4-Anilino-7,8-dialkoxybenzo[g]quinoline-3-carbonitriles as potent Src kinase inhibitors

Article abstract of DOI:10.1021/jm050512u

It has been previously reported that appropriately substituted 4-anilinoquinoline-3-carbonitriles are potent inhibitors of Src kinase, with biological activity in vitro and in vivo. Structural modifications to these compounds have been explored, providing

Full text of DOI:10.1021/jm050512u

J. Med. Chem. 2005, 48, 5909-5920
5909
4-Anilino-7,8-dialkoxybenzo[g]quinoline-3-carbonitriles as Potent Src Kinase
Inhibitors
Dan M. Berger,*,† Minu Dutia,† Gary Birnberg,† Dennis Powell,† Diane H. Boschelli,† Yanong D. Wang,†
Malini Ravi,† Deanna Yaczko,† Jennifer Golas,‡ Judy Lucas,‡ and Frank Boschelli‡
Chemical & Screening Sciences and Oncology, Wyeth Research, 401 N. Middletown Road, Pearl River, New York 10965
Received June 1, 2005
It has been previously reported that appropriately substituted 4-anilinoquinoline-3-carbonitriles
are potent inhibitors of Src kinase, with biological activity in vitro and in vivo. Structural
modifications to these compounds have been explored, providing the 4-anilinobenzo[g]quinoline-
3-carbonitriles as a series with enhanced Src inhibitory properties. The synthesis and structure-
activity relationships of these 4-anilino-7,8-dialkoxybenzo[g]quinoline-3-carbonitriles are
presented here. Analogues with cyclic basic amine groups attached via ethoxy linkages at the
C-8 position were the most active in vitro, with subnanomolar IC50 values against Src kinase
observed for a majority of the compounds synthesized. Compound 17d was more potent in
vitro than the analogously substituted 4-anilinoquinoline-3-carbonitrile SKI-606, which is
undergoing evaluation in clinical trials. The most potent analogue synthesized was 17a, with
an IC50 of 0.15 nM against Src kinase and with an IC50 of 10 nM against Src-transformed
fibroblasts. Molecular modeling studies provided a rationale for the exceptional activity observed
for these compounds, with favorable van der Waals interactions playing the major role.
Compound 17c was found to be highly selective for Src kinase when tested against a panel of
other kinases, with modest selectivity versus the Src family kinases Lyn and Fyn. Following
ip dosing at 50 mg/kg, analogues 17c and 17d were shown to have plasma levels that
significantly exceeded the cellular IC50 values against Src-transformed fibroblasts. In an Src-
transformed fibroblast xenograft model, both compounds exhibited a significant inhibition of
tumor growth.
Introduction
5,10-dihydropyrimido[4,5-b]quinolines,13 4-anilinoquin-
azolines,14 and 4-anilino-3-quinolinecarbonitriles.15a-j
Optimization of a series of 6,7-dialkoxy substituted
4-anilino-3-quinolinecarbonitriles provided SKI-606 as
a potent inhibitor of Src kinase, with oral activity in
xenograft models.15c,g Modifications of the 3-quinolin-
ecarbonitrile core were explored to provide 8-anilinoimi-
dazo[4,5-g]quinoline-7-carbonitriles,16 4-anilinobenzothi-
eno[3,2-b]pyridine-3-carbonitriles and 4-anilinobenzo-
furo[3,2-b]pyridine-3-carbonitriles,17 4-anilinobenzo[b]-
naphthyridine-3-carbonitriles,18 4-anilinobenzo[g]quin-
oline-3-carbonitriles,19 and 7-anilinothieno[3,2-b]pyridine-
6-carbonitriles20a,b as Src kinase inhibitors. Of these, the
most potent were the 4-anilinobenzo[g]quinoline-3-car-
bonitriles with 7,8-dimethoxy substituents. The 7,8-
dimethoxy substituted compound 1 (Chart 1) was more
active in Src enzyme and cellular assays than the
correspondingly substituted quinoline-3-carbonitrile.19
We set out to further improve the solubility and activity
of these 4-anilinobenzo[g]quinoline-3-carbonitriles by
adding cyclic basic amines via the C-7 or C-8 positions.
Preliminary presentations have highlighted the excep-
tional activity of several of these analogues against Src
kinase.21a-c
Protein tyrosine kinases (TKs) catalyze the phospho-
rylation of a tyrosine residue on a substrate protein, a
process important for cell growth and differentiation.
Src kinase is a member of a structurally homologous
group of nonreceptor TKs present in the cytoplasm
known as the Src family of kinases.1 Src itself partici-
pates in signaling pathways controlling proliferation,
differentiation, and migration.2 It has been demon-
strated that Src is overexpressed or constitutively active
in a variety of human tumors, including those derived
from colon, breast, pancreas, liver, brain, and bladder.3
Inhibition of Src kinase has been shown to decrease the
hypoxic induction of VEGF, a protein that is up-
regulated in angiogenesis.4a,b Thus, the evidence sug-
gests that the inhibition of Src activity may prove to be
useful for therapeutic intervention in cancer. Other
studies have provided evidence that Src inhibition might
be effective in the treatment of other diseases, such as
osteoporosis5a,b and stroke.6
Efforts by researchers to discover small-molecule
inhibitors of Src kinase have generated several classes
of structures as potent leads, including purines,7 pyrido-
[2,3-d] pyrimidines,8 pyrrolo[2,3-d]pyrimidines,9a-e pyra-
zolo[2,3-d]pyrimidines,9e,10 indolin-2-ones,11 2-methylpy-
rimidin-4-ylamino)thiazole-5-carboxamides,12a,b 4-anilino-
The present work describes the synthesis and detailed
SARs of these 4-anilino-7,8-dialkoxybenzo[g]quinoline-
3-carbonitriles. Structurally, they are described by the
general formula 2 shown in Chart 1.
* To whom correspondence should be addressed. Phone: (845) 602-
3435. Fax: (845) 602-5561. E-mail: Bergerd@wyeth.com.
† Chemical & Screening Sciences.
Chemistry
In our initial synthesis of the target compounds, we
chose a pathway that would provide the compounds as
‡ Oncology.
10.1021/jm050512u CCC: $30.25 © 2005 American Chemical Society
Published on Web 08/25/2005

5910 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19
Berger et al.
Chart 1
Scheme 1. Preparation of 16a-i (Table 2) and 17a-k
(Table 3)a
a regioisomeric mixture at the C-7 and C-8 positions. It
was anticipated that each product mixture would be
separable by chromatographic methods to provide two
analogues for biological evaluation. On the basis of the
fact that water-solubilizing substituents at the C-7
position of the 4-anilinoquinoline-3-carbonitriles provide
optimal in vitro activity,15b it seemed likely that the C-8
position would similarly be optimal for the 4-anilinoben-
zo[g]quinoline-3-carbonitriles. However, we decided to
confirm this before embarking on the challenge of
carrying out a regioselective synthesis of the target
compounds. To compensate for the larger benzo[g]-
quinoline-3-carbonitrile core, we chose to attach the
cyclic amine substituents via ethoxy linkers rather than
the propoxy linkers generally used for the quinoline-3-
carbonitrile scaffold.
The general method by which most of the 7,8-
substituted 4-anilinobenzo[g]quinoline-3-carbonitriles
were synthesized is outlined in Scheme 1. The reaction
of 2-methoxyphenol 3 with 2-chloroethyl p-toluene-
sulfonate provided 1-(2-chloroethoxy)-2-methoxybenzene
4. On heating with formaldehyde and hydrochloric acid
(gas), 4 was converted to the 4,5-dichloromethyl sub-
stituted intermediate 5, which on treatment with so-
dium acetate in acetic acid and subsequent hydrolysis
with ammonia-saturated methanol gave 6. Swern oxi-
dation of the diol 6 yielded phthalaldehyde 7. Reaction
of 7 with an excess of ethyl 3-nitropropanoate and
sodium ethoxide in ethanol provided an inseparable
mixture of 3-nitro-2-naphthoate22 intermediates, which
were reduced by catalytic hydrogenation to provide
3-amino-2-naphthoates 8 and 9 (approximately 1:1
mixture). Synthesis of the 3-nitro-2-naphthoate inter-
mediates was generally carried out on a small scale (<3
g) because larger scale reactions occasionally failed to
provide the desired products, instead resulting in the
complete decomposition of starting materials. Com-
pounds 8 and 9 were heated in dimethylformamide
dimethylacetal to provide the corresponding dimethyl-
aminomethyleneamine intermediates, which were im-
mediately reacted with the anion of acetonitrile to
provide benzoquinoline-3-carbonitriles 10 and 11.15b
Chlorination of 10 and 11 was carried out with phos-
phorus oxychloride to yield the 4-chloro substituted
intermediates 12 and 13. Compounds 14 and 15 were
a Reagents and reaction conditions: (a) chloroethyl tosylate,
K2CO3, 2-butanone, reflux, 2 days; (b) formaldehyde, ether, HCl
gas addition, 0 °C, 9 h, room temperature, 16 h; (c) NaOAc, acetic
acid, reflux, 2 h; (d) NH3, MeOH, 0-5 °C, 15 h; (e) oxalyl chloride,
DMSO, CH2Cl2, -78 °C, 30 min, then NEt3, 10 min; (f) NaOEt,
O2NCH2CH2CO2Et, EtOH, 0-5 °C, 10 min, then room tempera-
ture, 16 h; (g) H2 (Parr, 50 lb/in2), DMF, 2 h; (h) DMF-dimethyl
acetal, reflux, 16 h; (i) CH3CN, n-BuLi, THF, -78 °C, 30 min; (j)
POCl3, reflux, 20 min; (k) ArNH2, pyridine-HCl, 2-ethoxyethanol,
135 °C, 30 min to 1 h; (l) RRNH, NaI, neat, reflux 30 min, or in
CH3OCH2CH2OCH3, reflux, 3.5 h to 4 days.
obtained by heating 12 and 13 with the appropriate
aniline15c and pyridine hydrochloride in 2-ethoxyetha-
nol. The final products 16 and 17 were obtained follow-
ing addition of the cyclic basic amines at the C-7 and
C-8 positions. These compounds all proved to be readily
separable by silica gel chromatography (CH2Cl2/MeOH
or EtOAc/MeOH). In every case, analogues 17 were the
more polar of the two isomers. For selected analogues,
two-dimensional rotating-frame Overhauser enhance-
ment spectroscopy (ROESY) NMR data were obtained
on the isolated compounds to confirm the structural
assignments. While this work was in progress, a 12-
step reaction sequence was elucidated to regioselectively
provide compound 17c.23 This chemistry was utilized
for the synthesis of 17c and 17j as further confirmation
that the structures of the separated isomers had been
correctly assigned, as well as providing sufficient quan-
tities of 17c for in vivo studies.
