MS: tR
) 7.05 min; m/z: 421.2 [(M + H)+, 100] calcd for C26H25N5O4.
HRMS: (M + H)+ calcd for C26H25N5O4, 472.1979; found,
472.1978.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-cyanophenyl)-pyrazine (12).
Using method A (reaction time: 18 h) with 3-(4,4,5,5-tetramethyl-
1,3,2-borolan-2-yl)-benzonitrile (188 mg, 0.82 mmol), the title
compound was obtained (44 mg). Yield: 18%. 1H NMR (500 MHz,

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3267
DMSO-d6) δ: 3.64 (s, 3H), 3.84 (s, 6H), 7.23 (s, 2H), 7.73 (t, 1H,
J ) 7.9 Hz), 7.93 (d, 1H, J ) 7.7 Hz), 8.22 (s, 1H), 8.45 (d, 1H,
J ) 8.0 Hz), 8.57 (s, 1H), 8.64 (s, 1H), 9.68 (s, 1H, NH). 13C
NMR (62.9 MHz, DMSO-d6) δ: 55.62, 60.12, 96.00, 112.08,
118.46, 129.93, 130.10, 130.82, 132.25, 132.77, 134.84, 136.60,
137.79, 145.38, 151.43, 151.56, 152.82. LC-MS: tR ) 7.52 min;
m/z: 363.1 [(M + H)+, 100] calcd for C20H19N4O3. HRMS: (M +
H)+ calcd for C20H19N4O3, 363.1457; found, 363.1459.
10.07 (s, 1H), 10.21 (s, 1H). LC-MS: tR ) 6.20 min; m/z: 452.2
(M + H)+ calcd for C23H25N5O5.
3-(3,4,5-Trimethoxyphenylamino)-5-bromopyridine (17) (Method
C). To 1.00 g (4.2 mmol) 3,5-dibromopyridine dissolved in 40 mL
of dry toluene in an oven-dried flask were added under stirring:
Pd(0)2dba3 (82 mg, 0.09 mmol), BINAP (0.168 g, 0.27 mmol),
1.49 g (8.16 mmol) 3,4,5-trimethoxyaniline, and 0.914 g (9.52
mmol) sodium tert-butoxide. The reaction mixture was heated at
90 °C (bath temperature) for 18 h under N2. After cooling, the
toluene solution was filtered and evaporated in vacuo. The residue
was retaken in 30 mL of AcOEt, washed (2 × 30 mL brine), dried,
evaporated to a volume of 10 mL, and submitted to HPLC
(Kieselgel 60, 0.015-0.043; eluent: AcOEt:cycloxane 2:1). A
fraction of 167 mg of the title compound was collected. Yield:
2-(3,4,5-Trimethoxyphenylamino)-6-(4-carboxamidophenyl)pyra-
zine (13). In a tube suitable for microwave and under Ar were placed
3 (150 mg, 0.51 mmol), 4-aminocarbonylphenylboronic acid (108
mg, 0.66 mmol), Pd2(0)dba3 (30 mg, 0.051 mmol), 1,3-bis(2,6-di-
i-propylphenyl)-4,5-dihydro-imidazolium tetrafluoroborate (24 mg,
0.051 mmol), tetra-n-butylammonium bromide (16.8 mg, 0.051
mmol), and toluene (3 mL). The temperature was lowered to 0 °C
and dropwise addition of a suspension of potassium methoxide
(106.8 mg, 1.53 mmol) in MeOH (1.2 mL) followed. This mixture
was stirred for 5 min at 0 °C, and then microwave irradiation for
30 min at 65 °C followed. The reaction mixture was dissolved in
boiling AcOEt (75 mL) and filtered. The resulting organic layer
was evaporated under vacuum to give a solid that gave, upon
column chromatography (AcOEt), 124 mg of the title compound.
Yield: 64%. 1H NMR (250 MHz, DMSO-d6) δ: 3.65 (s, 3H), 3.85
(s, 6H), 7.27 (s, 2H), 7.47 (bs, 1H, CONH2), 8.02 (d, 2H, J ) 8.4
Hz), 8.08 (bs, 1H, CONH2), 8.21 (s, 1H), 8.23-8.34 (m, 3H), 8.61
(s, 1H), 9.64 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.61,
60.14, 95.93, 126.15, 127.99, 130.27, 132.14, 134.23, 134.89,
136.76, 139.12, 146.73, 151.47, 152.83, 167.31. LC-MS: tR ) 5.87
min; m/z: 381.1 (M + H)+ calcd for C20H21N4O4. HRMS: (M +
H)+ calcd for C20H21N4O4, 381.1563; found, 381.1570.
1
11.7%. H NMR (250 MHz, DMSO-d6) δ: 3.64 (s, 3H), 3.78 (s,
6H), 6.43 (s, 2H), 7.52 (t, 1H, Hpyr 4, J ) 2.1), 8.04 (d, 1H, J ) 1.9
Hz, HPyr 2 or 6), 8.14 (d, 1H, HPyr 6 or 2 J ) 2.3), 8.48 (s, 1H, NH).
LC-MS: tR ) 6.28 min; (C14H15BrN2O3) m/z: 339.06 [M+ + 1,
100].
3-(3,4,5-Trimethoxyphenylamino)-5-(3-acetamidophenyl)-pyri-
dine (18). Using method A (reaction time: 4 h) with 3-(3,4,5-
trimethoxyphenylamino)-5-bromo-pyridine, 25, (160 mg, 0.97
mmol), the title compound was obtained (129 mg). Yield: 70%.
1H NMR (250 MHz, DMSO-d6) δ: 2.06 (s, 3H), 3.63 (s, 3H), 3.76
(s, 6H), 6.46 (s, 2H), 7.32 (d, 1H, J ) 7.6 Hz), 7.41 (t, 1H, J ) 7.8
Hz), 7.54 (d, 1H), 7.58 (m, 1H), 7.95 (s, 1H), 8.24 (s, 1H), 8.34 (s,
1H), 8.46 (s, 1H, NH), 10.11 (s, 1H, NH). 13C NMR (62.9 MHz,
DMSO-d6) δ: 24.03, 55.76, 60.12, 95.99, 117.19, 118.61, 119.24,
121.39, 129.47, 132.19, 135.86, 137.60, 137.88, 138.02, 138.17,
140.01, 140.70, 153.47, 168.45. LC-MS: tR ) 4.69 min; m/z: 394.1
[(M + H)+, 100] calcd for C22H24N3O4. HRMS: (M + H)+ calcd
for C22H23N3O4, 394.1767; found, 394.1754.
2-N,N-Diacetylamido-3,5-dibromo-pyrazine (14). 2-Amino-3,5-
dibromo-pyrazine (1.01 g, 4.0 mmol) and sodium bicarbonate (1.02,
12mmol) were refluxed in acetyl chloride (20 mL) for 2 days. The
solvent was evaporated, and the residue was partitioned between
DCM and water. The organic layer was dried and evaporated to
2-(3,4,5-Trimethoxyphenylamino)-6-(4-acetamidophenylamino)-
pyrazine (19) (Method D). In a microwave tube, 3, (150 mg, 0.51
mmol), 4-aminoacetanilide (114 mg, 0.76 mmol), Pd2(0)dba3 (23
mg, 0.025 mmol), 1,3-bis(2,6-di-i-propylphenyl)-4,5-dihydro-imi-
dazolium tetrafluoroborate (24 mg, 0.051 mmol), potassium tert-
butoxide (114 mg, 0.91 mmol), and dioxane (3.5 mL) were reacted
under stirring and Ar at 100 °C for 30 min with microwave heating.
The reaction mixture was filtered, and the insoluble solid was
washed with boiling AcOEt (40 mL). The solution was evaporated
under vacuum, and the resulting solid was chromatographed on
silica gel using AcOEt as eluent to produce 86 mg of title
compound. Yield: 29.8%. 1H NMR (500 MHz, DMSO-d6) δ: 2.01
(s, 3H), 3.62 (s, 3H), 3.63 (s, 6H), 6.85 (s, 2H), 7.40-7.52 (m,
6H), 9.01 (s, 1H), 9.04 (s, 1H), 9.80 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 23.82, 55.60, 60.13, 97.01, 119.17, 119.57, 121.33,
121.68, 132.23, 133.28, 136.07, 136.89, 150.22, 152.64, 167.70.
LC-MS: tR ) 5.43 min; m/z: 410.2 (M + H)+ calcd for C21H23N5O4.
2-(3,4,5-Trimethoxyphenylamino)-6-phenylamino-pyrazine (20).
Using method D with aniline (71 mg, 0.76 mmol), the title
compound was obtained (63 mg). Yield: 35%. 1H NMR (500 MHz,
DMSO-d6) δ: 3.62 (s, 3H), 3.63 (s, 6H), 6.86 (s, 2H), 6.91 (t, 1H,
J ) 7.3 Hz), 7.22 (t, 2H, J ) 9.4 Hz), 7.53 (s, 1H), 7.54 (s, 1H),
7.57 (d, 2H, J ) 7.6 Hz), 9.05 (s, 1H), 9.14 (s, 1H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 55.60, 60.11, 97.01, 118.59, 121.00,
121.87, 121.92, 128.65, 132.26, 136.85, 140.86, 150.00, 150.17,
152.84. LC-MS: tR ) 6.42 min; m/z: 353.1 (M + H)+ calcd for
C19H21N4O3. HRMS: (M + H)+ calcd for C19H21N4O3, 353.1614;
found, 353.1612.
1
give 1.31 g of title compound as crystals. Yield: 97%. H NMR
(500 MHz, DMSO-d6) δ: 2.26 (6H, s), 9.00 (1H, s). MS m/z
(electrospray): 358/60/2 (M + Na+).
2-Bromo-3-acetamido-6-(3-acetamidophenyl)-pyrazine (15). In
a microwave tube under stirring and an Ar atmosphere were placed
14 (200 mg, 0.59 mmol), 3-acetamidobenceneboronic acid (106
mg, 0.59 mmol), PdCl2 · dppf:DCM 1:1 complex (36 mg, 0.045
mmol), DME (3.0 mL), H2O (0.6 mL), and Na2CO3 (126 mg, 1.19
mmol). The tube was heated by microwave irradiation at 90 °C for
30 min. Two batches were combined, filtered, and the precipitate
washed with hot AcOEt (60 mL). The organic layer was evaporated
under vacuum, and the resulting crude was chromatographed on
1
silica (AcOEt) to give 26 mg of title compound. Yield: 7%. H
NMR (500 MHz, DMSO-d6) δ: 2.07 (s, 3H), 2.12 (s, 3H), 7.46 (t,
1H, J ) 7.9 Hz), 7.77 (d, 1H, J ) 7.8 Hz), 7.82 (d, 1H, J ) 7.7
Hz), 8.21 (bs, 1H), 9.05 (s, 1H), 10.17 (s, 1H), 10.46 (s, 1H). 13C
NMR (62.9 MHz, DMSO-d6) δ: 23.04, 23.99, 117.03, 120.73,
121.29, 129.54, 134.45, 136.31, 138.55, 140.15, 145.67, 148.90,
168.55, 168.92. LC-MS: tR ) min; m/z: 349.1 (M + H)+ calcd for
C14H13BrN4O2.