Molecular Modeling
To gain insight on how the binding of the 4-anili-
nobenzo[g]quinoline-3-carbonitriles to Src kinase would
compare to the 4-anilinoquinoline-3-carbonitriles, mo-

Carbonitriles as Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19 5911
Figure 1. Binding orientations of 17d (atoms are colored by atom type with the carbon atoms colored green) and SKI-606 (atoms
are colored orange) to the hinge region within the ATP-binding site of Src kinase.
Table 1. Calculated Binding Energies of 17d and SKI-606
total energy
(kcal)
screened electrostatic energy (kcal)
hydrophobic
energy (kcal)
van der Waals
energy (kcal)
rotational
penalty (kcal)
structure
(Coulomb + solvation)
17d
SKI-606
-28.0
-24.6
3.6
4.5
-8.2
-7.8
-29.0
-27.6
5.6
6.3
lecular modeling studies were carried out using a
homology model (a crystal structure of the active form
of c-Src was not available) based on the active form of
the Lck kinase domain (PDB ID: 1QPE). Compounds
17d and SKI-606 were chosen as representative ex-
amples possessing the same aniline and cyclic amine
substituents. The scoring function utilized (described in
the Experimental Section) consists of a Poisson-Boltz-
mann based treatment of the electrostatic interactions
in the protein-ligand complex. The binding mode
presented here is selected on the basis of binding
interactions that result in optimal energetics as given
in Table 1. The screened electrostatics term in this
model measures the gain in favorable electrostatic
interactions between protein and ligand and includes a
penalty for buried polar groups due to the cost of
removing the ligand from solvent. The hydrophobic term
is the energetic reward for the buried hydrophobic
interactions. With tighter protein-ligand binding, a
greater amount of surface area is expected to be buried
away from solvent, making the interaction more ener-
getically favorable. A rotational penalty term is com-
puted as follows: (0.7)(number of rotatable bonds). This
is an approximate measure of the internal coordinate
entropy of the ligand.
more favorable electrostatic energy could be associated
with the closer approach of the 17d core (versus the
SKI-606 core) with the protein’s hinge region residues.
Favorable interactions between the core ring nitrogen
and the backbone NH of Met 319 were significant in
our model. Additionally, the more optimal interaction
of the Tyr 318 ring with the core of 17d enhanced the
van der Waals energy term for the interaction of the
protein-ligand complex. In the low-energy pose shown
(Figure 2), the center aromatic ring of the tricyclic core
approached the Tyr 318 ring to within 3.5 Å. In contrast,
the closest approach of the bicyclic core of SKI-606 to
the Tyr 318 ring was 4.0 Å. Allowing for the movement
of both protein and ligand, a greater area of aromatic
surface on the tricyclic core was available for interaction
with the Tyr 318 aromatic ring in comparison with the
bicyclic core of SKI-606. Last, a slight enhancement of
binding energy was afforded by the smaller rotational
penalty assigned to the less flexible C-8 substituent of
ligand 17d.
Biological Results
SAR for the Src Kinase and Src Cellular Assays.
The inhibitory activity of compounds 16a-i (Table 2)
and 17a-m (Table 3) against Src kinase was mea-
sured using an enzyme linked immunosorbent assay
(ELISA).15b A Src-transformed fibroblast line was used
to measure antiproliferative cellular activity.15b A clear
trend immediately observed from the data was that the
analogues bearing cyclic basic amine groups attached
via ethoxy linkers at C-8 (compounds 17) were signifi-
cantly more potent Src kinase inhibitors than analogous
compounds with cyclic amines attached at the C-7
position (compounds 16). Furthermore, compounds 17
were all exceptionally active, with the majority of the
The largest differences in energy metrics for 17d and
SKI-606 were apparent in the screened electrostatics
and the van der Waals terms. Notably, the gain in van
der Waals contributions was more significant than the
corresponding gain in the screened electrostatics. That
is, a greater overall contribution from favorable non-
bonded interactions between protein and ligand was
predicted for 17d than for SKI-606.
The corresponding binding orientations for the two
ligands are shown in Figure 1. This showed that the

5912 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19
Berger et al.
Figure 2. van der Waals surface view of Src kinase domain with SKI-606 (all atoms are colored orange) and 17d (atoms are
colored by atom type with the carbon atoms colored green) docked. The surface is shown in translucent mode such that the
location of the Tyr 318 side chain is apparent. The atoms of the Tyr residue are colored by atom type with the carbon atoms
colored white.
Table 2. Inhibition of Src Kinase Activity and Cell
Proliferation by SKI-606, 1, and
Table 3. Inhibition of Src Kinase Activity and Cell
Proliferation by 4-Anilinobenzo[g]quinoline-3-carbonitriles
Substituted with Cyclic Basic Amine Groups Attached at C-8
(17a-k)a
4-Anilinobenzo[g]quinoline-3-carbonitriles Substituted with
Cyclic Basic Amine Groups Attached at C-7 (16a-i)a
IC50 (nM)
Src Src
ELISA cell
IC50 (nM)
Src
Src
compd
RRN
X
compd
NRR
X
ELISA cell
SKI-606
1
1.2
1.4
2.1
1.5
1.8
1.1
2.1 1600
2.5 110
2.6 2000
100
210
71
220
230
260b
17a
17b
17c
17d
17e
17f
17g
17h
17i
morpholine
2-CH3, 4-Cl, 5-OCH3
2-CH3, 4-Cl, 5-OCH3
2,4-diCl, 5-OCH3
0.15
0.31
0.46
0.29
0.72
0.33
0.55
0.48
0.69
0.22b
2.0
10
42
4-methylpiperazine
morpholine
16a
16b
16c
16d
16e
16f
16g
16h
16i
morpholine
2-CH3, 4-Cl, 5-OCH3
37
4-methylpiperazine 2-CH3, 4-Cl, 5-OCH3
4-methylpiperazine
2,4-diCl, 5-OCH3
61
350
19
28
22
270
50
48
morpholine
4-methylpiperazine 2,4-diCl, 5-OCH3
4-hydroxypiperidine 2,4-diCl, 5-OCH3
morpholine
4-methylpiperazine 3,4,5-triOCH3
morpholine
morpholine
2,4-diCl, 5-OCH3
4-hydroxypiperidine 2,4-diCl, 5-OCH3
morpholine
2-Cl, 4-CH3, 5-OCH3
4-hydroxypiperidine 2-Cl, 4-CH3, 5-OCH3
2-Cl, 4-CH3, 5-OCH3
morpholine
2-Cl, 4-F, 5-OCH3
3,4,5-triOCH3
3,4,5-triOCH3
2,4-diCl
4-methylpiperazine
morpholine
3,4,5-triOCH3
2,4-diCl
1.2
13
180
970
17j
17k
morpholine
a IC50 values reported represent the mean of at least two
separate determinations. b IC50 values are from a single determi-
nation.
a IC50 values reported represent the mean of at least two
separate determinations. b IC50 values are from a single determi-
nation.
1, while 16a and 16b had overall activity comparable
to that of 1.
synthesized analogues having subnanomolar activity
against Src kinase and having exceptional cellular
activity as well (IC50 < 100 nM). Overall, these com-
pounds were more potent than the correspondingly
substituted 4-anilinoquinoline-3-carbonitriles.15a-c For
example, analogue 17d was more potent in vitro than
SKI-606, the analogously substituted 3-quinolinecarbo-
nitrile. As analogously observed with the 4-anilino-
quinoline-3-carbonitriles, basic amine groups attached
to C-8 of the benzoquinoline template enhanced both
enzyme and cellular activity. Compounds 17a and 17b
were more potent against Src kinase and Src-trans-
formed fibroblasts than the 7,8-dimethoxy substituted
The synthesized analogues utilized substituted anilines
that were previously explored in SAR studies of the
4-anilinoquinoline-3-carbonitriles.15a,c,f The differences
between the activities of these analogues within a series
were relatively small, although the 2-methyl-4-chloro-
5-methoxyaniline group of 17a appeared to provide the
best activity (Src enzyme IC50 ) 0.15 nM; Src cell IC50
) 10 nM). The 2,4,5-trisubstituted anilines and the
3,4,5-trimethoxyaniline groups appeared to provide
comparable activity, with significantly better activity
against Src kinase than the corresponding 2,4-dichloro
substituted analogues 16i and 17k. Of the cyclic basic

Carbonitriles as Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19 5913
(IC50 > 10 µM), MK2 (IC50 > 20 µM), AKT (IC50 > 10
µM), and IKK (no inhibition at 3.6 µM). Other kinase
studies were carried out in cell-based systems with rat
fibroblasts.15b,g In Abl transformed rat fibroblasts, 17c
was found to have modest activity, with an IC50 of 0.49
µM.21b In contrast, SKI-606 was a more potent inhibitor,
with an IC50 of 0.09 µM. As was observed for SKI-606,
17c demonstrated good selectivity versus non-Src family
kinases but modest selectivity versus selected Src family
kinases. Thus, for example, 17c had IC50 values of 0.14
µM versus Lyn-transformed fibroblasts and 0.42 µM
versus Fyn-transformed fibroblasts, respectively. This
represents 3.8-fold and 11.4-fold selectivity of 17c for
Src over Lyn and Fyn in the cellular proliferation
assays.