2-(3,4,5-Trimethoxyphenylamino)-3-acetamido-6-(3-acetami-
dophenyl)-pyrazine (16). In a microwave tube under stirring and
Ar atmosphere were placed 15 (32 mg, 0.091 mmol), 3,4,5-
trimethoxyaniline (33.6 mg, 0.183 mmol), sodium tert-butoxide (18
mg, 0.1833 mmol), Pd(0)2dba3 (8.4 mg, 0.009 mmol), 4,5-
bis(diphenylphosphino)-9,9-dimethylxanthene (Xantphos) (5.3 mg,
0.009 mmol), and dry toluene (1.5 mL). The tube was heated by
microwave irradiation at 110 °C for 30 min. The reaction crude
was filtered and the precipitate washed with boiling AcOEt (30
mL). The organic layers were evaporated under vacuum, and the
resulting crude was chromatographed (AcOEt:EtOH 9:1) to produce
the title compound (7 mg). Yield: 17%. 1H NMR (500 MHz,
DMSO-d6) δ: 2.06 (s, 3H), 2.17 (s, 3H), 3.64 (s, 3H), 3.77 (s, 9H),
7.24 (s, 2H), 7.42 (t, 1H, J ) 8.0 Hz), 7.64 (d, 1H, J ) 7.7 Hz),
7.73 (d, 1H, J ) 7.9 Hz), 8.22 (bs, 1H), 8.28 (s, 1H), 8.52 (s, 1H),
2-(3,4,5-Trimethoxyphenylamino)-6-(4-pyridinylamino)-pyra-
zine (21). Using method D with 4-aminopyridine (127 mg, 1.35
mmol), the title compound was obtained (25 mg). Yield: 10%. 1H
NMR (500 MHz, DMSO-d6) δ: 3.64 (s, 3H), 3.68 (s, 6H), 6.83 (s,
2H), 7.55 (d, 2H, J ) 6.3 Hz), 7.62 (s, 1H), 7.69 (s, 1H), 8.25 (d,
2H, J ) 6.3 Hz), 9.23 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6)
δ: 55.66, 60.14, 97.52, 112.07, 122.57, 123.96, 132.67, 136.40,
147.39, 148.88, 149.83, 150.18, 152.93. LC-MS: tR ) 3.70 min;
m/z: 354.2 (M + H)+ calcd for C18H19N5O3.

3268 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-acetamidophenylami-
no)pyrazine (22). Using method D with 3-aminoacetanilide (114
mg, 0.76 mmol), the title compound was obtained (12 mg). Yield:
6%. 1H NMR (500 MHz, DMSO-d6) δ: 2.01 (s, 3H), 3.61 (s, 3H),
3.62 (s, 6H), 6.88 (s, 2H), 7.04 (d, 1H, J ) 8.4 Hz), 7.11 (t, 1H, J
) 7.9 Hz), 7.50 (d, 1H, J ) 7.9 Hz), 7.53 (s, 1 H), 7.56 (s, 1 H),
7.67 (bs, 1 H), 9.05 (s, 1 H), 9.13 (s, 1 H), 9.81 (s, 1 H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 23.98, 55.58, 60.10, 96.74, 109.169,
112.22, 113.55, 121.87, 122.04, 128.74, 132.14, 136.92, 139.68,
141.08, 149.99, 150.18, 152.64, 168.15. LC-MS: tR ) 5.67 min;
m/z: 410.1 (M + H)+ calcd for C21H23N5O4.
(500 MHz, DMSO-d6) δ: 3.06 (s, 9H), 3.45 (s, 3H), 6.69 (s, 2H),
7.37 (dd, 1H, J1 ) 2.3 Hz, J2 ) 8.9 Hz), 7.49-7.54 (m, 2H), 7.72
(d, 1H, J ) 2.0 Hz), 7.84-7.89 (m, 2H), 7.92-7.99 (m, 3H), 9.55
(s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 54.81, 59.95, 95.64,
116.83, 121.05, 121.18, 125.53, 126.65, 127.15, 127.61, 128.51,
129.61, 130.45, 132.06, 133.78, 136.25, 150.57, 151.52, 152.57,
157.47. LC-MS: tR ) 8.62 min; m/z: 404.1 (M + H)+ calcd for
C23H22N3O4. HRMS: (M + H)+ calcd for C23H22N3O4, 404.1610;
found, 404.1590.
2-(3,4,5-Trimethoxyphenylamino)-6-(phenyloxy)-pyrazine (27).
Following procedure E (no K2CO3 added) with phenol (48 mg, 0.51
mmol), the title compound was obtained (127 mg). Yield: 71%.
1H NMR (500 MHz, DMSO-d6) δ: 3.31 (s, 9H), 3.54 (s, 3H), 6.77
(s, 2H), 7.16-7.23 (m, 3H), 7.41 (t, 2H, J ) 7.9 Hz), 7.75 (s, 1H),
7.95 (s, 1H), 9.53 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ:
55.33, 60.03, 95.77, 120.55, 121.17, 124.47, 128.42, 129.74, 132.15,
136.29, 150.52, 152.69, 153.86, 153.86, 157.23. LC-MS: tR ) 7.65
min; m/z: 354.1 (M + H)+ calcd for C19H19N3O4. HRMS: (M +
H)+ calcd for C19H20N3O4, 354.1454; found, 354.1444.
2-(3,4,5-Trimethoxybenzylamino)-6-(4-oxyphenylamino)-pyra-
zine (23). 4-Hydroxyaniline (58.0 mg, 0.53 mmol), DMF (5 mL),
potassium tert-butoxide (67 mg, 0.60 mmol), and K2CO3 (42 mg,
0.3 mmol) were stirred in Ar. After 5 min, 2-chloro-6-(3,4,5-
trimethoxyphenylamino)-pyrazine (150 mg, 0.51 mmol) was added
and the reaction mixture heated by microwaves (30 min, 110 °C).
The reaction mixture was filtered, poured in water (20 mL), and
extracted with AcOEt (2 × 20 mL) The organic layer was pooled,
dried, and evaporated to a volume of 2-3 mL. After purification
by flash chromatography (Isolute column, Flash SiII, 50 g, 150 mL;
eluent: AcOEt), 48.9 mg of the title compound were obtained. Yield:
2-(3,4,5-Trimethoxyphenylamino)-6-(4-pyridinyloxy)-pyrazine
(28). Using method E with 4-hydroxypyridine (48 mg, 0.51 mmol),
1
the title compound was obtained (89 mg). Yield: 50%. H NMR
1
26%. H NMR (500 MHz, DMSO-d6) δ: 3.61 (s, 12H), 6.67 (d,
(500 MHz, DMSO-d6) δ: 3.64 (s, 3H), 3.79 (s, 6H), 6.29 (d, 2H,
J ) 7.9 Hz), 7.07 (s, 2H), 8.19 (s, 1H), 8.32 (s, 1H), 8.43 (d, 2H,
J ) 8.0 Hz), 9.91 (bs, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ:
55.61, 60.11, 96.50, 117.94, 121.98, 132.71, 132.79, 135.77, 136.52,
145.43, 150.31, 152.85, 178.22. LC-MS: tR ) 5.52 min; m/z: 355.1
(M + H)+ calcd for C18H19N4O4. HRMS: (M + H)+ calcd for
C18H19N4O4, 355.1406; found, 355.1418.
2H, J ) 8.5 Hz), 6.86 (s, 2H), 7.31 (d, 2H), 7.42 (s, 2H), 8.75 (s,
1H, NH), 8.95 (s, 1H, NH), 9.07 (s, 1H, OH). LC-MS: tR ) 4.48
min; m/z: 353.1 [(M + H)+, 100] calcd for C19H21N4O3. HRMS:
(M + H)+ calcd for C19H21N4O3, 353.1614; found, 3353.1612.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-fluorophenylamino)-pyra-
zine (24). Using method D with 3-fluoroaniline (150 mg, 1.35
mmol), the title compound was obtained (60 mg). Yield: 23%. 1H
NMR (500 MHz, DMSO-d6) δ: 3.62 (s, 3H), 3.65 (s, 6H),
6.65-6.72 (m, 1H), 6.81 (s, 2H), 7.24 (m, 2H), 7.56 (s, 1H), 7.58
2-(3,4,5-Trimethoxyphenylamino)-6-(3-pyridinyloxy)-pyrazine
(29). Using method E with 3-hydroxypyridine (48 mg, 0.51 mmol),
1
the title compound was obtained (46 mg). Yield: 26%. H NMR
(s, 1H), 7.67 (d, 1H, J ) 13.0 Hz), 9.12 (s, 1H), 9.41 (s, 1H). 13
C
(250 MHz, DMSO-d6) δ: 3.41 (s, 6H), 3.54 (s, 3H), 6.71 (s, 2H),
7.47 (m, 1H, J1 ) 0.6 Hz, J2 ) 4.7 Hz, J3 ) 8.4 Hz), 7.69 (m, 1H,
J1 ) 1.4 Hz, J2 ) 2.8 Hz, J3 ) 8.4 Hz), 7.82 (s, 1H), 7.98 (s, 1H),
8.43 (dd, 1H, J1 ) 1.4 Hz, J2 ) 4.7 Hz), 8.54 (dd, 1H, J1 ) 0.5
Hz, J2 ) 2.8 Hz), 9.61 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6)
δ: 55.32, 60.04, 95.95, 120.94, 124.52, 128.42, 128.91, 132.34,
136.10, 142.68, 145.66, 150.31, 150.38, 152.71, 156.84. LC-MS:
tR ) 6.27 min; m/z: 355.1 (M + H)+ calcd for C18H19N4O4. HRMS:
(M + H)+ calcd for C18H19N4O4, 355.1406; found, 355.1416.
2-(3,4,5-Trimethoxyphenylamino)-6-(naphthalen-1-ylthio)-pyra-
zine (30). Using method E with 1-thionaphthol (81 mg, 0.51 mmol),
NMR (62.9 MHz, DMSO-d6) δ: 55.45, 60.07, 97.24, 106.71,
107.05, 114.04, 122.16, 122.49, 130.14, 132.43, 136.55, 142.71,
142.89, 149.52, 150.10, 152.89. LC-MS: tR ) 6.75 min; m/z: 371.1
[(M + H)+, 100] calcd for C19H20FN4O3. HRMS: (M + H)+ calcd
for C19H20FN4O3, 371.1519; found, 371.1528.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-fluorophenylamino)-pyra-
zine (25). Using method D with 4-fluoroaniline (150 mg, 1.35
mmol), the title compound was obtained (72 mg). Yield: 28%. 1H
NMR (250 MHz, DMSO-d6) δ: 3.62 (s, 9H), 6.83 (s, 2H), 7.06 (t,
2H, J ) 9.0 Hz), 7.51 (s, 1H), 7.57 (d, 2H, J ) 9.1 Hz), 9.05 (s,
1H), 9.14 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.56, 60.12,
97.01, 114.94, 115.29, 120.38, 120.50, 121.62, 121.83, 132.29,
137.19, 137.22, 150.15, 152.84. LC-MS: tR ) 6.38 min; purity >
99%. HRMS: (M + H)+ calcd for C19H20FN4O3, 371.1519; found,
371.1522.
1
the title compound was obtained (68 mg). Yield: 32%. H NMR
(500 MHz, DMSO-d6) δ: 3.60 (s, 3H), 3.61 (s, 6H), 6.94 (s, 2H),
7.25 (s, 1H), 7.57-7.63 (m, 3H), 7.91 (s, 1H), 7.95 (dd, 1H, J1 )
1.2 Hz, J2 ) 7.1 Hz), 8.03-8.07 (m, 1H), 8.11 (d, 1H, J ) 8.3
Hz), 8.26-8.30 (m, 1H), 9.54 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 55.55, 60.09, 96.41, 124.82, 126.23, 126.58, 126.74,
127.65, 128.90, 129.90, 130.63, 131.09, 132.52, 133.33, 133.97,
134.60, 136.13, 151.27, 152.55, 152.74. LC-MS: tR ) 8.54 min;
m/z: 420.1 (M + H)+ calcd for C23H22N3O3S. HRMS: (M + H)+
calcd for C23H22N3O3S, 420.1382; found, 420.1378.
2-(3,4,5-Trimethoxyphenylamino)-6-(5,6,7,8-tetrahydronaphtha-
len-1-yloxy)-pyrazine (31). Using method E with 5,6,7,8-tetrahydro-
1-naphthol (75 mg, 0.51 mmol), the title compound was obtained
(27 mg). Yield: 13%. 1H NMR (500 MHz, DMSO-d6) δ: 1.65-1.69
(m, 4H), 2.53-2.56 (m, 2H), 2.72-2.77 (m, 2H), 3.42 (s, 6H),
3.54 (s, 3H), 6.74 (s, 2H), 6.89 (d, 1H, J ) 7.9 Hz), 6.95 (d, 1H,
J ) 6.6 Hz), 7.11 (t, 1H, J ) 7.8 Hz), 7.71 (s, 1H), 7.91 (s, 1H),
9.55 (bs, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 22.01, 22.26,
23.12, 28.78, 55.36, 60.03, 95.72, 118.40, 120.56, 125.66, 126.09,
127.60, 129.06, 132.10, 136.39, 138.76, 150.59, 151.46, 152.70,
157.65. LC-MS: tR ) 8.62 min; m/z: 408.2 (M + H)+ calcd for
C23H26N3O4. HRMS: (M + H)+ calcd for C23H26N3O4, 408.1923;
found, 408.1916.
2-(3,4,5-Trimethoxyphenylamino)-6-(naphthalen-1-yloxy)-pyra-
zine (26) (Method E). In a microwave tube were mixed under Ar
and stirring, 1-naphthol (73 mg, 0.51 mmol), K2CO3 (35 mg, 0.25
mmol), potassium tert-butoxide (60 mg, 0.53 mmol), and DMF dry
(3.5 mL). This mixture was stirred for 10 min, 2 (150 mg, 0.51
mmol) was added and the tube was heated for 30 min at 120 °C.
The reaction crude was mixed with 40 mL of solution NaOH 1.5
M and extracted with DCM (3 × 40 mL). The organic layers were
combined, dried (MgSO4), filtered, and evaporated under vacuum
to give a solid that was chromatographed using DCM:AcOEt (7:3)
1
as eluent to produce the title compound (66 mg). Yield: 32%. H
NMR (500 MHz, DMSO-d6) δ: 3.19 (s, 6H), 3.47 (s, 3H), 6.58 (s,
2H), 7.32 (d, 1H, J ) 8.5 Hz), 7.50-7.60 (m, 3H), 7.83 (d, 1H, J
) 8.3 Hz), 7.91 (s, 1H), 7.96 (s, 1H), 7.98-8.03 (m, 2H), 9.49 (s,
1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.17, 59.96, 95.53,
117.11, 120.88, 121.28, 124.80, 126.03, 126.54, 126.65, 126.72,
127.96, 128.32, 132.07, 134.45, 136.16, 149.40, 150.61, 152.58,
158.19. LC-MS: tR ) 8.50 min; m/z: (M + H)+ calcd for
C23H22N3O4. HRMS: (M + H)+ calcd for C23H22N3O4, 404.1610;
found. 404.1651.
2-(3,4,5-Trimethoxyphenylamino)-6-(1H-indol-4-yloxy)-pyra-
zine (32). Using method E with 4-hydroxyindole (67 mg, 0.51
mmol), the title compound was obtained (34 mg). Yield: 17%. 1H
NMR (500 MHz, DMSO-d6) δ: 3.24 (s, 6H), 3.49 (s, 3H),
6.10-6.12 (m, 1H), 6.66 (s, 2H), 6.79 (d, 1H, J ) 7.1 Hz), 7.07 (t,
2-(3,4,5-Trimethoxyphenylamino)-6-(naphthalen-2-yloxy)-pyra-
zine (26a). Using method E, with 2-naphthol (73 mg, 0.51 mmol),
1
the title compound was obtained (78 mg). Yield: 38%. H NMR

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3269
1H, J ) 7.8 Hz), 7.25-7.29 (m, 2H), 7.79 (s, 1H), 7.92 (s, 1H),
9.44 (s, 1H), 11.26 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ:
55.12, 59.96, 95.43, 98.04, 108.42, 109.87, 120.50, 120.91, 121.37,
125.36, 127.87, 131.91, 136.34, 138.13, 146.33, 150.59, 152.58,
157.84. LC-MS: tR ) 7.22 min; m/z: 393.1 (M + H)+ calcd for
C21H21N4O4. HRMS: (M + H)+ calcd for C21H21N4O4, 393.1563;
found, 393.1534.
C13H8ClN2O3. HRMS: (M + H)+ calcd for C13H8ClN2O3, 275.0223;
found, 275.0231.
2-Chloro-6-(5-tetral-1-onyloxy)-pyrazine (37). Method F was
followed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and
5-hydroxy-tetralone (238 mg, 1.49 mmol). Purification by column
chromathography (AcOEt:cyclohexane, 1:1) afforded the title
1
compound (366 mg). Yield: 100%. H NMR (500 MHz, DMSO-
d6) δ: 1.97-2.07 (m, 2H), 2.64 (t, 2H, J ) 7.1 Hz), 2.77 (t, 2H, J
2-Chloro-6-(quinolin-4-yloxy)-pyrazine (33) (Method F). In a
carousel tube suitable for parallel synthesis were added 2,6-
dichloropyrazine (250 mg, 1.69 mmol), 4-hydroxyquinoline (270
mg, 1.49 mmol), dry degassed DMF (10 mL), and potassium tert-
butoxide (165 mg, 1.49 mmol). This mixture was stirred under Ar
at 90 °C (block) for 5 h. The reaction mixture was diluted with
AcOEt (20 mL), washed with brine (2 × 20 mL), dried, and
evaporated under vacuum. Purification by column chromatography
(AcOEt:cyclohexane, 1:1) afforded the title compound (60 mg).
) 6.8 Hz), 7.46-7.45 (m, 2H), 7.86 (dd, 1H, Jo ) 7.4 Hz, Jm
)
1.6 Hz), 8.54 (s, 1H), 8.63 (s, 1H). 13C NMR (62.9 MHz, DMSO-
d6) δ: 21.69, 22.73, 38.05, 124.03, 126.82, 127.38, 133.36, 134.03,
136.79, 137.60, 144.40, 149.80, 157.96, 196.73. LC-MS: tR ) 6.94
min; m/z: 275.0 (M + H)+ calcd for C14H12ClN2O2. HRMS: (M +
H)+ calcd for C14H12ClN2O2, 275.0587; found, 275.0595.
2-Chloro-6-(6-tetral-1-onyloxy)-pyrazine (38). Method F was
followed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and
6-hydroxy-tetralone (238 mg, 1.49 mmol). Crystallization from
1
Yield: 14%. H NMR (500 MHz, DMSO-d6) δ: 7.45 (d, 1H, J )
1
AcOEt furnished the title compound (365 mg). Yield: 100%. H
4.9 Hz), 7.67 (t, 1H, J ) 7.6 Hz), 7.86 (t, 1H, J ) 7.8 Hz), 8.12
(d, 2H, J ) 8.7 Hz), 8.67 (s, 1H), 8.82 (s, 1H), 8.94 (d, 1H, J )
4.9 Hz). 13C NMR (62.9 MHz, DMSO-d6) δ: 111.15, 121.39,
121.52, 127.20, 129.11, 130.74, 134.54, 139.24, 144.48, 149.59.
151.42, 156.17, 157.21. LC-MS: tR ) 5.45 min; m/z: 258.0 (M +
H)+ calcd for C13H9ClN3O. HRMS: (M + H)+ calcd for
C13H9N3OCl, 258.0434; found, 258.0427.
NMR (500 MHz, DMSO-d6) δ: 2.02-2.12 (m, 2H), 2.63 (t, 2H, J
) 5.9 Hz), 2.97 (t, 2H, J ) 6.5 Hz), 7.20-7.26 (m, 2H), 7.96 (d,
1H, J ) 8.3 Hz), 8.59 (s, 1H), 8.62 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 22.96, 29.92, 38.37, 119.23, 120.46, 128.70, 129.18,
134.08, 138.33, 144.44, 147.14, 156.34, 157.61, 196.35. LC-MS:
tR ) 7.03 min; m/z: 275.0 (M + H)+ calcd for C14H12ClN2O2.
HRMS: (M + H)+ calcd for C14H12ClN2O2, 275.0587; found,
275.0592.
2-Chloro-6-(indanone-4-oxy)-pyrazine (39). Method F was fol-
lowed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and 4-hy-
droxyindanone (219 mg, 1.49 mmol). Crystallization from AcOEt
afforded the title compound (314 mg). Yield: 90%. 1H NMR (500
MHz, DMSO-d6) δ: 2.70 (t, 2H, J ) 5.7 Hz), 2.94 (t, 2H, J ) 5.7
Hz), 7.55-7.64 (m, 3H), 8.57 (s, 1H), 8.66 (s, 1H). 13C NMR (62.9
MHz, DMSO-d6) δ: 22.43, 35.68, 120.55, 127.24, 129.29, 133.55,
137.88, 139.26, 144.38, 146.50, 149.95, 157.62, 205.32. LC-MS:
tR ) 6.48 min; m/z: 275.0 (M + H)+ calcd for C13H10ClN2O2.
HRMS: (M + H)+ calcd for C13H10ClN2O2, 261.0431; found,
261.0432.
2-Chloro-6-(indanone-5-oxy)-pyrazine (40). Method F was fol-
lowed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and 5-hy-
droxyindanone (219 mg, 1.49 mmol). Crystallization from AcOEt
produced the title compound (350 mg). Yield: 100%. 1H NMR (500
MHz, DMSO-d6) δ: 2.67 (t, 2H, J ) 5.9 Hz), 3.13 (t, 2H, J ) 5.5
Hz), 7.31 (dd, 1H, Jm ) 1.9 Hz, Jo ) 8.3 Hz), 7.47 (d, 1H, J ) 1.3
Hz), 7.73 (d, 1H, J ) 8.3 Hz), 8.59 (s, 1H), 8.64 (s, 1H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 25.45, 36.11, 120.56, 124.83, 134.07,
138.34, 144.42, 157.76, 204.61. LC-MS: tR ) 6.34 min; m/z: 261.0
(M + H)+ calcd for C13H10ClN2O2.
2-Chloro-6-(indanone-6-oxy)-pyrazine (41). Method F was fol-
lowed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and 6-hy-
droxyindanone (219 mg, 1.49 mmol). Crystallization from AcOEt
produced the title compound (330 mg). Yield: 94%. 1H NMR (500
MHz, DMSO-d6) δ: 2.74 (t, 2H, J ) 5.8 Hz), 3.15 (t, 2H, J ) 6.0
Hz), 7.50 (d, 1H, J ) 2.2 Hz), 7.60 (dd, 1H, Jm ) 2.3 Hz, Jo ) 8.3
Hz), 7.70 (d, 1H, J ) 8.3 Hz), 8.53 (s, 1H), 8.59 (s, 1H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 25.13, 36.59, 115.03, 128.01, 128.48,
133.84, 137.57, 138.21, 144.20, 152.51, 158.25, 205.56. LC-MS:
tR ) 6.48 min; m/z: 261.0 (M + H)+ calcd for C13H10ClN2O2.
HRMS: (M + H)+ calcd for C13H10ClN2O2, 261.0431; found,
261.0422.
2-Chloro-6-(quinoline-5-oxy)-pyrazine (33a). Method F was
followed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and
5-hydroxyquinoline (270 mg, 1.49 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 1:1) gave the title compound
(264 mg). Yield: 61%. 1H NMR (500 MHz, DMSO-d6) δ:
7.54-7.58 (m, 2H), 7.84 (t, 1H, J ) 8.1 Hz), 8.01 (d, 1H, J ) 8.5
Hz), 8.37 (d, 1H, J ) 8.0 Hz), 8.55 (s, 1H), 8.73 (s, 1H), 8.98 (dd,
1H, J ) 1.4 Hz, J ) 4.0 Hz). 13C NMR (62.9 MHz, DMSO-d6) δ:
118.19, 121.82, 121.99, 126.94, 129.37, 129.99, 133.76, 137.84,
144.32, 147.74, 148.57. 151.27, 158.57. LC-MS: tR ) 5.71 min;
m/z: 258.0 (M + H)+ calcd for C13H9ClN3O. HRMS: (M + H)+
calcd for C13H9N3OCl, 258.0434; found, 258.0433.