Additional Cellular Assays. The antiproliferative
activities of compounds 17c and 17d were determined
using HT29 cells, whose growth in vivo is reduced by
administration of SKI-60615c and antisense Src.25 When
the HT29 cells were grown on plastic, little inhibition
was observed at concentrations lower than 5 µM for both
compounds. A similar result was observed when 17c and
17d were tested against Colo205 cells.24 However,
submicromolar concentrations of 17c and 17d were
sufficient to significantly decrease HT29 cellular pro-
liferation on soft agar (Figure 4). This activity mirrors
that observed with SKI-606.15c
To determine the effect of 17c on Src activity in
human colon tumor cells, we examined the effect of 17c
on tyrosine phosphorylation of focal adhesion kinase
(FAK) in Colo205 cells. FAK is phosphorylated by Src
on a number of tyrosines, including Y925. As shown in
Figure 5, phosphorylation of FAK on Y925 in Colo205
colon tumor cells was reduced in a dose-dependent
manner by 17c at 100, 500, and 1000 nM, consistent
with the inhibition of Src activity observed in fibro-
blasts.
Plasma Level Determination. To determine whether
17c and 17d would have sufficient exposure levels to
warrant investigation of their antitumor properties in
vivo, the plasma levels of both compounds were deter-
mined. The mice were dosed ip at 50 mg/kg with each
compound, and plasma levels were measured at four
time points (Table 4). The results show that both
compounds had substantial blood levels up to 24 h after
dosing, with 17c initially showing higher exposure levels
than 17d but a more rapid drop in plasma concentration
over time. Both compounds had sustained plasma levels
that significantly exceeded the IC50 for Src-transformed
cellular proliferation for at least 8 h and were therefore
evaluated in vivo.
Xenograft Studies. Src inhibitors 17c and 17d were
evaluated in xenograft models employing Src-trans-
formed fibroblasts.15c When these compounds were
administered daily at 50 mg/kg ip from day 0 to day 7,
significant tumor growth inhibition was observed (Fig-
ure 6). The two compounds showed similar efficacy, with
a T/C of 25% for 17c and a T/C of 20% for 17d at day 7.
While significantly reducing tumor growth, some ab-
dominal bloating and moderate weight loss (∼15%) was
observed with this dosing regimen.
Figure 3. Src-transformed rat fibroblasts were treated with
17c for 4 h at the indicated concentrations. (A) Blots of whole
cell lysates were probed with antibodies to phosphotyrosine
(top), pY416 Src (middle), and Src (bottom). The lane labeled
“Rat” has cell lysate from untransformed rat fibroblasts. (B)
Blots of whole cell lysates probed with antibody to pY421
cortactin (top) or cortactin (bottom).
amine groups that were explored, morpholine provided
better cellular potency (16a, 16c, 16h, 17a, 17c, 17f,
17j) than the 4-methylpiperazine or the 4-hydroxypip-
eridine substituted analogues.
Inhibition of Src-Dependent Tyrosine Protein
Phosphorylation. Compound 17c, which possesses the
2,4-dichloro-5-methoxyanilino C-4 substituent of SKI-
606, was selected as a representative analogue for
further evaluation on cellular phosphorylation of Src-
dependent proteins in Src-transformed fibroblasts. Im-
munoblots of whole cell extracts from cells treated with
17c were probed with phosphotyrosine antibodies. As
shown in Figure 3A, 17c reduced tyrosine phosphory-
lation of many cellular proteins with a clear dose
response over a concentration range of 50-500 nM. To
examine the effects of 17c treatment on a specific Src
substrate protein, we examined tyrosine phosphoryla-
tion of cortactin (Figure 3B), whose phosphorylation was
also reduced by treatment with 50-500 nM 17c. In
contrast, the autophosphorylation of Src was diminished
at relatively high concentrations of this compound.
Similar observations were obtained with SKI-606.15c,24
These studies demonstrated that 17c inhibited Src-
dependent tyrosine phosphorylation of cellular proteins
at concentrations that correlated with inhibition of
anchorage-independent growth.
Conclusion
Kinase Selectivity. Analogue 17c was found to be
selective for Src versus several other kinases, including
ErbB-2 (IC50 ) 2.5 µM), EGFr (IC50 ) 2.7 µM), cdk4
On the basis of the activities observed with modified
4-anilinoquinoline-3-carbonitrile cores, the 4-anilinoben-

5914 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19
Berger et al.
Figure 4. Inhibition of HT29 colony formation in soft agar by 17c and 17d. HT29 tumor cells were suspended in a 0.4% agar-
medium mix and plated onto a 0.7% hard agar-medium underlay. After the top layer had solidified, medium containing compound
was added such that the final equilibrium concentration of compound was the indicated value. Fresh medium was added every
week. This photograph was taken 4 weeks after plating.
Figure 5. Phosphorylation of FAK on Y925 in Colo205 cells
is diminished by 17c. Colo205 cells were treated with 17c for
4 h at the indicated concentration. Blots of whole cell lysates
were probed with antibody to pY925 FAK (top) or FAK
(bottom).
Table 4. Nude Mouse Plasma Levels Following a Single Dose
of 50 mg/kg ip
compd 0.5 h (µg/mL) 4 h (µg/mL) 8 h (µg/mL) 24 h (µg/mL)
17c
17d
2.1 ( 0.7
1.3 ( 0.5
1.9 ( 1.3
0.3 ( 0.2
0.02 ( 0.02
0.63 ( 0.06 0.70 ( 0.08 0.24 ( 0.09
Figure 6. Inhibition of Src-transformed fibroblast tumor
growth in nude mice by 17c (9) and 17d (b) administered at
50 mg/kg ip in 2% Tween-80 in 5% dextrose/water once a day
versus vehicle ([) alone, at days 0-7.
zo[g]quinoline-3-carbonitrileswere chosen as the most
promising series to optimize further. We have demon-
strated here that adding the appropriate water-solubi-
lizing substituents to these tricyclic benzo[g]quinolin-
ecarbonitriles provided a series of analogues with
exceptional potency as Src kinase inhibitors. This
represents the only core modification we have explored
that resulted in a series with greater activity than the
similarly substituted 6,7-dialkoxy-4-anilinoquinoline-3-
carbonitriles. A 12-step reaction sequence provided
target compounds 16 and 17 as a separable mixture of
products. Variation at the C-4 and C-7/C-8 positions was
introduced in the final two steps from advanced inter-
mediates 12 and 13.
As anticipated, the SAR of the tricyclic series largely
mirrored that of the 4-anilinoquinoline-3-carbonitrile
compounds. Thus, the C-8 position proved to be optimal
for the attachment of a cyclic basic amine group. As was
observed for the 4-anilinoquinoline-3-carbonitriles, cyclic
amines (17a and 17b) enhanced both enzyme and
cellular activity versus an analogue possessing a meth-
oxy group at C-8 (1). While compounds with several
2,4,5-substituted anilines at the C-4 position provided
similar activity to those possessing the 3,4,5-trimethoxy-
aniline moieties, these were all more active than the
2,4-dichloroaniline substituted 16i and 17k.
Molecular modeling studies indicated that two modes
of protein-ligand interactions, namely, nonbonded and
electrostatic interactions, appear to be the major con-
tributors to differences in the binding of 17d and SKI-
606 with the Src protein pocket. Since our computa-
tional studies indicated that the van der Waals energy
metrics were more discriminating, we attributed the
enhanced potency of 17d primarily to the more optimal
van der Waals protein-ligand interactions. Addition-
ally, in structure-based studies, it appeared that the
more flexible tailpiece at the 7-position of SKI-606 was
often engaged in favorable intramolecular nonbonded

Carbonitriles as Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19 5915
by filtration and washed with acetic acid. The filtrate was
evaporated to approximately 30 mL, then poured into water
and extracted with ether. The organic phase was washed with
aqueous sodium carbonate, water, and brine. After drying over
sodium sulfate, the solution was filtered and evaporated to
give 5.69 g (86%) of 2-[(acetyloxy)methyl]-4-(2-chloroethoxy)-
interactions, while the corresponding tailpiece of 17d
was more significantly involved in favorable nonbonded
interactions with the nearby protein residues. Taking
these factors into account, these studies provide a clear
rationale for the strong binding of analogues such as
17d to Src kinase.
1
5-methoxybenzyl acetate as a white solid, mp 79-80 °C. H
NMR (CDCl3): δ 6.96 (s, 1H), 6.94 (s, 1H), 5.14 (s, 2H), 5.12
(s, 2H), 4.29 (t, J ) 6.2 Hz, 2H), 3.89 (s, 3H), 3.84 (t, J ) 6.2
Hz, 2H), 2.09 (s, 3H), 2.08 (s, 3H). MS (EI), m/z: 329.72 (M+).
Anal. (C15H19ClO6) C, H.
Because 17c and 17d demonstrated potent cell activ-
ity and good blood exposures on ip dosing at 50 mg/kg,
we carried out in vivo studies against Src-transformed
fibroblasts in nude mice. Both analogues showed good
activity in this model when dosed daily at 50 mg/kg (T/C
of 25% and 20% at day 7, respectively). We have
therefore shown that these compounds have sufficient
bioavailability when dosed ip to provide significant in
vivo activity at 50 mg/kg. In addition to having potent
biological activity, these 4-anilinobenzo[g]quinolinecar-
bonitriles have provided important structural informa-
tion with regard to discovering compounds with en-
hanced binding to Src kinase.