2-Chloro-6-(quinoline-6-oxy)-pyrazine (34). Method F was fol-
lowed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and 6-hy-
droxyquinoline (270 mg, 1.49 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 1:1) afforded the title com-
1
pound (324 mg). Yield: 75%. H NMR (500 MHz, DMSO-d6) δ:
7.56-7.59 (m, 1H), 7.70 (dd, 1H, Jm ) 2.6 Hz, Jo ) 9.1 Hz), 7.87
(d, 1H, J ) 2.4 Hz), 8.12 (d, 1H, J ) 9.0 Hz), 8.37 (d, 1H, J ) 8.1
Hz), 8.56 (s, 1H), 8.65 (s, 1H), 8.93 (dd, 1H, J ) 1.0 Hz, J ) 4.9
Hz). 13C NMR (62.9 MHz, DMSO-d6) δ: 117.82, 122.01, 124.48,
128.50, 131.00, 133.86, 135.69, 137.88, 144.41, 145.64, 150.58.
150.38, 158.23. LC-MS: tR ) 6.07 min; m/z: 258.0 (M + H)+ calcd
for C13H9ClN3O. HRMS: (M + H)+ calcd for C13H9N3OCl,
258.0434; found, 258.0433.
2-Chloro-6-(isoquinoline-5-oxy)-pyrazine (35). Method F was
followed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and
5-hydroxyisoquinoline (270 mg, 1.49 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 1:1) furnished the title
compound (263 mg). Yield: 61%. 1H NMR (500 MHz, DMSO-d6)
δ: 7.72-7.78 (m, 3H), 8.11 (d, 1H, J ) 7.8 Hz), 8.53 (d, 1H, J )
5.9 Hz), 8.55 (s, 1H), 8.75 (s, 1H), 9.43 (s, 1H). 13C NMR (62.9
MHz, DMSO-d6) δ: 114.05, 122.20, 125.66, 127.72, 128.98, 129.26,
133.78, 137.86, 143.56, 144.26, 146.92. 152.51, 158.45. LC-MS:
tR ) 4.94 min; m/z: 258.0 (M + H)+ calcd for C13H9ClN3O. HRMS:
(M + H)+ calcd for C13H9N3OCl, 258.0434; found, 258.0448.
2-Chloro-6-(7-coumarinyloxy)-pyrazine (36). Using method F
with 2,6-dichloropyrazine (200 mg, 1.35 mmol) and 17-hydroxy
coumarine (242 mg, 1.49 mmol), 370 mg of the desired compound
resulted after purification. Yield: 100%. 1H NMR (500 MHz,
DMSO-d6) δ: 6.50 (d, 1H, J ) 9.6 Hz), 7.29 (dd, 1H, Jo ) 8.2 Hz,
Jm ) 2.2 Hz), 7.44 (d, 1H, J ) 2.2 Hz), 7.84 (d, 1H, J ) 8.4 Hz),
8.10 (d, 1H, J ) 9.6 Hz), 8.60 (s, 1H), 8.65 (s, 1H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 109.02, 115.37, 115.89, 117.16, 129.86,
134.01, 138.39, 143.79, 144.37, 155.03. 155.35, 157.61, 159.66,
161.27. LC-MS: tR ) 6.30 min; m/z: 275.0 (M + H)+ calcd for
2-Chloro-6-(2-phenyl-phenyl-oxy)-pyrazine (42). Method F was
followed (reaction time: 20 h) with 2,6-dichloropyrazine (200 mg,
1.34 mmol) and 2-phenyl-phenol (268 mg, 1.49 mmol). Purification
by column chromatography (AcOEt:cyclohexane, 1:2) produced the
title compound (238 mg). Yield: 65%. 1H NMR (500 MHz, DMSO-
d6) δ: 7.31-7.61 (m, 9H), 8.43 (s, 1H), 8.46 (s, 1H). LC: tR
)
8.40 min; m/z: 283.1 (M + H)+ calcd for C16H11ClN2O.
2-Chloro-6-(2-fluorophenyl-oxy)-pyrazine (43). Method F was
followed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and
2-fluorophenol (166 µL, 1.86 mmol). Purification by column
chromathography (AcOEt:cyclohexane, 1:1) afforded the title
compound (302 mg). Yield: 80%. 1H NMR (500 MHz, DMSO-d6)

3270 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
δ: 7.31-7.45 (m, 4H), 8.56 (s, 1H), 8.67 (s, 1H). 13C NMR (62.9
MHz, DMSO-d6) δ: 116.94, 123.93, 125.47, 127.64, 133.10, 138.02,
144.33, 152.71, 154.68, 157.49. LC-MS: tR ) 7.19 min; m/z: 225.0
(M + H)+ calcd for C10H6ClFN2O. HRMS: (M + H)+ calcd for
C10H6ClFN2O, 225.0231; found, 225.0222.
8.53 (s, 1H), 8.67 (s, 1H), 10.23 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 23.89, 120.29, 121.38, 133.46, 136.96, 137.22,
144.37, 158.58, 168.22. LC-MS: tR ) 5.90 min; m/z: 264.0 (M +
H)+ calcd for C12H11ClN3O2. HRMS: (M + H)+ calcd for
C12H11ClN3O2, 264.0540; found, 264.0545.
2-Chloro-6-(3-fluorophenyl-oxy)-pyrazine (44). Method F was
followed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and
3-fluorophenol (166 µL, 1.86 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 1:1) afforded the title com-
2-Chloro-6-(5-acetamido-naphthyl-1-oxy)-pyrazine (51). Method
F was followed with 2,6-dichloropyrazine (200 mg, 1.34 mmol)
and 5-acetamido-1-naphthol (270 mg, 1.49 mmol). Crystallization
1
from AcOEt gave the title compound (379 mg). Yield: 90%. H
1
pound (347 mg). Yield: 92%. H NMR (500 MHz, DMSO-d6) δ:
NMR (500 MHz, DMSO-d6) δ: 2.27 (s, 3H), 7.54 (t, 1H, J ) 8.0
Hz), 7.59 (d, 1H, J ) 7.5 Hz), 7.68 (t, 1H, J ) 8.0 Hz), 7.80 (d,
1H, J ) 7.2 Hz), 7.83 (d, 1H, J ) 6.4 Hz), 8.13 (d, 1H, J ) 8.4
Hz), 8.59 (s, 1H), 8.78 (s, 1H), 10.10 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 23.47, 117.94, 118.14, 120.98, 122.31, 125.65,
126.51, 127.12, 129.19, 133.58, 134.22, 137.59, 144.42, 148.21,
158.81, 168.99. LC-MS: tR ) 6.30 min; m/z: 313.1 (M)+ calcd for
C16H12ClN3O2.
7.13-7.17 (m, 2H), 7.25 (d, 1H, JF ) 10.0 Hz), 7.52 (q, 1H, J )
8.1 Hz), 8.54 (s, 1H), 8.56 (s, 1H). 13C NMR (62.9 MHz, DMSO-
d6) δ: 109.15, 112.56, 117.28, 131.17, 133.77, 137.95, 144.34,
153.74, 157.85, 161.35. LC-MS: tR ) 7.31 min; m/z: 224.0 (M)+
calcd for C10H6ClFN2O.
2-Chloro-6-(3-chlorophenyl-oxy)-pyrazine (45). Method F was
followed with 2,6-dichloropyrazine (250 mg, 1.69 mmol) and
3-chlorophenol (193 µL, 1.86 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 1:1), afforded the title
compound (346 mg). Yield: 85%. 1H NMR (500 MHz, DMSO-d6)
δ: 7.32 (dd, 1H, Jo ) 8.2 Hz, Jm ) 1.9 Hz), 7.43 (d, 1H, J ) 8.0
Hz), 7.50 (d, 1H, Jm ) 1.9 Hz), 7.56 (t, 1H, J ) 8.1 Hz), 8.59 (s,
1H), 8.62 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 118.73,
120.09, 121.50, 125.83, 131.84, 133.76, 137.92, 144.29, 153.14,
157.88. LC-MS: tR ) 7.89 min; m/z: 240.9 (M + H)+ calcd for
C10H7Cl2N2O. HRMS: (M + H)+ calcd for C10H7Cl2N2O, 240.9935;
found, 240.9933.
2-Chloro-6-(3-acetamidophenyl-oxy)-pyrazine (46). Method F
was followed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and
3-acetamidophenol (202 mg, 1.49 mmol). Crystallization from
AcOEt e gave the title compound (350 mg). Yield: 99%. 1H NMR
(500 MHz, DMSO-d6) δ: 2.06 (s, 3H), 6.90-6.95 (m, 1H), 7.76
(dd, 2H, Jm ) 2.0 Hz, Jo ) 7.6 Hz), 7.59 (s, 1H), 8.52 (s, 1H),
8.54 (s, 1H), 10.12 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ:
24.01, 111.39, 115.40, 116.02, 130.00, 133.66, 137.57, 140.82,
144.42, 152.57, 158.23, 168.55. LC-MS: tR ) 6.02 min; m/z: 264.0
(M + H)+ calcd for C12H11ClN3O2. HRMS: (M + H)+ calcd for
C12H11ClN3O2, 264.0540; found, 264.0525.
2-Chloro-6-(4-acetamidophenyl-oxy)-pyrazine (47). Method F
was followed with 2,6-dichloropyrazine (200 mg, 1.34 mmol) and
4-acetamidophenol (202 mg, 1.49 mmol). Crystallization from
AcOEt afforded the title compound (327 mg). Yield: 92%. 1H NMR
(500 MHz, DMSO-d6) δ: 2.12 (s, 3H), 7.25 (d, 2H, J ) 8.9 Hz),
7.71 (d, 2H, J ) 8.9 Hz), 8.55 (s, 1H), 8.57 (s, 1H), 10.10 (s, 1H).
13C NMR (62.9 MHz, DMSO-d6) δ: 23.89, 120.29, 121.38, 133.46,
136.96, 137.22, 144.37, 158.58, 168.22. LC-MS: tR ) 5.90 min;
m/z: 264.0 (M + H)+ calcd for C12H11ClN3O2. HRMS: (M + H)+
calcd for C12H11ClN3O2, 264.0540; found, 264.0545.
2-(3,4,5-Trimethoxyphenylamino)-6-(7-coumarinyloxy)-pyra-
zine (52) (Method G). In a tube suitable for parallel synthesis,
2-chloro-6-(coumarin-1-yl-6-oxy)-pyrazine (200 mg, 0.72 mmol)
was reacted with 3,4,5-trimethoxyaniline (158 mg, 0.86 mmol) in
dry, degassed toluene (10 mL) in the presence of Pd(0)2 ·dba3 (20
mg, 0.022 mmol, 3 mol %), 2-dicyclohexylphosphino-2′-(N,N-
dimethylamino)-biphenyl (16 mg, 0.044 mmol, 6% mol) and sodium
tert-butoxide (138 mg, 1.44 mmol). This mixture was stirred at 90
°C for 20 h under a N2 atmosphere. The reaction content was
evaporated to dryness in vacuum, and the solid residue was retaken
in hot AcOEt (2 × 10 mL). The solutions were filtered, pooled,
washed with 0.1N HCl (10 mL) and water, dried (MgSO4), and
evaporated again. The solid residue was purified by column
chromatography (AcOEt:EtOH, 1:1) to give the title compound (102
mg). Yield: 34%. 1H NMR (500 MHz, DMSO-d6) δ: 3.31 (s, 6H),
3.39 (s, 3H), 6.47 (d, 1H, J ) 9.6 Hz), 6.77 (s, 2H), 7.22 (dd, 1H,
Jo ) 8.5 Hz, Jm ) 2.3 Hz), 7.34 (d, 1H, J ) 2.2 Hz), 7.72 (d, 1H,
J ) 8.5 Hz), 7.84 (s, 1H), 8.03 (s, 1H), 8.10 (d, 1H, J ) 9.62 Hz),
9.63 (s, 1H, NH). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.16, 60.04,
96.23, 108.23, 114.70, 115.49, 117.27, 121.24, 129.45, 129.66,
132.09, 143.89, 150.52, 152.67, 154.59, 156.51, 156.75, 159.74.
LC-MS: tR ) 6.93 min; m/z: 422.1 (M + H)+ calcd for C22H20N3O6.
HRMS: (M + H)+ calcd for C22H20N3O6, 422.1352; found,
422.1341.
2-(3,4,5-Trimethoxyphenylamino)-6-(quinoline-4-oxy)-pyra-
zine (53). Using method G with 2-chloro-6-(quinolyl-4-oxy)-
pyrazine (150 mg, 0.58 mmol) and 3,4,5-trimethoxyaniline (128
mg, 0.70 mmol), the title compound (48 mg) was obtained after
crystallization from AcOEt. Yield: 20%. 1H NMR (500 MHz,
DMSO-d6) δ: 3.22 (s, 6H), 3.49 (s, 3H), 6.67 (s, 2H), 7.16 (d, 1H,
J ) 5.0 Hz), 7.66 (t, 1H, J ) 7.7 Hz), 7.84 (t, 1H), 8.02 (s, 1H),
8.08 (d, 1H, J ) 8.1 Hz), 8.09 (s, 1H), 8.16 (d, 1H, J ) 8.1 Hz),
8.80 (d, 1H, J ) 5.0 Hz), 9.71 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 55.01, 59.98, 95.78, 110.29, 121.47, 121.63, 121.81,
126.91, 128.99, 130.28, 132.21, 135.94, 149.51, 150.67, 151.32,
152.58, 156.37, 157.59. LC-MS: tR ) 6.80 min; m/z: 406.7 (M +
H)+ calcd for C22H22N4O4. HRMS: (M + H)+ calcd for
C22H21N4O4, 405.1563; found, 405.1564.
2-Chloro-6-(acetophenone-3-oxy)-pyrazine (48). Method F was
followed with 2,6-dichloropyrazine (220 mg, 1.49 mmol) and
3-hydroxyacetophenone (200 mg, 1.47 mmol). Crystallization from
1
AcOEt gave the title compound (370 mg). Yield: 96%. H NMR
(500 MHz, DMSO-d6) δ: 2.61 (s, 3H), 7.57 (dd, 1H, Jm ) 1.2 Hz,
Jo ) 9.0 Hz), 7.65 (t, 1H, J ) 7.8 Hz), 7.82 (d, 1H, Jm ) 2.0 Hz),
7.88 (dd, 1H, Jm ) 1.3 Hz, Jo ) 7.5 Hz), 8.55 (s, 1H), 8.61 (s,
1H). LC-MS: tR ) 6.52 min; m/z: 249.0 (M + H)+ calcd for
C12H9ClN2O2 248.0.
2-(3,4,5-Trimethoxyphenylamino)-6-(quinoline-5-oxy)-pyra-
zine (54). Method G was followed with 2-chloro-6-(quinolyl-5-oxy)-
pyrazine (150 mg, 0.58 mmol) and 3,4,5-trimethoxyaniline (128
mg, 0.70 mmol). Crystallization (AcOEt) afforded the title com-
2-Chloro-6-(acetophenone-4-oxy)-pyrazine (49). Method F was
followed with 2,6-dichloropyrazine (220 mg, 1.49 mmol) and
4-hydroxyacetophenone (200 mg, 1.47 mmol). Crystallization from
AcOEt furnished the title compound (365 mg). Yield: 99%. 1H
NMR (500 MHz, DMSO-d6) δ: 2.61 (s, 3H), 7.42 (d, 2H, J ) 8.7
Hz), 8.08 (d, 2H, J ) 8.7 Hz), 8.59 (s, 1H), 8.63 (s, 1H). LC-MS:
tR ) 6.52 min; m/z: 249.0 (M + H)+ calcd for C12H9ClN2O2.
2-Chloro-6-(6-acetamido-naphthyl-1-oxy)-pyrazine (50). Method
F was followed with 2,6-dichloropyrazine (250 mg, 1.68 mmol)
and 6-acetamido-1-naphthol (350 mg, 1.72 mmol). Crystallization
from AcOEt produced the title compound (526 mg). Yield: 100%.
1H NMR (500 MHz, DMSO-d6) δ: 2.12 (s, 3H), 7.29 (d, 1H, J )
7.4 Hz), 7.53 (t, 1H, J ) 7.9 Hz), 7.57-7.62 (m, 1H), 7.80 (d, 1H,
J ) 8.2 Hz), 7.85 (d, 1H, J ) 9.1 Hz), 8.42 (d, 1H, J ) 1.3 Hz),
1
pound (77 mg). Yield: 33%. H NMR (500 MHz, DMSO-d6) δ:
3.18 (s, 6H), 3.47 (s, 3H), 6.57 (s, 2H), 7.43 (d, 1H, J ) 7.5 Hz),
7.53-7.57 (m, 1H), 7.78 (t, 1H, J ) 8.5 Hz), 7.93 (d, 1H, J ) 8.3
Hz), 8.37 (d, 1H, J ) 8.0 Hz), 7.94 (s, 1H), 7.99 (s, 1H), 8.40 (d,
1H, J ) 8.3 Hz), 8.96 (dd, 1H, Ja ) 1.5 Hz, Jb ) 3.7 Hz), 9.56 (s,
1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.12, 59.97, 95.56,
117.40, 120.93, 121.78, 122.04, 125.78, 128.72, 129.39, 130.08,
132.11, 136.08, 148.60, 149.19, 150.56, 151.17, 152.56, 157.89.
LC-MS: tR ) 7.00 min; m/z: 405.1 (M + H)+ calcd for C22H22N4O4.
HRMS: (M + H)+ calcd for C22H21N4O4, 405.1563; found,
405.1561.

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3271
2-(3,4,5-Trimethoxyphenylamino)-6-(quinoline-6-oxy)-pyra-
zine (55). Method G was followed with 2-chloro-6-(quinolyl-6-oxy)-
pyrazine (150 mg, 0.58 mmol) and 3,4,5-trimethoxyaniline (128
mg, 0.70 mmol). Crystallization (AcOEt) gave the title compound
(149 mg). Yield: 64%. 1H NMR (500 MHz, DMSO-d6) δ: 3.06 (s,
6H), 3.45 (s, 3H), 6.68 (s, 2H), 7.50-7.55 (m, 1H), 7.63 (d, 1H, J
) 9.1 Hz), 7.80 (s, 1H), 7.87 (s, 1H), 8.00 (s, 1H), 8.08 (d, 1H, J
) 9.1 Hz), 8.33 (d, 1H, J ) 8.2 Hz), 8.89 (d, 1H, J ) 2.5 Hz),
9.60 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 54.84, 59.97,
95.87, 116.89, 121.19, 121.88, 124.52, 128.62, 128.83, 130.72,
132.23, 135.36, 136.17, 145.26, 149.89, 150.57, 151.60, 152.58,
157.24. LC-MS: tR ) 6.88 min; m/z: 405.1 (M + H)+ calcd for
C22H21N4O4. HRMS: (M + H)+ calcd for C22H21N4O4, 405.1563;
found, 405.1571.
2-(3,4,5-Trimethoxyphenylamino)-6-(isoquinoline-5-oxy)-pyra-
zine (56). Method G was followed with 2-chloro-6-(isoquinolyl-5-
oxy)-pyrazine (150 mg, 0.58 mmol) and 3,4,5-trimethoxyaniline
(128 mg, 0.70 mmol). Crystallization (AcOEt) and purification by
column chromatography (AcOEt) afforded the title compound (67
mg). Yield: 29%. 1H NMR (500 MHz, DMSO-d6) δ: 3.20 (s, 6H),
3.48 (s, 3H), 6.55 (s, 2H), 7.62 (d, 1H, J ) 7.5 Hz), 7.71 (t, 1H, J
) 7.9 Hz), 7.80 (d, 1H, J ) 5.8 Hz), 7.95 (s, 1H), 7.99 (s, 1H),
8.03 (d, 1H, J ) 8.1 Hz), 8.52 (d, 1H, J ) 5.8 Hz), 9.41 (s, 1H),
9.55 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.27, 59.99,
95.91, 114.19, 120.90, 121.33, 124.41, 127.71, 128.77, 129.24,
129.31, 132.35, 136.03, 143.38, 148.42, 150.55, 152.42, 152.60,
157.78. LC-MS: tR ) 6.42 min; m/z: 405.1 (M + H)+ calcd for
C22H22N4O4. HRMS: (M + H)+ calcd for C22H21N4O4, 405.1563;
found, 405.1579.
(169 mg, 0.92 mmol). Purification by column chromatography
(AcOEt) afforded the title compound (61 mg). Yield: 20%. 1H NMR
(500 MHz, DMSO-d6) δ: 2.68 (t, 2H, J ) 5.6 Hz), 3.08 (t, 2H, J
) 5.0 Hz), 3.39 (s, 6H), 3.55 (s, 3H), 6.77 (s, 2H), 7.21 (dd, 1H,
Jm ) 1.9 Hz, Jo ) 8.3 Hz), 7.40 (s, 1H), 7.68 (d, 1H, J ) 8.3 Hz),
7.84 (s, 1H), 8.02 (s, 1H), 9.62 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 25.36, 36.04, 55.15, 60.05, 96.21, 118.03, 120.08,
121.40, 124.56, 129.45, 132.44, 132.96, 136.09, 150.61, 152.68,
156.57, 157.62, 159.52, 204.62. LC-MS: tR ) 7.11 min; m/z: 408.1
[(M + H)+ calcd for C22H22N3O5. HRMS: (M + H)+ calcd for
C22H22N3O5, 408.1559; found, 408.1561.
2-(3,4,5-Trimethoxyphenylamino)-6-(indanone-6-oxy)-pyra-
zine (61). Method G was followed with 2-chloro-6-(indanone-6-
oxy)-pyrazine (200 mg, 0.77 mmol) and 3,4,5-trimethoxyaniline
(169 mg, 0.92 mmol). Purification by column chromatography
1
(AcOEt) furnished the title compound (30 mg). Yield: 10%. H
NMR (500 MHz, DMSO-d6) δ: 2.73 (t, 2H, J ) 5.4 Hz), 3.13 (t,
2H, J ) 5.8 Hz), 3.37 (s, 6H), 3.54 (s, 3H), 6.72 (s, 2H), 7.37 (d,
1H, J ) 2.2 Hz), 7.52 (dd, 1H, Jm ) 2.3 Hz, Jo ) 7.2 Hz), 7.64 (d,
1H, J ) 8.3 Hz), 7.81 (s, 1H), 7.98 (s, 1H), 9.57 (s, 1H). LC: tR )
6.94 min; m/z: 408.1 (M + H)+ calcd for C22H22N3O5.
2-(3,4,5-Trimethoxyphenylamino)-6-[(2-phenyl)-phenyl-oxy]-
pyrazine (62). Method G was followed with 2-chloro-6-[(2-phenyl)-
phenyl-oxy]-pyrazine (110 mg, 0.39 mmol) and 3,4,5-trimethoxy-
aniline (84 mg, 0.46 mmol). Purification by column chromatography
1
(AcOEt) afforded the title compound (101 mg). Yield: 60%. H
NMR (500 MHz, DMSO-d6) δ: 3.41 (s, 6H), 3.55 (s, 3H), 6.79 (s,
2H), 7.22-7.49 (m, 9H), 7.66 (s, 1H), 7.88 (s, 1H), 9.51 (s, 1H).
13C NMR (62.9 MHz, DMSO-d6) δ: 55.33, 60.03, 95.51, 121.09,
122.25, 125.32, 127.35, 127.99, 128.29, 128.57, 128.91, 130.83,
132.05, 133.73, 136.40, 137.20, 150.59, 152.71, 157.61. LC-MS:
tR ) 8.25 min; m/z: 430.1 (M + H)+ calcd for C25H24N3O4. HRMS:
(M + H)+ calcd for C25H24N3O4, 430.1767; found, 430.1758.