A solution of 14.0 g of the 2-[(acetyloxy)methyl]-4-(2-chlo-
roethoxy)-5-methoxybenzyl acetate (42.3 mmol) in 600 mL of
methanol was stirred and cooled in an ice bath while ammonia
gas was bubbled in, until the solution was saturated. The flask
was stoppered and stored in the refrigerator for 15 h. The
reaction mixture was evaporated to give a white solid that was
dried and chromatographed on a silica gel column, eluting with
2:1 hexanes/ethyl acetate, to give 9.87 g (95%) of 6 as a white
solid, mp 93-94 °C. 1H NMR (CDCl3): δ 6.94 (s, 1H), 6.93 (s,
1H), 4.68 (br s, 4H), 4.29 (t, J ) 6.2 Hz, 2H), 3.88 (s, 3H), 3.83
(t, J ) 6.2 Hz, 2H), 2.77 (br s, 1H), 2.71 (br s,1H). MS, m/z:
264.10 (M + NH4)+. Anal. (C11H15ClO4) C, H.
4-(2-Chloroethoxy)-5-methoxyphthalaldehyde (7). To
a 500 mL three-neck round-bottom flask fitted with mechanical
stirrer, thermometer, and addition funnel was added 100 mL
of dry methylene chloride and 8 mL (91.7 mmol) of oxalyl
chloride under nitrogen. This was cooled to -78 °C in a dry
ice/acetone bath. Then 13.6 mL (191.6 mmol) of DMSO in 25
mL of dry methylene chloride was added dropwise. After
complete addition it was further stirred for 5 min. Then 9.87
g (40.0 mmol) of 6 in 10 mL of dry methylene chloride (with
enough DMSO added to dissolve the solid) was added drop-
wise. The reaction mixture was stirred for an additional 30
min, and then 100 mL of triethylamine was added slowly at
-78 °C. After being stirred for 10 min, the solution was allowed
to warm to room temperature, and then 200 mL of ice/water
was added. Following separation of the layers, the aqueous
layer was extracted with methylene chloride (2 × 100 mL).
The organic layers were combined, dried over MgSO4, filtered,
and evaporated to give the crude product as a solid. This solid
was slurried with cold methanol and filtered, washed with cold
methanol, then dried in vacuo to give 6.37 g (66%) of 7 as a
yellow solid, mp 113-114 °C. 1H NMR (CDCl3): δ 7.49 (s, 1H),
7.47 (s, 1H), 4.43 (t, J ) 5.9 Hz, 2H), 4.03 (s, 3H), 3.91 (t, J )
5.9 Hz, 2H). MS, m/z: 242.0 (M + H)+. Anal. (C11H11ClO4) C,
H.
Experimental Section
General Methods. Melting points were determined in open
capillary tubes on a Meltemp melting point apparatus and are
uncorrected. 1H NMR spectra were determined at 400 MHz
on a DRX-400 spectrometer or at 300 MHz on an NT-300 WB
spectrometer, with chemical shifts (δ) reported in parts per
million referenced to Me4Si. Electrospray (ES) mass spectra
were recorded in positive mode on a Micromass Platform or
an LCT spectrometer. Electron impact (EI) mass spectra were
obtained on a Finnigan MAT-90 spectrometer. All reagents
and solvents obtained from commercial suppliers were used
without further purification. Reactions were carried out under
nitrogen as an inert atmosphere. Flash chromatography was
carried out with Baker 40 µM silica gel.
1-(2-Chloroethoxy)-2-methoxybenzene (4). A mixture of
52.88 g (0.426 mol) of guaiacol, 100 g (0.426 mol) of chloroethyl
tosylate, 88.3 g (0.639 mol) of powdered potassium carbonate,
and 600 mL of 2-butanone was stirred mechanically and
heated at reflux for 2 days. The mixture was filtered, and the
solid was rinsed with 2-butanone. After evaporation of the
filtrate, the residue was taken up in ether and washed with 1
N NaOH to remove unreacted guaiacol. The ether layer was
dried over sodium sulfate, filtered, and evaporated to give an
oil that slowly crystallized. Isolation from cold cyclohexane
gave 41.47 g (52%) of 4 as a white solid, mp 42-43 °C. 1H
NMR (CDCl3): δ 6.85-7.02 (m, 4H), 4.28 (t, J ) 6.3 Hz, 2H),
3.87 (s, 3H), 3.84 (t, J ) 6.3 Hz, 2H). MS, m/z: 187.4 (M +
H)+. Anal. (C9H11ClO2) C, H.
1-(2-Chloroethoxy)-4,5-bis(chloromethyl)-2-methoxy-
benzene (5). To a solution of 55.99 g (300 mmol) of 4 in 250
mL of 1,4-dioxane was added 40 mL of concentrated hydro-
chloric acid while stirring at 0 °C. While HCl gas was bubbled
in, 30 mL of 35% formalin was added. After 45 min, another
equal volume of formalin was added. The addition of HCl gas
was continued for 6 h, and the ice bath was removed after 2 h
and allowed to warm to ambient temperature. The reaction
mixture was stirred overnight at ambient temperature. The
green reaction mixture was then cooled in an ice bath, and
the resulting solid was filtered and washed with cold dioxane/
water (2.5:1). Silica gel chromatography of the crude solid,
eluting with 2:1 hexanes/dichloromethane, provided 36.35 g
(42%) of 5 as a white solid, mp 117-118 °C. 1H NMR (CDCl3):
δ 6.92 (s, 1H), 6.91 (s, 1H), 4.70 (s, 2H), 4.69 (s, 2H), 4.29 (t,
J ) 6.2 Hz, 2H), 3.90 (s, 3H), 3.84 (t, J ) 6.2 Hz, 2H). MS,
m/z: 282.0 (M + H)+. Anal. (C11H13Cl3O2) C, H.
Ethyl 3-Amino-7-(2-chloroethoxy)-6-methoxy-2-naph-
thoate (8) and Ethyl 3-Amino-6-(2-chloroethoxy)-7-meth-
oxy-2-naphthoate (9). A mixture of 25 g (0.21 mol) of
3-nitropropionic acid, 300 mL of absolute ethanol, and 10 drops
of concentrated sulfuric acid was refluxed overnight. The
reaction mixture was evaporated, and the residue was parti-
tioned between water and diethyl ether. The ether layer was
washed with water, aqueous sodium bicarbonate solution, and
brine and then dried over sodium sulfate. Following the
removal of ether in vacuo, the product was distilled as a clear
liquid to provide 21.54 g (69%) of ethyl 3-nitropropionate as a
1
clear oil, bp 160-165 °C at 120 mmHg. H NMR (CDCl3): δ
4.66 (t, J ) 6.1 Hz, 2H), 4.18 (q, J ) 7.1 Hz, 2H), 2.98 (t, J )
6.1 Hz, 2H), 1.28 (t, J ) 7.1 Hz, 3H).
To a solution of 2.43 g (16.5 mmol) of ethyl 3-nitropropionate
in 15 mL of absolute ethanol cooled in an ice bath was added
20 mL of 1 N sodium ethoxide in ethanol dropwise over 10
min, keeping the temperature at 0-5 °C. A slurry of 2.43 g
(10.0 mmol) of 7 in 5 mL of ethanol was added. The ice bath
was removed, and the reaction mixture was stirred for 16 h.
The mixture was transferred to a beaker with 300 mL of water
and neutralized with acetic acid to pH 4. A solid was collected
and washed first with water, then with 40 mL of cold ethanol.
The solid was dried in vacuo to provide 2.48 g (70%) of ethyl
7-(2-chloroethoxy)-6-methoxy-3-nitro-2-naphthoate and ethyl
6-(2-chloroethoxy)-7-methoxy-3-nitro-2-naphthoate (∼1:1 mix-
ture) as a yellow solid, mp 119-129 °C dec. 1H NMR (CDCl3):
[4-(2-Chloroethoxy)-2-(hydroxymethyl)-5-methoxyphen-
yl]methanol (6). To a solution of 5.67 g (20 mmol) of 5 in 75
mL of acetic acid was added a solution of 3.5 g of anhydrous
sodium acetate (42.7 mmol) in 100 mL of acetic acid. This
mixture was refluxed with stirring for 2 h. Solids were removed

5916 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19
Berger et al.
δ 8.27 (s, 1H), 8.05 (s, 1H), 7.24, 7.23 and 7.22 (3s, 2H), 4.37-
4.45 (m, 4H), 4.03 (s, 3H), 3.95 (t, J ) 6.0 Hz, 2H), 1.38 (t, J
) 7.1 Hz, 3H). MS, m/z: 354.2 (M + H)+. Anal. (C16H16ClNO6)
C, H, N.
10 mg of pyridine hydrochloride, 150 mg (0.874 mmol) of
4-chloro-5-methoxy-2-methylaniline,15c and 5 mL of 2-ethoxy-
ethanol was stirred and heated to 135 °C. After 1 h, the
mixture was cooled to room temperature, the reaction was
quenched with 0.2 mL of triethylamine, and the mixture was
concentrated in vacuo. The residue was dissolved in 1:1
hexane/ethyl acetate with added dichloromethane and chro-
matographed on silica gel, eluting with 1:1 hexane/ethyl
acetate and then ethyl acetate to provide 0.282 g of 14a and
15a (∼1:1 mixture) as a dull-yellow solid (81%), mp 132-168
A 1.60 g (4.52 mmol) portion of ethyl 7-(2-chloroethoxy)-6-
methoxy-3-nitro-2-naphthoate and ethyl 6-(2-chloroethoxy)-7-
methoxy-3-nitro-2-naphthoate (∼1:1 mixture) was heated in
100 mL of absolute ethanol until it dissolved. The solution was
allowed to cool to room temperature, and 0.2 g of 10%
palladium on carbon was added. Hydrogenation was carried
out in a Parr apparatus at 50 psi for 2 h. The reaction mixture
was filtered through Celite, and the filter cake was rinsed with
ethanol. The filtrate and washes were combined and evapo-
rated to give 8 and 9 (∼1:1 mixture) as a greenish yellow solid,
1
°C dec. H NMR (CDCl3): δ 8.66 and 8.65 (2s, 1H), 8.41 and
8.40 (2s, 1H), 8.19 and 8.17 (2s, 1H), 7.36 (s, 1H), 7.25 and
7.24 (2s, 1H), 7.12 (s, 1H), 7.02 (br s, 1H), 6.76 (s, 1H), 4.48
and 4.41 (2t, J ) 6.1 Hz, 2H), 4.07 and 4.01 (2s, 3H), 3.95 and
3.98 (2t, J ) 6 0.1 Hz, 2H), 3.77 (s, 3H), 2.28 (s, 3H). MS, m/z:
482.0 (M + H)+. Anal. (C25H21Cl2N3O3‚H2O) C, H, N.
1
1.28 g (87%), mp 104-108 °C. H NMR (CDCl3): δ 8.34 and
8.32 (2s, 1H), 7.06 and 7.03 (2s, 1H), 6.85 and 6.84 (2s, 1H),
6.82 (s, 1H), 5.52 (br s, 2H), 4.30-4.43 (m, 4H), 3.97 and 3.93
(2s, 3H), 3.89 (t, J ) 6.6 Hz, 2H), 1.44 (t, J ) 7.1 Hz, 3H). MS,
m/z: 324.3 (M + H)+. Anal. (C16H18ClNO4) C, H, N.
4-(2,4-Dichloro-5-methoxyanilino)-8-methoxy-7-(chlo-
roethoxy)benzo[g]quinoline-3-carbonitrile (14b) and
4-(2,4-Dichloro-5-methoxyanilino)-7-methoxy-8-(chloro-
ethoxy)benzo[g]quinoline-3-carbonitrile (15b). A mixture
of 1.10 g (3.17 mmol) of 12 and 13 (∼1:1 mixture), 50 mg of
pyridine hydrochloride, 742 mg (3.86 mmol) of 2,4-dichloro-5-
methoxyaniline,15c and 25 mL of 2-ethoxyethanol was stirred
and heated to 135 °C. After 1 h, the mixture was cooled to
room temperature, the reaction was quenched with 1.0 mL of
triethylamine, and the mixture was concentrated in vacuo. The
residue was dissolved in 95:5 methylene chloride/methanol and
chromatographed on silica gel, eluting with 1:1 hexane/ethyl
acetate. The product was precipitated from ethyl acetate to
provide 0.760 g (48%) of 14b and 15b (∼1:1 mixture) as a dull-
7-(2-Chloroethoxy)-8-methoxy-4-oxo-1,4-dihydrobenzo-
[g]quinoline-3-carbonitrile (10) and 8-(2-Chloroethoxy)-
7-methoxy-4-oxo-1,4-dihydrobenzo[g]quinoline-3-carbo-
nitrile (11). A 648 mg portion (2.0 mmol) of ethyl 3-amino-
7-(2-chloroethoxy)-6-methoxy-2-naphthoate and ethyl 3-amino-
6-(2-chloroethoxy)-7-methoxy-2-naphthoate (∼1:1 mixture) and
5 mL of dimethylformamide dimethylacetal were heated to
reflux using an oil bath. The mixture was kept at reflux
overnight. Solvent was removed in vacuo to provide crude ethyl
6-(2-chloroethoxy)-3-{[(E)-(dimethylamino)methylidene]amino}-
7-methoxy-2-naphthoate and ethyl 7-(2-chloroethoxy)-3-{[(E)-
(dimethylamino)methylidene]amino}-6-methoxy-2-naphtho-
ate (∼1:1 mixture) as a dark-red mixture that was used in the
next step without further purification.
To 2.5 mL of dry tetrahydrofuran at -78 °C was added 1.8
mL of 2.5 M n-butyllithium (4.4 mmol). Then 0.24 mL (4.6
mmol) of dry acetonitrile in 4.5 mL of dry tetrahydrofuran was
added dropwise over 10 min. This was stirred an additional
15 min at -78 °C. Then the ethyl 6-(2-chloroethoxy)-3-{[(E)-
(dimethylamino)methylidene]amino}-7-methoxy-2-naphtho-
ate and ethyl 7-(2-chloroethoxy)-3-{[(E)-(dimethylamino)-
methylidene]amino}-6-methoxy-2-naphthoate (∼1:1 mixture)
was dissolved in 3 mL of tetrahydrofuran and added dropwise
over 15 min. The reaction mixture was stirred for 30 min at
-78 °C. Then the reaction was quenched with 0.57 mL (10
mmol) of glacial acetic acid, and the mixture was warmed to
room temperature. A 10 mL portion of water was added to
the yellow mixture. The solids were filtered, washed with
water, and dried to give 0.502 g of 10 and 11 (∼1:1 mixture)
as a yellow-green solid (76%) that was used in the next step
without further purification, mp 260-73 °C dec. 1H NMR
(DMSO-d6): δ 8.68 (s, 1H), 8.62 and 8.61 (2s, 1H), 7.95 and
7.94 (2s, 1H), 7.62 and 7.61 (2s, 1H), 7.49 and 7.47 (2s, 1H),
4.37-4.47 (m, 2H), 4.00-4.11 (m, 2H), 3.96 and 3.93 (2s, 3H).
MS, m/z: 329.5 (M + H)+.
1
yellow solid, mp 239-255 °C dec. H NMR (CDCl3 + DMSO-
d6): δ 8.94 (br s, 1H), 8.77 (br s, 1H), 8.59 (br s, 1H), 8.36 (br
s, 1H), 7.54 (br s, 1H), 7.28 (br s, 1H), 7.26 (br s, 1H), 6.96 (br
s, 1H), 4.40-4.49 (m, 2H), 3.94-4.07 (m, 5H), 3.86 (s, 3H).
MS, m/z: 502.2 (M + H)+. Anal. (C24H18Cl3N3O3‚0.3H2O) C,
H, N.
8-(2-Chloroethoxy)-4-(2-chloro-5-methoxy-4-methyl-
anilino)-7-methoxybenzo[g]quinoline-3-carbonitrile (14c)
and 7-(2-Chloroethoxy)-4-(2-chloro-5-methoxy-4-methyl-
anilino)-8-methoxybenzo[g]quinoline-3-carbonitrile (15c).
Following the route used to prepare 14a and 15a, 14c and
15c (∼1:1 mixture) were obtained as a yellow solid in 64% yield
from the reaction of 12 and 13 with 2-chloro-5-methoxy-4-
methylaniline,15c mp 204-212 °C. 1H NMR (DMSO-d6):
δ
10.05 (s, 2H), 9.04 (s, 2H), 8.44 (s, 2H), 6.35 (s, 2H), 7.55 (s,
2H), 7.36 (s, 4H), 7.14 (s, 2H), 4.44 (s, 4H), 4.07 (s, 4H), 3.98
(s, 6H), 3.78 (s, 6H), 2.19 (s, 6H). MS, m/z: 482.0 (M + H)+.
Anal. (C25H21Cl2lN3O3) C, H, N.
8-(2-Chloroethoxy)-4-(2-chloro-4-fluoro-5-methoxy-
anilino)-7-methoxybenzo[g]quinoline-3-carbonitrile (14d)
and 7-(2-Chloroethoxy)-4-(2-chloro-4-fluoro-5-methoxy-
anilino)-8-methoxybenzo[g]quinoline-3-carbonitrile (15d).
Following the route used to prepare 14a and 15a, 14d and
15d (∼1:1 mixture) were obtained as a yellow solid in 59%
yield from the reaction of 12 and 13 with 2-chloro-4-fluoro-5-
4-Chloro-8-methoxy-7-(2-chloroethoxy)benzo[g]quino-
line-3-carbonitrile (12) and 4-Chloro-7-methoxy-8-(2-
chloroethoxy)benzo[g]quinoline-3-carbonitrile (13). To
a slurry of 1.11 g (3.38 mmol) of 10 and 11 (∼1:1 mixture)
and 5 mL of phosphorus oxychloride was added 0.15 mL of
anhydrous dimethylformamide. This was stirred and heated
to reflux for 20 min using an oil bath, followed by concentration
in vacuo. The dark residue was quenched with 30 mL of cold
water. The solid formed was collected, washed with water, and
dried to give 1.02 g (87%) of 12 and 13 (∼1:1 mixture) as a
1
methoxyaniline,15c mp 193-204 °C. H NMR (CDCl3): δ 8.70
(s, 1H), 8.44 and 8.43 (2s, 1H), 8.28 and 8.27 (2s, 1H), 7.31
and 7.28 (2s, 1H), 7.20 (s, 1H), 7.11 (s, 1H), 6.89 and 6.87 (2s,
1H), 4.49 and 4.42 (2t, J ) 6.0 Hz, 2H), 4.08 and 4.03 (2s, 3H),
3.98 and 3.96 (2t, J ) 6.0 Hz, 2H), 3.79 (s, 3H). MS, m/z: 486.0,
488.1 (M + H)+. Anal. (C24H18Cl2FN3O3‚0.3H2O) C, H, N.
7-(2-Chloroethoxy)-8-methoxy-4-(3,4,5-trimethoxy-
anilino)benzo[g]quinoline-3-carbonitrile (14e) and 8-(2-
Chloroethoxy)-7-methoxy-4-(3,4,5-trimethoxyanilino)-
benzo[g]quinoline-3-carbonitrile (15e). A mixture of 500
mg of 12 and 13 (1.4 mmol), 277 mg of 3,4,5-trimethoxyaniline15a
(1.5 mmol), and 200 mg of pyridine hydrochloride (1.7 mmol)
in 14 mL of 2-ethoxyethanol was heated at reflux for 0.5 h.