2-(3,4,5-Trimethoxyphenylamino)-6-(2-fluorophenyl-oxy)-pyra-
zine (63). Method G was followed with 2-chloro-6-(2-fluorophe-
nyloxy)-pyrazine (200 mg, 0.89 mmol) and 3,4,5-trimethoxyaniline
(196 mg, 1.07 mmol). Purification by column chromatography
(AcOEt:cyclohexane, 1:1) afforded the title compound (234 mg).
Yield: 71%. 1H NMR (500 MHz, DMSO-d6) δ: 3.42 (s, 6H), 3.54
(s, 3H), 6.69 (s, 2H), 7.24-7.41 (m, 4H), 7.83 (s, 1H), 7.96 (s,
1H), 9.57 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.73, 60.04,
95.77, 116.92, 120.15, 123.79, 125.26, 126.52, 128.47, 132.28,
136.13, 140.27, 150.39, 152.71, 154.80, 156.98, 170.32. LC-MS:
tR ) 7.50 min; m/z: 372.1 (M + H)+ calcd for C19H19FN3O4.
HRMS: (M + H)+ calcd for C19H19FN3O4, 372.1360; found,
372.1363.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-fluorophenyl-oxy)-pyra-
zine (64). Method G was followed with 2-chloro-6-(3-fluorophe-
nyloxy)-pyrazine (200 mg, 0.89 mmol) and 3,4,5-trimethoxyaniline
(196 mg, 1.07 mmol). Purification by column chromatography
(AcOEt:cyclohexane, 1:1) gave the title compound (200 mg). Yield:
61%. 1H NMR (500 MHz, DMSO-d6) δ: 3.44 (s, 6H), 3.55 (s, 3H),
6.77 (s, 2H), 7.02-7.09 (m, 2H), 7.16 (d, 1H, JF ) 8.8 Hz), 7.43
(q, 1H, J ) 8.5 Hz), 7.78 (s, 1H), 7.98 (s, 1H), 9.61 (s, 1H). 13C
NMR (62.9 MHz, DMSO-d6) δ: 55.23, 60.04, 95.80, 108.46,
111.40, 116.74, 121.14, 128.99, 130.94, 132.18, 136.21, 150.49,
152.68, 154.98, 156.73, 161.60, 163.55, 170.32. LC-MS: tR ) 7.82
min; m/z: 372.1 (M + H)+ calcd for C19H19FN3O4. HRMS: (M +
H)+ calcd for C19H19FN3O4, 372.1360; found, 372.1357.
2-(3,4,5-Trimethoxyphenylamino)-6-(5-tetral-1-one-oxy)-pyra-
zine (57). Method G was followed with 2-chloro-6-(tetralon-1-yl-
5-oxy)-pyrazine (200 mg, 0.73 mmol) and 3,4,5-trimethoxyaniline
(160 mg, 0.87 mmol). Purification by column chromatography
1
(AcOEt) afforded the title compound (193 mg). Yield: 63%. H
NMR (500 MHz, DMSO-d6) δ: 1.99-2.02 (m, 2H), 2.60 (t, 2H, J
) 5.9 Hz), 2.77 (t, 2H, J ) 5.9 Hz), 3.41 (s, 6H), 3.55 (s, 3H),
6.69 (s, 2H), 7.40-7.44 (m, 2H), 7.88 (d, 1H, J ) 6.8 Hz), 7.81
(s, 1H), 7.95 (s, 1H), 9.52 (s, 1H). 13C NMR (62.9 MHz, DMSO-
d6) δ: 21.95, 22.97, 55.52, 60.06, 96.28, 120.55, 123.11, 127.20,
128.06, 136.67, 152.72, 197.00. LC-MS: tR ) 7.49 min; m/z: 422.1
(M + H)+ calcd for C23H24N3O5. HRMS: (M + H)+ calcd for
C23H24N3O5, 422.1716; found, 422.1717.
2-(3,4,5-Trimethoxyphenylamino)-6-(6-tetral-1-one-oxy)-pyra-
zine (58). Method G was followed with 2-chloro-6-(tetralon-1-yl-
6-oxy)-pyrazine (200 mg, 0.73 mmol) and 3,4,5-trimethoxyaniline
(160 mg, 0.87 mmol). Purification by column chromatography
(AcOEt) gave the title compound (182 mg). Yield: 59%. 1H NMR
(500 MHz, DMSO-d6) δ: 2.02-2.09 (m, 2H), 2.62 (t, 2H, J ) 6.8
Hz), 2.92 (t, 2H, J ) 5.8 Hz), 3.40 (s, 6H), 3.55 (s, 3H), 6.79 (s,
2H), 7.13 (dd, 1H, Jo ) 8.5 Hz, Jm ) 2.4 Hz), 7.18 (d, 1H, J ) 2.2
Hz), 7.82 (s, 1H), 7.92 (d, 1H, J ) 8.5 Hz), 8.02 (s, 1H), 9.63 (s,
1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 22.72, 28.90, 55.15,
59.71, 60.04, 96.06, 118.66, 119.71, 121.39, 128.62, 129.42, 132.32,
136.12, 147.04, 150.57, 152.66, 156.43, 158.01, 196.28. LC: tR )
7.49 min; m/z: 422.2 (M + H)+ calcd for C23H24N3O5.
2-(3,4,5-Trimethoxyphenylamino)-6-(indanone-4-oxy)-pyra-
zine (59). Method G was followed with 2-chloro-6-(indanone-4-
oxy)-pyrazine (200 mg, 0.77 mmol) and 3,4,5-trimethoxyaniline
(169 mg, 0.92 mmol). Purification by column chromatography
(AcOEt) produced the title compound (71 mg). Yield: 23%. 1H
NMR (500 MHz, DMSO-d6) δ: 2.69 (t, 2H, J ) 3.7 Hz), 2.98 (t,
2H, J ) 5.2 Hz), 3.43 (s, 6H), 3.59 (s, 3H), 6.75 (s, 2H), 7.56-7.62
(m, 3H), 7.90 (s, 1H), 8.04 (s, 1H), 9.62 (s, 1H). 13C NMR (62.9
MHz, DMSO-d6) δ: 22.68, 35.68, 55.45, 60.04, 96.06, 119.46,
120.72, 126.71, 128.54, 129.16, 132.51, 136.11, 139.04, 146.34,
150.51, 151.32, 152.70, 156.89, 205.47. LC: tR ) 7.10 min; m/z:
408.1 (M + H)+ calcd for C22H22N3O5. HRMS: (M + H)+ calcd
for C22H22N3O5, 408.1559; found, 408.1552.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-chlorophenyl-oxy)-pyra-
zine (65). Method G was followed with 2-chloro-6-(3-chlorophe-
nyloxy)-pyrazine (200 mg, 0.83 mmol) and 3,4,5-trimethoxyaniline
(183 mg, 1.00 mmol). Purification by column chromatography
(AcOEt:cyclohexane, 1:1) produced the title compound (165 mg).
Yield: 43%. 1H NMR (500 MHz, DMSO-d6) δ: 3.43 (s, 6H), 3.54
(s, 3H), 6.76 (s, 2H), 7.18 (d, 1H, J ) 6.5 Hz), 7.28 (d, 1H, J )
6.7 Hz), 7.36 (s, 1H), 7.43 (t, 1H, J ) 8.1 Hz), 7.79 (s, 1H), 7.98
(s, 1H), 9.62 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 55.32,
60.04, 95.75, 119.55, 120.80, 121.27, 124.58, 128.98, 131.14,
132.17, 133.71, 136.20, 138.57, 150.47, 152.68, 153.24, 154.69,
156.67, 170.32. LC-MS: tR ) 8.19 min; m/z: 388.1 (M + H)+ calcd
2-(3,4,5-Trimethoxyphenylamino)-6-(indanone-5-oxy)-pyra-
zine (60). Method G was followed with 2-chloro-6-(indanone-5-
oxy)-pyrazine (200 mg, 0.77 mmol) and 3,4,5-trimethoxyaniline

3272 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
for C19H18ClN3O4. HRMS: (M + H)+ calcd for C19H18ClN3O4,
388.1064; found, 388.1051.
(M + H)+ calcd for C25H25N4O5. HRMS: (M + H)+ calcd for
C25H25N4O5, 461.1825; found, 461.1839.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-hydroxyiminoeth-2-yl-
phenoxy)-pyrazine (71) (Method H). 2-(3,4,5-Trimethoxypheny-
lamino)-6-(acetophenone-3-oxy)-pyrazine (90 mg, 0.22 mmol) and
hydrazine hydrate solution 50% (40 µL, 0.96 mmol) were refluxed
for 1.5 h in EtOH (5 mL). The reaction mixture was evaporated to
dryness. Purification by column chromatography (AcOEt:EtOH, 9:1)
furnished the title compound (37 mg). Yield: 41%. 1H NMR (500
MHz, DMSO-d6) δ: 2.61 (s, 3H), 3.39 (s, 6H), 3.54 (s, 3H),
7.18-7.21 (m, 1H), 7.43-7.50 (m, 3H), 7.79 (s, 1H), 7.97 (s, 1H),
9.55 (s, 1H), 11.29 (s, 1H). LC-MS: tR ) 7.29 min; m/z: 410.1(M)+
calcd for C21H22N4O5. HRMS: (M + H)+ calcd for C21H23N4O5,
411.1668; found, 411.1678.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-acetamidophenyl-oxy)-
pyrazine (66). Method G was followed with 2-chloro-6-(3-aceta-
midophenyl-oxy)-pyrazine (150 mg, 0.57 mmol) and 3,4,5-
trimethoxyaniline (125 mg, 0.68 mmol). Crystallization (AcOEt)
gave the title compound (81 mg). Yield: 35%. 1H NMR (500 MHz,
DMSO-d6) δ: 2.03 (s, 3H), 3.44 (s, 6H), 3.56 (s, 3H), 6.80 (s, 2H),
6.82-6.87 (m, 1H), 7.31-7.49 (m, 2H), 7.50 (s, 1H), 7.76 (s, 1H),
7.97 (s, 1H), 9.55 (s, 1H), 10.04 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 23.98, 55.30, 60.03, 95.75, 110.84, 114.83, 121.21,
128.57, 129.75, 132.14, 136.27, 140.69, 150.60, 152.69, 154.20,
157.21, 168.47. LC-MS: tR ) 6.86 min; m/z: 411.2 (M + H)+ calcd
for C21H23N4O5. HRMS: (M + H)+ calcd for C21H23N4O5,
411.1668; found, 411.1677.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-hydroxyiminoeth-2-yl-
phenoxy)-pyrazine (72). Method H was followed with 2-(3,4,5-
trimethoxyphenylamino)-6-(acetophenone-4-oxy)-pyrazine (90 mg,
0.22 mmol) and hydrazine hydrate solution 50% (40 µL, 0.96
mmol). Purification by column chromatography (AcOEt:EtOH, 9:1)
2-(3,4,5-Trimethoxyphenylamino)-6-(4-acetamidophenyl-oxy)-
pyrazine (67). Method G was followed with 2-chloro-6-(4-aceta-
midophenyl-oxy)-pyrazine (150 mg, 0.57 mmol) and 3,4,5-
trimethoxyaniline (125 mg, 0.68 mmol). Crystallization (AcOEt)
furnished the title compound (87 mg). Yield: 37%. 1H NMR (500
MHz, DMSO-d6) δ: 2.07 (s, 3H), 3.43 (s, 6H), 3.55 (s, 3H), 6.77
(s, 2H), 7.13 (d, 2H, J ) 8.9 Hz), 7.60 (d, 2H, J ) 8.9 Hz), 7.73
(s, 1H), 7.93 (s, 1H), 9.51 (s, 1H), 9.98 (s, 1H). 13C NMR (62.9
MHz, DMSO-d6) δ: 23.87, 55.29, 60.04, 95.71, 120.14, 120.92,
128.01, 132.13, 136.35, 148.77, 150.49, 152.70, 157.62, 168.11.