When the mixture cooled, a solid precipitated from solution,
which was collected by filtration. Subsequent washing with
water and hexanes provided 700 mg (99% yield) of a yellow
1
greenish yellow solid, mp 195-209 °C dec. H NMR (CDCl3):
δ 8.88 (s, 1H), 8.66 and 8.65 (2s, 1H), 8.52 and 8.51 (2s, 1H),
7.33 (s, 1H), 7.30 and 7.29 (2s, 1H), 4.46-4.52 (m, 2H), 4.09
and 4.08 (2s, 3H), 3.96-4.00 (m, 2H). MS, m/z: 347.3 (M +
H)+. Anal. (C17H12Cl2N2O2) C, H, N.
4-(4-Chloro-5-methoxy-2-methylanilino)-8-methoxy-7-
(chloroethoxy)benzo[g]quinoline-3-carbonitrile (14a) and
4-(4-Chloro-5-methoxy-2-methylanilino)-7-methoxy-8-
(chloroethoxy)benzo[g]quinoline-3-carbonitrile (15a). A
mixture of 248 mg (0.714 mmol) of 12 and 13 (∼1:1 mixture),

Carbonitriles as Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19 5917
solid mixture of 14e and 15e (∼1:1 mixture), which was used
4-(2,4-Dichloro-5-methoxyanilino)-8-methoxy-7-[2-(4-
morpholinyl)ethoxy]benzo[g]quinoline-3-carbonitrile
(16c) and 4-(2,4-Dichloro-5-methoxyanilino)-7-methoxy-
8-[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-3-carbo-
nitrile (17c). A mixture of 0.436 g (0.87 mmol) of 14b and
15b (∼1:1 mixture), 2.0 mL (23.0 mmol) of morpholine, and
0.05 g of sodium iodide (0.34 mmol) in 2.0 mL of ethylene glycol
dimethyl ether was heated at 90 °C for 3.5 h under nitrogen.
The mixture was cooled, solvent was removed in vacuo, and
the resulting residue was stirred with saturated aqueous
sodium bicarbonate. The crude solid was collected by filtration,
washed with water, and dried in vacuo. Purification was
carried out by silica gel chromatography, eluting with a
gradient of 97:3 to 90:10 ethyl acetate/methanol to yield 0.241
g (50%) of 16c as a yellow solid, mp 210-212 °C, and 0.203 g
(42%) of 17c as a yellow solid, mp 207-214 °C.
in the next step without further purification.
7-(2-Chloroethoxy)-4-(2,4-dichloroanilino)-8-methoxy-
benzo[g]quinoline-3-carbonitrile (14f) and 8-(2-Chloro-
ethoxy)-4-(2,4-dichloroanilino)-7-methoxybenzo[g]quin-
oline-3-carbonitrile (15f). A mixture of 300 mg of 12 and
13 (0.87 mmol), 153 mg of 2,4-dichloroaniline (0.94 mmol), and
100 mg of pyridine hydrochloride (0.87 mmol) in 10 mL of
2-ethoxyethanol was heated at reflux for 1 h. The reaction
mixture was cooled to room temperature and added to satu-
rated aqueous sodium bicarbonate. The solid was collected by
filtration and partitioned between ethyl acetate and 1 N
aqueous sodium hydroxide. The organic layer was washed with
1 N aqueous sodium hydroxide, dried over magnesium sulfate,
filtered, and concentrated in vacuo. The crude solid was passed
through a silica gel column, eluting with 99:1 dichloromethane/
methanol to provide 194 mg of 14f and 15f (∼1:1 mixture),
which was directly used in the next step.
16c. 1H NMR (DMSO-d6 + TFA): δ 9.35 (s, 1H), 9.25 (s 1H),
8.43 (s, 1H), 7.91 (s, 1H), 7.78 (s 1H), 7.63 (s, 1H), 7.53 (s, 1H),
4.65 (m, 2H), 4.06 (s, 3H), 4.04-3.97 (m, 2H), 3.91 (s, 3H),
3.84-3.63 (m, 6H), 3.34 (t, J ) 10.6 Hz, 2H). MS, m/z: 553.3
(M + H)+. Anal. (C28H26Cl2N4O4‚0.15CH3CO2C2H5) C, H, N.
17c. 1H NMR (DMSO-d6 + TFA): δ 9.35 (s, 1H), 9.24 (s 1H),
8.42 (s, 1H), 7.91 (s, 1H), 7.82 (s 1H), 7.61 (s, 1H), 7.48 (s, 1H),
4.66 (m, 2H), 4.07 (s, 3H), 4.04-3.97 (m, 2H), 3.90 (s, 3H),
3.83-3.63 (m, 6H), 3.34 (m, 2H). MS, m/z: 553.3 (M + H)+.
Anal. (C28H26Cl2N4O4‚2.0H2O) C, H, N.
4-(4-Chloro-5-methoxy-2-methylanilino)-8-methoxy-7-
[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-3-carboni-
trile (16a) and 4-(4-Chloro-5-methoxy-2-methylanilino)-
7-methoxy-8-[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-
3-carbonitrile (17a). A mixture of 318 mg (0.66 mmol) of 14a
and 15a (∼1:1 mixture), 100 mg of sodium acetate and 5 mL
of morpholine was stirred and heated to 130 °C using an oil
bath. After 30 min the mixture was allowed to cool to room
temperature. The reaction mixture was concentrated in vacuo
and the residue was purified by silica gel chromatography,
eluting with 95:5 methylene chloride/methanol to yield 153 mg
(43%) of 16a as a yellow solid, mp 191-194 °C dec, and 82 mg
(23%) of 17a as a yellow wax.
4-(2,4-Dichloro-5-methoxyanilino)-8-methoxy-7-[2-(4-
methyl-1-piperazinyl)ethoxy]benzo[g]quinoline-3-carbo-
nitrile (16d) and 4-(2,4-Dichloro-5-methoxyanilino)-7-
methoxy-8-[2-(4-methyl-1-piperazinyl)ethoxy]benzo-
[g]quinoline-3-carbonitrile (17d). Following the route used
to prepare 16c and 17c, the reaction of 14b and 15b with
1-methylpiperazine provided 16d as a yellow solid in 33% yield
(after purification by silica gel chromatography, eluting with
dichloromethane/methanol), mp 141-150 °C, and 17d as a
yellow solid in 45% yield, mp 132-135 °C.
1
16a. H NMR (CDCl3): δ 8.65 (s, 1H), 8.39 (s, 1H), 8.17 (s,
1H), 7.35 (br s, 1H), 7.27 (s, 1H), 7.22 (s, 1H), 7.10 (s, 1H),
6.75 (s, 1H), 4.28 (t, J ) 5.7 Hz, 2H), 4.05 (s, 3H), 3.75 (s, 3H),
3.78-3.73 (m, 4H), 2.94 (t, J ) 5.7 Hz, 2H), 2.67-2.62 (br s,
4H), 2.27 (s, 3H). MS, m/z: 533.1 (M + H)+. Anal. (C29H29-
ClN4O4‚0.5H2O) C, H, N.
16d. 1H NMR (DMSO-d6 + TFA): δ 9.34 (s, 1H), 9.23 (s
1H), 8.42 (s, 1H), 7.91 (s, 1H), 7.77 (s 1H), 7.61 (s, 1H), 7.51
(s, 1H), 4.63 (m, 2H), 4.03 (s, 3H), 3.90 (s, 3H), 3.81-3.31 (m,
10H), 2.94 (s, 3H). MS, m/z: 566.3 (M + H)+. Anal. (C29H29-
Cl2N5O3‚0.9CH2Cl2) C, H, N.
1
17a. HNMR(CDCl3): δ 8.57 (s, 1H), 8.40 (s, 1H), 8.30 (s,
1H), 7.76 (br s, 1H), 7.25 (s, 1H), 7.18 (s, 1H), 7.08 (s, 1H),
6.80 (s, 1H), 4.32 (t, J ) 5.7 Hz, 2H), 3.92 (s, 3H), 3.75 (s, 3H),
3.77-3.74 (m, 4H), 2.95 (t, J ) 5.7 Hz, 2H), 2.67-2.60 (br s,
4H), 2.19 (s, 3H). MS 533.1 (M + H)+. Anal. (C29H29ClN4O4‚
H2O) C, H, N.
17d. 1H NMR (DMSO-d6 + TFA): δ 9.35 (s, 1H), 9.23 (s
1H), 8.42 (s, 1H), 7.91 (s, 1H), 7.80 (s 1H), 7.60 (s, 1H), 7.48
(s, 1H), 4.64 (m, 2H), 4.03 (s, 3H), 3.90 (s, 3H), 3.81-3.4 (m,
10H), 2.94 (s, 3H). MS, m/z: 566.3 (M + H)+ Anal. (C29H29-
Cl2N5O3‚0.5CH2Cl2) C, H, N.
4-(4-Chloro-5-methoxy-2-methylanilino)-7-methoxy-
8-[2-(4-methyl-1-piperazinyl)ethoxy]benzo[g]quinoline-
3-carbonitrile (16b) and 4-(4-Chloro-5-methoxy-2-methyl-
anilino)-8-methoxy-7-[2-(4-methyl-1-piperazinyl)ethoxy]-
benzo[g]quinoline-3-carbonitrile (17b). A mixture of 205
mg (0.425 mmol) of 14a and 14b (∼1:1 mixture), 0.15 mL (1.35
mmol) of 1-methylpiperazine, and 50 mg (0.34 mmol) of sodium
iodide in 5 mL of ethylene glycol dimethyl ether was heated
at 90 °C for 4 days under nitrogen. The mixture was cooled,
the solvent was removed in vacuo, and the resulting residue
was stirred with saturated aqueous sodium bicarbonate. The
crude solid was collected by filtration, washed with water, and
dried in vacuo. Purification was carried out by silica gel
chromatography, eluting with a gradient of 92:8 to 85:15
methylene chloride/methanol to yield 100 mg (43%) of 16b as
a yellow solid, mp 121-135 °C, and 68 mg (29%) of 17b as a
yellow solid, mp 122-137 °C.