LC-MS: tR ) 6.69 min; m/z: 411.2 (M + H)+ calcd for C21H23N4O5.
HRMS: (M + H)+ calcd for C21H23N4O5, 411.1668; found,
411.1654.
1
afforded the title compound (30 mg). Yield: 33%. H NMR (500
MHz, DMSO-d6) δ: 2.56 (s, 3H), 3.44 (s, 6H), 3.60 (s, 3H), 6.83
(s, 2H), 7.26 (d, 2H, J ) 8.7 Hz), 7.74 (d, 2H, J ) 8.7 Hz), 7.84
(s, 1H), 8.02 (s, 1H), 9.63 (s, 1H), 11.25 (s, 1H). LC-MS: tR
)
7.35 min; m/z: 410.1(M)+ calcd for C21H22N4O5. HRMS: (M +
H)+ calcd for C21H23N4O5, 411.1668; found, 411.1636.
2-(3,4,5-Trimethoxyphenylamino)-6-(indanone-oxime-4-oxy)-
pyrazine (73). Method H was followed (reaction time: 5 h) with
2-(3,4,5-trimethoxyphenylamino)-6-(indanone-4-oxy)-pyrazine (70
mg, 0.17 mmol) and hydrazine hydrate solution (32 µL, 0.51 mmol).
The title compound was obtained (30 mg) by crystallization
(AcOEt). Yield: 42%. 1H NMR (500 MHz, DMSO-d6) δ: 2.78 (m,
2H), 3.36 (m, 2H), 3.38 (s, 6H), 3.53 (s, 3H), 6.72 (s, 2H), 7.18 (d,
1H, J ) 7.6 Hz), 7.33 (t, 1H, J ) 7.7 Hz), 7.44 (d, 1H, J ) 7.3
Hz), 7.80 (s, 1H), 7.96 (s, 1H), 9.57 (s, 1H), 11.02 (s, 1H). 13C
NMR (62.9 MHz, DMSO-d6) δ: 24.55, 25.42, 55.43, 60.03, 95.87,
117.18, 119.46, 120.75, 121.77, 126.71, 128.22, 128.35, 128.78,
129.15, 132.31, 136.10, 138.94, 139.31, 150.59, 152.71, 157.08,
160.49. LC-MS: tR ) 7.24 min; m/z: 423.2 (M + H)+ calcd for
C22H23N4O5. HRMS: (M + H)+ calcd for C22H23N4O5, 423.1668;
found, 423.1654.
2-(3,4,5-Trimethoxyphenylamino)-6-(3-methcarbonylphenyl-
oxy)-pyrazine (68). Method G was followed with 2-chloro-6-
(acetophenone-3-oxy)-pyrazine (200 mg, 0.80 mmol) and 3,4,5-
trimethoxyaniline (176 mg, 0.96 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 9:1) gave the title compound
1
(85 mg). Yield: 27%. H NMR (500 MHz, DMSO-d6) δ: 2.61 (s,
3H), 3.39 (s, 6H), 3.54 (s, 3H), 6.74 (s, 2H), 7.49 (dd, 1H, Jm
2.3 Hz, Jo ) 9.3 Hz), 7.59 (t, 1H, J ) 8.0 Hz), 7.73 (d, 1H, Jm
)
)
2.0 Hz), 7.81 (s, 1H), 7.82 (d, 1H, J ) 6.8 Hz), 7.99 (s, 1H), 9.57
(s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 26.77, 55.32, 60.03,
96.13, 119.90, 121.13, 124.40, 128.74, 130.20, 132.38, 136.13,
138.50, 145.11, 150.48, 152.68, 153.27, 154.02, 156.99, 197.24.
LC-MS: tR ) 7.14 min; m/z: 396.2 (M + H)+ calcd for C21H22N3O5.
HRMS: (M + H)+ calcd for C21H22N3O5, 396.1559; found,
396.1554.
2-(3,4,5-Trimethoxyphenylamino)-6-(tetralonyl-1-oxime-5-oxy)-
pyrazine (74). Method H was followed (reaction time: 4.5 h) with
2-(3,4,5-trimethoxy phenylamino)-6-(tetralon-1-yl-5-oxy)-pyrazine
(100 mg, 0.24 mmol) and hydrazine hydrate solution (44 µL, 0.70
mmol). The title compound was obtained (55 mg) by crystallization
2-(3,4,5-Trimethoxyphenylamino)-6-(4-methcarbonylphenyl-
oxy)-pyrazine (69). Method G was followed with 2-chloro-6-
(acetophenone-4-oxy)-pyrazine (150 mg, 0.60 mmol) and 3,4,5-
trimethoxyaniline (148 mg, 0.72 mmol). Purification by column
chromatography (AcOEt:cyclohexane, 9:1) afforded the title com-
1
(AcOEt). Yield: 53%. H NMR (500 MHz, DMSO-d6) δ: 1.69 (t,
2H, J ) 5.9 Hz), 2.64-2.65 (m, 4H), 3.40 (s, 6H), 3.53 (s, 3H),
6.71 (s, 2H), 7.13 (d, 1H, J ) 7.9 Hz), 7.25 (t, 2H, J ) 7.9 Hz),
7.76 (d, 1H, J ) 5.2 Hz), 7.77 (s, 1H), 7.93 (s, 1H), 9.53 (s, 1H),
11.27 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 20.29, 22.82,
55.54, 60.04, 95.92, 120.21, 120.53, 121.36, 126.70, 127.76, 132.33,
150.55, 151.00, 152.08, 152.72, 157.64, 196.90. LC-MS: tR ) 7.60
min; m/z: 437.2 (M + H)+ calcd for C23H25N4O5. HRMS: (M +
H)+ calcd for C23H25N4O5, 437.1825; found, 437.1812.
1
pound (85 mg). Yield: 34%. H NMR (500 MHz, DMSO-d6) δ:
2.56 (s, 3H), 3.43 (s, 6H), 3.59 (s, 3H), 6.82 (s, 2H), 7.37 (d, 2H,
J ) 8.7 Hz), 7.89 (s, 1H), 8.07 (d, 2H, J ) 6.7 Hz), 8.08 (s, 1H),
9.68 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ: 26.57, 55.33,
60.02, 95.58, 120.24, 121.36, 129.41, 130.24, 132.30, 133.08,
136.12, 150.55, 152.67, 156.50, 157.95, 196.62. LC-MS: tR ) 7.21
min; m/z: 396.2 (M + H)+ calcd for C21H22N3O5. HRMS: (M +
H)+ calcd for C21H22N3O5, 396.1559; found, 396.1567.
2-(3,4,5-Trimethoxyphenylamino)-6-(tetralonyl-1-oxime-6-oxy)-
pyrazine (75). Method H was followed (reaction time: 4.5 h) with
2-(3,4,5-trimethoxy phenylamino)-6-(tetralon-1-yl-6-oxy)-pyrazine
(100 mg, 0.24 mmol) and hydrazine hydrate solution (44 µL, 0.70
mmol). Crystallization (AcOEt) afforded the title compound (32
2-(3,4,5-Trimethoxyphenylamino)-6-(5-methcarbonylaminon-
aphthyl-oxy)-pyrazine (70). Method G was followed with 2-chloro-
6-(5-acetamidonaphthyl-oxy)-pyrazine (200 mg, 0.64 mmol) and
3,4,5-trimethoxyaniline (140 mg, 0.76 mmol). Purification by
column chromatography (AcOEt) produced the title compound (72
mg). Yield: 24%. 1H NMR (500 MHz, DMSO-d6) δ: 2.23 (s, 3H),
3.32 (s, 6H), 3.49 (s, 3H), 6.60 (s, 2H), 7.35 (d, 1H, J ) 7.4 Hz),
7.52 (t, 1H, J ) 7.9 Hz), 7.56 (t, 1H, J ) 7.9 Hz), 7.75 (d, 1H, J
) 7.3 Hz), 7.84 (d, 1H, J ) 8.4 Hz), 7.93 (s, 1H), 7.97 (s, 1H),
7.99 (d, 1H, J ) 7.7 Hz), 9.50 (s, 1H), 9.97 (s, 1H). 13C NMR
(62.9 MHz, DMSO-d6) δ: 23.45, 55.11, 59.96, 95.56, 117.23,
118.39, 119.74, 120.35, 120.93, 122.37, 125.70, 126.16, 127.61,
128.38, 129.24, 132.03, 134.05, 136.17, 145.13, 149.70, 150.64,
152.59, 153.26, 158.19, 168.99. LC-MS: tR ) 7.01 min; m/z: 461.2
1
mg). Yield: 30%. H NMR (500 MHz, DMSO-d6) δ: 1.75 (t, 2H,
J ) 5.2 Hz), 2.61-2.68 (m, 4H), 3.40 (s, 6H), 3.54 (s, 3H), 6.79
(s, 2H), 7.03-7.18 (m, 2H), 7.78 (s, 1H), 7.87 (d, 1H, J ) 8.4
Hz), 7.96 (s, 1H), 9.60 (s, 1H), 11.10 (s, 1H). 13C NMR (62.9 MHz,
DMSO-d6) δ: 20.93, 22.78, 28.91, 55.22, 60.04, 95.92, 118.72,
120.02, 121.17, 124.89, 127.66, 128.57, 132.37, 136.27, 150.58,
151.89, 152.69, 157.10, 196.27. LC-MS: tR ) 7.72 min; m/z: 437.2
(M + H)+ calcd for C23H25N4O5. HRMS: (M + H)+ calcd for
C23H25N4O5, 437.1825; found, 437.1822.
2-(3,4,5-Trimethoxyphenylamino)-6-(4-methaminocarbonyl-naph-
thyloxy)-pyrazine (76). Using method E with N-(4-hydroxynaph-
thalen-1-yl)acetamide (200 mg, 0.99 mmol), the title compound

Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11 3273
was obtained (31 mg). Yield: 6.7%. 1H NMR (500 MHz, DMSO-
d6) δ: 2.21 (s, 3H), 3.20 (s, 9H), 3.48 (s, 3H), 6.61 (s, 2H), 7.31
(d, 1H, J ) 8.2 Hz), 7.59 (t, 2H, J ) 7.6 Hz), 7.70 (d, 1H, J ) 8.3
Hz), 7.91 (s, 1H), 7.96-7.99 (m, 2H), 8.13 (d, 1H, J ) 7.8 Hz),
9.49 (s, 1H), 9.93 (s, 1H). 13C NMR (62.9 MHz, DMSO-d6) δ:
23.44, 55.10, 59.95, 95.38, 116.77, 120.89, 121.48, 121.61, 123.18,
126.38, 126.53, 127.11, 128.23, 128.71, 130.90, 131.98, 136.18,
146.44, 150.64, 152.59, 158.26, 168.93. LC-MS: tR ) 7.29 min;
m/z: 461.1 (M + H)+ calcd for C25H25N4O5. HRMS: (M + H)+
calcd for C25H25N4O5, 461.1825; found, 461.1811.
1/1000, Perkin-Elmer, Turku, Finland) were preincubated for 30
min and 50 µL added to the washed plates, which were incubated
for a further hour. The plates were washed as before, and 100 µL
DELFIA enhancement solution (Perkin-Elmer, Turku, Finland)
added. The plates were sealed and incubated for 30 min and
europium counts measured on a Victor 2 reader (Perkin-Elmer,
Turku, Finland).
Acknowledgment. This work is supported by Cancer
Research UK (refs: C309/A2187 and C107/A3096), the Wellcome
Trust, and The Institute of Cancer Research. We thank Professor
Paul Workman for helpful discussions and support and John
Harris, Biofocus, for useful discussions. We also thank Meirion
Richards for his help in evaluating the purity of the compounds
and the accurate mass.