4-(2,4-Dichloro-5-methoxyphenylamino)-7-[2-(4-hydroxy-
piperidin-1-yl)ethoxy]-8-methoxybenzo[g]quinoline-3-
carbonitrile (16e) and 4-(2,4-Dichloro-5-methoxyphenyl-
amino)-8-[2-(4-hydroxypiperidin-1-yl)ethoxy]-7-methoxy-
benzo[g]quinoline-3-carbonitrile (17e). Following the route
used to prepare 16b and 17b, the reaction of 14b and 15b
with 4-hydroxypiperidine provided 16e as a yellow solid in 7%
yield, mp 150-160 °C, and 17e as a yellow solid in 25% yield,
mp 193-198 °C dec.
16e. 1H NMR(DMSO-d6): δ 9.37 (s, 1 H), 9.28 (s, 1H), 8.46
(s, 1H), 7.89 (s, 1H), 7.78 (s, 1H), 7.64 (s, 1H), 7.53 (d, J )
3.90 Hz, 1H), 4.68-4.60 (m, 2H), 4.08 (s, 3H), 4.02-3.97 (m,
1H), 3.92 (s, 3H), 3.77-3.67 (m, 3H), 3.57-3.38 (m, 2H), 3.27-
3.17 (m, 1H), 2.10-1.79 (m, 3H), 1.74-1.63 (m, 1H). MS, m/z:
566.7 (M + H)+. Anal. (C29H28Cl2N4O4‚2.5H2O) C, H, N.
17e. 1H NMR (400 MHz, DMSO-d6): δ 9.39 (s, 1H), 9.29 (s,
1H), 8.46 (s, 1H), 7.89 (s, 1H), 7.83 (d, J ) 3.8 Hz, 1H), 7.64
(s, 1H), 7.49 (s, 1H), 4.70-4.62 (m, 2H), 4.06 (s, 3H), 4.03-
3.96 (m, 1H), 3.93 (s, 3H), 3.78-3.67 (m, 3H), 3.57-3.49 (m,
1H), 3.49-3.39 (m, 1H), 3.29-3.19 (m, 1H), 2.11-2.03 (m, 1H),
2.03-1.92 (m, 1H), 1.88-1.82 (m, 1H), 1.75-1.64 (m, 1H). MS,
m/z: 566.8 (M + H)+. Anal. (C29H28Cl2N4O4‚1.1CH3OH) C, H,
N.
1
16b. H NMR (DMSO-d6): δ 9.86 (s, 1H), 9.01 (s 1H), 8.42
(s, 1H), 8.32 (s, 1H), 7.57 (s, 1H), 7.43 (s 1H), 7.31 (s, 1H),
7.16 (s, 1H), 4.27 (t, J ) 5.6 Hz, 2H), 3.96 (s, 3H), 3.81 (s, 3H),
2.81 (t, J ) 5.8 Hz, 2H), 2.54 (m, 4H), 2.37 (br s, 4H), 2.18 (s,
3H), 2.14 (s, 3H). MS, m/z: 546.4 (M + H)+. Anal. (C30H32-
ClN5O3‚1.6CH2Cl2) C, H, N.
17b. 1HNMR (DMSO-d6): δ 9.86 (s, 1H), 9.0 (s 1H), 8.43 (s,
1H), 8.34 (s, 1H), 7.53 (s, 1H), 7.43 (s 1H), 7.33 (s, 1H), 7.16
(s, 1H), 4.27 (t, J ) 5.6 Hz, 2H), 3.97 (s, 3H), 3.81 (s, 3H), 2.81
(t, J ) 5.8 Hz, 2H), 2.55 (m, 4H), 2.36 (br s, 4H), 2.17 (s, 3H),
2.14 (s, 3H). MS, m/z: 546.4 (M + H)+. Anal. (C30H32-
ClN5O3‚1.0CH2Cl2) C, H, N.
4-(2-Chloro-5-methoxy-4-methylanilino)-8-methoxy-7-
[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-3-carboni-
trile (16f) and 4-(4-Chloro-5-methoxy-2-methylanilino)-
7-methoxy-8-[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-
3-carbonitrile (17f). Following the route used to prepare 16c

5918 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19
Berger et al.
and 17c, the reaction of 14c and 15c with morpholine provided
16d as a yellow solid in 48% yield, mp 240-241 °C, and 17d
as a yellow solid in 47% yield, mp 220-222 °C.
16h. 1H NMR (DMSO-d6): δ 9.92 (s, 1H), 8.91 (s, 1H), 8.47
(s, 1H), 8.32 (s, 1H), 7.52 (s, 1H), 7.35 (s, 1H), 6.67 (s, 2H),
4.28 (t, J ) 4.3 Hz, 2H), 3.97 (s, 3H), 3.78 (s, 6H), 3.69 (s, 3H),
3.61 (t, J ) 3.5 Hz, 4H), 2.81 (t, J ) 4.5 Hz, 2H), 2.54 (t, J )
3.5 Hz, 4H). MS, m/z: 544.9 (M + H)+. Anal. (C31H35N5O5‚
1.0CH2Cl2) C, H, N.
1
16f. H NMR (DMSO-d6 + TFA-d): δ 9.36 (s, 1H), 9.24 (s
1H), 8.43 (s, 1H), 7.81 (s, 1H), 7.51 (s, 2H), 7.35 (s 1H), 4.65
(m, 2H), 4.07 (s, 3H), 4.03 (s, 1H), 3.83 (s, 3H), 3.76-3.64 (m,
7H), 3.35 (m, 2H), 2.27 (s, 3H). MS, m/z: 533.0 (M + H)+. Anal.
(C29H29ClN4O4) C, H, N.
1
17j. H NMR (DMSO-d6): δ 9.92 (s, 1H), 8.94 (s, 1H), 8.47
(s, 1H), 8.30 (s, 1H), 7.56 (s, 1H), 7.31 (s, 1H), 6.67 (s, 2H),
4.29 (t, J ) 4.4 Hz, 2H), 3.95 (s, 3H), 3.78 (s, 6H), 3.69 (s, 3H),
3.61 (t, J ) 3.4 Hz, 4H), 2.82 (t, J ) 4.4 Hz, 2H), 2.53 (t, J )
3.5 Hz, 4H). MS, m/z: 544.9 (M + H)+. Anal. (C31H35N5O5‚
1.5H2O) C, H, N.
1
17f. H NMR (DMSO-d6 + TFA-d): δ 9.38 (s, 1H), 9.25 (s
1H), 8.42 (s, 1H), 7.83 (s, 1H), 7.51 (s, 1H), 7.46 (s, 1H), 7.35
(s 1H), 4.67 (m, 2H), 4.07 (s, 1H), 4.05 (s, 3H), 3.84 (s, 3H),
3.79-3.65 (m, 7H), 3.35 (m, 2H), 2.27 (s, 3H). MS, m/z: 533.0
(M + H)+. Anal. (C29H29ClN4O4‚2.2H2O) C, H, N.
4-(2,4-Dichloroanilino)-8-methoxy-7-[2-(4-morpholinyl)-
ethoxy]benzo[g]quinoline-3-carbonitrile (16i) and 4-(2,4-
Dichloroanilino)-7-methoxy-8-[2-(4-morpholinyl)ethoxy]-
benzo[g]quinoline-3-carbonitrile (17k). A 194 mg (0.41
mmol) portion of 14f and 15f was combined with 1.0 mL of
morpholine (11.5 mmol) and 10 mg of sodium iodide (0.07
mmol) in 3 mL of ethylene glycol dimethyl ether. The suspen-
sion was heated at reflux for 8 h, then cooled to room
temperature and poured into saturated aqueous sodium
bicarbonate. The solid was collected by filtration, washing with
water. Flash column chromatography, eluting with a gradient
of 95:5 to 4:1 ethyl acetate/methanol, gave 62 mg (29%) of 16i
as a yellow solid, mp 115-119 °C dec, and 52 mg (24%) of 17k
as a yellow solid, mp 250-252 °C dec.
4-(2-Chloro-5-methoxy-4-methylanilino)-8-[2-(4-hydroxy-
piperidin-1-yl)ethoxy]-7-methoxybenzo[g]quinoline-3-car-
bonitrile (17g). Following the route used to prepare 16b and
17b, the reaction of 14c and 15c with 4-hydroxypiperidine
provided 17g as a yellow solid in 51% yield, mp 210-215 °C.
1H NMR (DMSO-d6): δ 9.38 (s, 1H), 9.24 (s, 1H), 8.42 (s, 1H),
7.82 (d, J ) 3.5 Hz, 1H), 7.51 (s, 1H), 7.47 (s, 1H), 7.34 (s,
1H), 4.64 (q, J ) 5.5 Hz, 2H), 4.05 (s, 3H), 3.99-3.95 (m, 1H),
3.84 (s, 3H), 3.76-3.65 (m, 3H), 3.55-3.48 (m, 1H), 3.46-3.36
(m, 1H), 3.27-3.16 (m, 1H), 2.27 (s, 3H), 2.10-1.79 (m, 3H),
1.72-1.61 (m, 1H). MS, m/z: 546.8 (M + H)+. Anal. (C30H31-
ClN4O4‚1.25H2O‚0.25HCl) C, H, N.
4-(2-Chloro-4-fluoro-5-methoxyanilino)-7-methoxy-8-
[2-(4-morpholinyl)ethoxy]benzo[g]quinoline-3-carboni-
trile Hydrochloride (17h). A mixture of 243 mg (0.50 mmol)
of 14d and 15d (∼1:1 mixture) and 100 mg (0.67 mmol) of
sodium iodide in 5 mL of morpholine was heated to reflux for
30 min. After cooling to room temperature, the product mixture
was evaporated, then chromatographed on silica gel, eluting
with 9:1 ethyl acetate/methanol. This gave 76 mg (26%) of 17h
as a yellow solid, mp 206-222 °C dec. 1H NMR (DMSO-d6): δ
9.36 (s, 1H), 9.26 (s, 1H), 8.43 (s, 1H), 7.84 (s, 1H), 7.78 (d, J
) 10.9 Hz, 1H), 7.68 (d, J ) 8.3 Hz, 1H), 7.47 (s, 1H), 4.66 (m,
2H), 4.10-3.98 (m, 6H), 3.89 (s, 3H), 3.81-3.73 (m, 4H), 3.73-
3.61 (m, 3H), 3.24-3.44 (s, 2H). MS, m/z: 537.1 (M + H)+.
Anal. (C28H26ClFN4O4‚1.5H2O‚1.2HCl) C, H, N.
8-Methoxy-7-[2-(4-methyl-1-piperazinyl)ethoxy]-4-(3,4,5-
trimethoxyanilino)benzo[g]quinoline-3-carbonitrile (16
g) and 7-Methoxy-8-[2-(4-methyl-1-piperazinyl)ethoxy]-
4-(3,4,5-trimethoxyanilino)benzo[g]quinoline-3-carboni-
trile (17i). A mixture of 350 mg (0.70 mmol) of 14e and 15e
(∼1:1 mixture) was combined with 1.4 g (14 mmol) of N-
methylpiperazine and 10 mg (0.07 mmol) of sodium iodide in
5 mL of ethylene glycol dimethyl ether. The suspension was
heated at reflux for 26 h, then cooled to room temperature and
poured into saturated aqueous sodium bicarbonate. The solid
was collected by filtration, washing with water. Flash column
chromatography, eluting with a gradient of 95:5 to 4:1 meth-
ylene chloride/methanol, followed by washing with ethyl
acetate and hexane, gave 142 mg (36%) of 16g as a light-yellow
solid, mp 115-120 °C, and 127 mg (32% yield) of 17i as a light-
yellow solid, mp 95-97 °C.
16i. 1H NMR (DMSO-d6, TFA): δ 9.29 (s, 1H), 9.20 (s, 1H),
8.40 (s, 1H), 7.94 (d, J ) 2.8 Hz, 1H), 7.76 (s, 1H), 7.74 (d, J
) 10.4 Hz, 1H), 7.65 (dd, J ) 8.4, 2.0 Hz, 1H), 4.62 (m, 2H),
7.52 (s, 1H), 4.09-3.98 (m, 5H), 3.79-3.57 (m, 6H), 3.38-3.23
(m, 2H). MS, m/z: 522.7 (M + H)+. Anal. (C27H24Cl2N4O3‚
1.0H2O) C, H, N.
17k. 1H NMR (DMSO-d6, TFA): δ 9.30 (s, 1H), 9.22 (s, 1H),
8.40 (s, 1H), 7.94 (d, J ) 2.4 Hz, 1H), 7.81 (s, 1H), 7.75 (d, J
) 8.4 Hz, 1H), 7.65 (dd, J ) 8.4, 2.4 Hz, 1H), 7.47 (s, 1H), 4.64
(m, 2H), 4.08-3.97 (m, 5H), 3.79-3.58 (m, 6H), 3.37-3.24 (m,
2H). MS, m/z: 522.7 (M + H)+. Anal. (C27H24Cl2N4O3‚1.0H2O)
C, H, N.
Molecular Modeling. The structures of 17d and SKI-606
were minimized using the MMFF force field in Sybyl, a
software package of Tripos (Sybyl 6.9; Tripos, Inc., St. Louis,
MO), and conformationally expanded using the Openeye
Scientific Software package, Omega (Open Eye Scientific
Software, Santa Fe, NM) with a 1.0 Å rmsd cutoff to filter out
degenerate conformers. A homology model of the active form
of the Src kinase domain was constructed using the active form
of the Lck kinase domain as a template. The software package
of Andrej Sali, MODELLER (Accelrys, Inc., San Diego, CA),
was used to build the homology model. The Openeye Scientific
Software package FRED (Open Eye Scientific Software, Santa
Fe, NM) was used for the docking studies. In summary, FRED
does not use stochastic sampling to dock the ligand conformers;
rather, a shape-based matching of all possible orientations of
each conformer is optimized in accordance with the selected
scoring function. The PLP or piecewise linear potential func-
tion was used to score the poses docked using FRED, consisting
of a steric matching function with a less stringent charge
matching function of protein-ligand interactions. The top 10
poses obtained from the FRED docking procedure are rescored
with a Poisson-Boltzmann molecular mechanics scoring func-
tion (PBMM-SA).26 The free energy of protein-ligand binding
is desribed as
16g. 1H NMR (DMSO-d6): δ 9.91 (s, 1H), 8.93 (s, 1H), 8.49
(s, 1H), 8.34 (s, 1H), 7.53 (s, 1H), 7.34 (s, 1H), 6.70 (s, 2H),
4.25 (t, J ) 5.5 Hz, 2H), 3.97 (s, 3H), 3.78 (s, 6H), 3.69 (s, 3H),
2.81 (t, J ) 5.5 Hz, 2H), 2.55-2.38 (m, 8H), 2.18 (s, 3H). MS,
m/z: 558.3 (M + H)+. Anal. (C31H35N5O5‚1.05EtOAc) C, H, N.
1
17i. H NMR (DMSO-d6): δ 9.92 (s, 1H), 8.95 (s, 1H), 8.49
(s, 1H), 8.32 (s, 1H), 7.57 (s, 1H), 7.31 (s, 1H), 6.70 (s, 2H),
4.28 (t, J ) 5.9 Hz, 2H), 3.96 (s, 3H), 3.79 (s, 6H), 3.69 (s, 3H),
2.81 (t, J ) 5.5 Hz, 2H), 2.55-2.38 (m, 8H), 2.17 (s, 3H). MS,
m/z: 558.2 (M + H)+. Anal. (C31H35N5O5‚5.0H2O) C, H, N.
8-Methoxy-7-[2-(4-morpholinyl)ethoxy]-4-[(3,4,5-tri-
methoxyphenyl)amino]benzo[g]quinoline-3-carbonitrile
(16h) and 7-Methoxy-8-[2-(4-morpholinyl)ethoxy]-4-(3,4,5-
trimethoxyanilino)benzo[g]quinoline-3-carbonitrile (17j).
Following the route used to prepare 16g and 17i, the reaction
of 14e and 15e with morpholine provided 16h as a yellow solid
in 37% yield, mp 135-138 °C, and 17j as a yellow solid in 37%
yield, mp 127-130 °C.
solv
gas
∆G
) ∆G
- ∆GPsolv - ∆GLsolv + ∆GPL
bind
bind solv
where the first term at the right of the equal sign is the gas-
phase protein-ligand binding energy and is obtained via
molecular mechanics calculations. The remainder of the terms
represent the solvated free energy terms for the protein and
ligand individually as well as the protein-ligand complex. The
solvated free energy terms are evaluated explicitly via Pois-
son-Boltzmann/surface area calculations. Solvation and hy-
drophobic effects are handled by this scoring function, which
is used to rescore or fine-tune the docked views.

Carbonitriles as Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 19 5919
hibitors of the tyrosine kinase c-Src. Bioorg. Med. Chem. Lett.
2000, 10, 945-949. (c) Widler, L.; Green, J.; Missbach, M.; Susa,
M.; Altmann, E. 7-Alkyl- and 7-cycloalkyl-5-arylpyrrolo[2,3-d]-
pyrimidines, potent inhibitors of the tyrosine kinase c-src.
Bioorg. Med. Chem. Lett. 2001, 11, 849-852. (d) Altmann, E.;
Missbach, M.; Green, J.; Susa, M.; Wagenknecht, H.-A.; Widler,
L. 7-Pyrrolidinyl- and 7-piperidinyl-5-arylpyrrolo[2,3-d]pyrim-
idines, potent inhibitors of the tyrosine kinase c-src. Bioorg. Med.
Chem. Lett. 2001, 11, 853-856. (e) Sundaramoorthi, R.; Shakes-
peare, W. C.; Keenan, T. P.; Metcalf, C. A., III; Wang, Y.; Mani,
U.; Taylor, M.; Liu, S.; Bohacek, R. S.; Narula, S. S.; Dalgarno,
D. C.; van Schravandijk, M. R.; Violette, S. M.; Liou, S.; Adams,
S.; Ram, M. K.; Keats, J. A.; Weigele, M.; Sawyer, T. K. Bioorg.
Med. Chem. Lett. 2003, 13, 3063-3066.
Biological Materials and Methods. Lysate preparation
was carried out as previously described.15b
Antibodies. The 4G10 antibody to phosphotyrosine, mono-
clonal antibody to Src (GD11), and cortactin were obtained
from Upstate Biotechnologies. Antibodies to focal adhesion
kinase (FAK), PY925 FAK, PY421 cortactin, and PY418 Src
were obtained from BioSource.
Acknowledgment. The authors acknowledge the
Wyeth Analytical Chemistry Department for providing
the spectral data, and Carlo Etienne for in vivo technical
assistance. Additionally, we thank Dr. Tarek Mansour
for his support.
(10) Hanke, J. H.; Gardner, J. P.; Dow, R. L.; Changelian, P. S.;
Brissette, W. H.; Weringer, E. J.; Pollok, B. A.; Connelly, P. A.
Discovery of a novel, potent, and Src family-selective tyrosine
kinase inhibitor. Study of Lck- and FynT-dependent T cell
activation. J. Biol. Chem. 1996, 271, 695-701.
(11) Guan, H.; Laird, A. D.; Blake, R. A.; Tang, C.; Liang, C. Design
and synthesis of aminopropyl tetrahydroindole-based indolin-
2-ones as selective and potent inhibitors of Src and Yes tyrosine
kinase. Bioorg. Med. Chem. Lett. 2004, 14, 187-190.
Supporting Information Available: Elemental analysis
data for compounds 4-15, 16a-i, and 17a-k. This material
is available free of charge via the Internet at http://
pubs.acs.org.
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