2[(3-Oxazol-5-yl)phenylamino]-6-(5-methaminocarbonylnaph-
thyl-oxy)pyrazine (77). Using method D (conventional heating,
reaction time: 20 h) with 2-chloro-6-(5-acetamidonapht-1-yloxy)-
pyrazine (150 mg, 0.48 mmol) and 3-(1,3-oxazol-5-yl-aniline) (96
mg, 0.60 mmol), the title compound was obtained (71 mg). Yield:
34%. 1H NMR (500 MHz, DMSO-d6) δ: 2.22 (s, 3H), 6.95 (t, 1H,
J ) 7.9 Hz), 7.06 (d, 1H, J ) 8.5 Hz), 7.15 (d, 1H, J ) 7.6 Hz),
7.44-7.46 (m, 2H), 7.49-7.52 (m, 2H), 7.57 (d, 1H, J ) 7.6 Hz),
7.74 (d, 1H, J ) 6.6 Hz), 7.77 (d, 1H, J ) 8.2 Hz), 7.94 (s, 1H),
7.96 (s, 1H), 8.02 (d, 1H, J ) 8.4 Hz), 8.36 (s, 1H), 9.68 (bs, 1H),
9.99 (bs, 1H). 13C NMR (125.8 MHz, DMSO-d6) δ: 23.44, 113.35,
117.19, 117.82, 117.99, 118.40, 120.07, 120.86, 121.74, 122.02,
125.49, 126.10, 127.53, 127.61, 127.69, 129.22, 134.08, 140.57,
149.18, 149.98, 150.28, 151.61, 154.70, 158.28, 168.90. LC: tR )
7.20 min; m/z: 438.1 (M + H)+ calcd for C25H20N5O3. HRMS: (M
+ H)+ calcd for C25H20N5O3, 438.1560; found, 438.1564.
2-(3-oxo-1,3-Dihydroisobenzofuran-5-ylamino)-6-(5-methami-
nocarbonylnaphthyl-oxy) pyrazine (78). Using method D with
2-chloro-6-(5-acetamidonaphthyl-oxy)-pyrazine (150 mg, 0.48 mmol)
and 6-amino-1,3-dihydroisobenzofuran-1-one (89 mg, 0.60 mmol),
the title compound was obtained (8 mg). Yield: 4%. 1H NMR (500
MHz, DMSO-d6) δ: 2.21 (s, 3H), 5.24 (s, 2H), 7.17 (d, 1H, J )
8.5 Hz), 7.45-7.51 (m, 3H), 7.63 (d, 1H, J ) 8.2 Hz), 7.68-7.75
(m, 3H), 7.92 (s, 1H), 7.98 (s, 1H), 8.05 (d, 1H, J ) 8.7 Hz), 9.92
(bs, 1H), 10.00 (bs, 1H). LC-MS: tR ) 6.72 min; m/z 427.1 (M +
H)+ calcd for C24H18N4O4.
Supporting Information Available: Tables of purity and HPLC
traces for final compounds. This material is available free of charge
via the Internet at http://pubs.acs.org.
References
(1) Wellbrock, C.; Karasarides, M.; Marais, R. The RAF proteins take
center stage. Nat. ReV. Mol. Cell Biol. 2004, 5, 875–885.
(2) Davies, H.; Bignell, G. R.; Cox, C.; Stephen, P.; et al. Mutation of
the BRAF gene in human cancers. Nature 2002, 417, 949–954.
(3) Tuveson, D. A.; Weber, B. L.; Herlyn, M. BRAF as a potential
therapeutic target in melanoma and other malignancies. Cancer Cell
2003, 4, 95–98.
(4) Garnett, M. J.; Marais, R. Guilty as charged: B-RAF is a human
oncogene. Cancer Cell 2004, 4, 313–319.
(5) Chiloeches, A.; Marais, R. Mitogen-activated protein kinase cascades
as therapeutic targets in cancer. Targets for Cancer Chemotherapy:
Transcription Factors and Other Nuclear Proteins; Humana Press Inc.:
Totowa, NJ, 2002; pp 179-206.
(6) Wood, E.; Crosby, R. M.; Dickerson, S.; Frye, S. V.; Griffin, R.; et
al. A prodrug approach to the design of cRaf1 kinase inhibitors with
improved cellular activity. Anti-Cancer Drug Des. 2001, 16, 1–6.
(7) Hall-Jackson, C. A.; Eyers, P. A.; Cohen, P.; Boyle, F. T.; Hewitt,
N.; et al. Paradoxical activation of Raf by a novel Raf inhibitor. Chem.
Biol. 1999, 6, 559–568.
(8) Wilhelm, S. M.; Carter, C.; Tang, L. Y.; Wilkie, D.; McNabola, A.;
et al. BAY 43-9006 exhibits broad spectrum oral antitumor activity
and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases
involved in tumor progression and angiogenesis. Cancer Res. 2004,
64, 7099–7109.
(9) Bollag, G.; Freeman, S.; Lyons, G. F.; Post, L. E. Raf pathway
inhibition in oncology. Curr. Opin. InVestig. Drugs 2004, 4, 1436–
1441.
(10) Smith, R. A.; Dumas, J.; Adnane, L.; Wilhelm, S. M. Recent advances
in the research and development of RAF kinase inhibitors. Curr. Top.
Med. Chem. 2006, 6, 1071–1089.
(11) Batt, D. B.; Bold, G.; Kim, S.; Ramsey, T. M.; Sabio, M. L. Use of
Isoquinoline DeriVatiVes for Treating Cancer and MAP Kinase Related
Diseases; Novartis AG: Basel, Switzerland, 2004; p 85.
(12) Fink, C. A.; Perez, L. B.; Ramsey, T. M.; Yusuf, N.; Versace, R. W.
1,4-Disubstituted Isoquinoline DeriVatiVes as RAF-Kinase Inhibitors
Useful for the Treatment of ProliferatiVe Diseases; Novartis AG: Basel,
Switzerland, 2005; p 144.
(13) Takle, A. K.; Brown, M. J. B.; Davies, S.; Dean, D. K.; Francis, G.;
et al. The identification of potent and selective imidazole-based
inhibitors of B-RAF kinase. Bioorg. Med. Chem. Lett. 2006, 16, 378–
381.
(14) King, A. J.; Patrick, D. R.; Batorsky, R. S.; Ho, M. L.; Do, H. T.; et
al. Demonstration of a genetic therapeutic index for tumors expressing
oncogenic BRAF by the kinase inhibitor SB-590885. Cancer Res.
2006, 66, 11100–11105.
(15) Ahmad, T.; Eisen, T. Kinase inhibition with BAY 43-9006 in renal
cell carcinoma. Clin. Cancer Res. 2004, 10, 6388S–6392S.
(16) Niculescu-Duvaz, I.; Roman, E.; Wittaker, S. R.; Friedlos, F.; Kirk,
R.; et al. Novel inhibitors of B-RAF based on a disubstituted pyrazine
scaffold. Generation of a nanomolar lead. J. Med. Chem. 2006, 49,
407–416.
2[(3-Oxazol-5-yl)phenylamino]-6-(4-methaminocarbonylnaph-
thyl-oxy)pyrazine (79). Using method D with 2-chloro-6-(4-
acetamidonaphthalen-1-yloxy)pyrazine (150 mg, 0.48 mmol) and
3-(1,3-oxazol-5-yl-aniline) (115 mg, 0.71 mmol), the title compound
1
was obtained (97 mg). Yield: 46%. H NMR (500 MHz, DMSO-
d6) δ: 2.22 (s, 3H), 6.95 (t, 1H, J ) 7.9 Hz), 7.10 (d, 1H, J ) 8.2
Hz), 7.14 (d, 1H, J ) 7.7 Hz), 7.41 (d, 1H, J ) 8.2 Hz), 7.44 (bs,
2H), 7.54 (t, 1H, J ) 7.5 Hz), 7.60 (t, 1H, J ) 7.4 Hz), 7.70 (d,
1H, J ) 8.1 Hz), 7.91 (d, 1H, J ) 8.5 Hz), 7.95 (bs, 2H), 8.16 (d,
1H, J ) 8.4 Hz), 8.37 (s, 1H), 9.70 (bs, 1H), 9.99 (bs, 1H). 13C
NMR (125.8 MHz, DMSO-d6) δ: 23.43, 113.25, 117.10, 117.70,
117.81, 120.71, 121.66, 121.77, 123.32, 126.27, 126.53, 127.31,
127.45, 127.53, 128.92, 129.39, 131.23, 140.63, 146.10, 149.99,
150.27. LC-MS: tR ) 7.29 min; m/z: 438.1 (M + H)+ calcd for
C19H20N5O3. HRMS: (M + H)+ calcd for C19H20N5O3, 438.1566;
found, 438.1565.
V600EBRAF Kinase Assay. Full-length rabbit MEK1 protein was
expressed with a GST tag at the N-terminus and a C-terminal
histidine tag in Escherichia coli JM109 bacteria and purified by
nickel-agarose affinity chromatography. V600EBRAF was generated
by infection of SF9 insect cells cultured in SF-900 II medium
(Invitrogen, Paisley, Scotland) with a baculovirus containing full-
length human BRAF with an N-terminal histidine tag. Assay buffer
was 20 mM MOPS, pH 7.2 containing 5 mM EGTA, 10 mM
MgCl2, 0.1% ꢀ-ME and 25 mM ꢀ-glycerophosphate. All incuba-
tions were at room temperature with shaking. One µg GST-MEK1,
0.07 µL of insect cell V600EBRAF lysate, and 0.5µL of inhibitor at
the required concentrations (0.001 to 100 µM final concentration)
were added to the wells of a glutathione-coated plate and the plate
preincubated for 10 min. ATP in assay buffer (10 µL), to give a
final concentration of 100 µM, was added to each well and the
plates incubated for 45 min. The plates were then washed 3× with
200 µL of 0.1% Tween 20/water. Primary antibody (rabbit
antiphospho MEK1/2 diluted 1/2000, Cell Signaling Technologies,
Hitchin, UK) and Eu-labeled antirabbit secondary antibody (diluted
(17) Wan, P. T. C.; Garnett, M. J.; Roe, M. S.; Lee, S.; Niculescu-Duvaz,
D.; et al. Mechanism of activation of of the RAF/ERK signaling
pathway by oncogenic mutations of B-RAF. Cell 2004, 116, 855–
867.
(18) Liao, J. J.-L. Molecular recognition of protein kinase binding pockets
for design of potent and selective kinase inhibitors. J. Med. Chem.
2007, 50, 409–424.

3274 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 11
Niculescu-DuVaz et al.
(19) Wolfe, J. P.; Tomori, H.; Sadighi, J. P.; Yin, J.; Buchwald, S. L.
Simple, efficient catalyst system for the palladium-catalyzed amination
of aryl chlorides, bromides and triflates. J. Org. Chem. 2000, 65, 1158–
1174.
(20) Grasa, G. A.; Viciu, M. S.; Huang, J.; Nolan, S. P. Amination reaction
of aryl halides with nitrogen-containing reagents mediated by pal-
ladium/imidazolium salts. J. Org. Chem. 2001, 66, 7729–7737.
(21) Kotha, S.; Lahiri, K.; khashinath, D. Recent applications of the
Suzuki-Miyaura cross coupling reactions in organic synthesis. Tet-
rahedron 2002, 58, 9633–9695.
(22) Littke, A.; Fu, G. C. Palladium catalysed coupling reaction of aryl
chlorides. Angew. Chem., Int. Ed. 2002, 41, 4176–4211.
(23) Tokarski, J. S.; Newitt, J. A.; Chang, C. Y. J.; Cheng, J. D.; Witteking,
M.; et al. The structure of Dasatinib (BMS-354825) bound to activated
ABL kinase domain elucidates its inhibitory activity against Imatinib-
resistant ABL mutants. Cancer Res. 2006, 66, 5790–5797.
JM070776B