The instant application is divided out of Indian Patent Application No.
1564/KOLNP/2005.
BACKGROUND OF THE INVENTION
5
Stroke is the third leading cause of death and the major cause of disability in the US,
where approximately 750,000 strokes occur each year. Ischemic stroke comprises
about 80% of this number, with primary intracerebral hemorrhagic stroke about 15-
20%. To date, the only approved efficacious treatment for acute ischemic cerebral
10 infarction is thrombolytic therapy by means of intravenous administration of t-PA,
recombinant tissue plasminogen activator. The usefulness of this therapy is
extremely limited. It must be given within a three hour window after the onset of
symptoms, while a majority of patients seek and/or receive treatment after a
substantial delay. In addition, treatment with t-PA carries an increased risk of
15 causing intracerebral hemorrhage, a potentially devastating complication. Presence
of hemorrhage must be ruled out prior to treatment and blood pressure must be
carefully managed and monitored during and after treatment with t-PA. Currently, no
neuroprotective therapy is available for treatment of ischemic stroke, hemorrhagic
stroke or brain trauma. New treatments for stroke and other conditions associated
20 with vascular permeability are greatly needed.
DESCRIPTION OF THE INVENTION
In accordance with the present invention are provided compounds of the structural
formula:
25
wherein:
X is N, CH

n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutically
acceptable salts thereof, with the proviso that when n is 1, X is not N.
Examples of alkyl of 1 to 3 carbon atoms include methyl, ethyl, n-propyl and i-propyl.
In some preferred embodiments of the invention, R' is methyl.
In other preferred embodiments of the invention, R is methyl or ethyl.
In still other embodiments of the invention, n is 2 or 3.
X is preferably N in some preferred embodiments of the invention.
In yet other preferred embodiments X is CH.
Pharmaceutically acceptable salts are those derived from such organic and inorganic
acids as: acetic, lactic, carboxylic, citric, cinnamic, tartaric, succinic, fumaric, maleic,
malonic, mandelic, malic, oxalic, propionic, hydrochloric, hydrobromic, phosphoric,
nitric, sulfuric, glycolic, pyruvic, methanesulfonic, ethanesulfonic, toluenesulfonic,
salicylic, benzoic, and similarly known acceptable acids.
Specific compounds of the invention include:
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(4-methyl-1-piperazinyl)-
propoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 7-[3-(4-ethyl-1-piperazinyl)propoxy]- 6-
methoxy-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[2-(4-methyl-1-piperazinyl)-
ethoxyl-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 7-[2-(4-ethyl-1 -piperazinyl)ethoxy]- 6-
methoxy-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 6-methoxy-7-[(1-methylpiperidin-4-yl)-
methoxyl)-3-quinolinecarbonitrile;
4-[(2,4-Dich!oro-5-methoxyphenyl)amino]- 6-methoxy-7-[2-(1-methylpiperidin-4-yl)-
ethoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile;

4-[(2,4-Dichloro-5-methoxyphenyi)amino]-7-[(1-ethylpiperidin-4-yl)methoxy]-6-
methoxyquinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-methylpiperazin-1-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[(1-methylpiperidin-4-yl)-
methoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-ethylpiperazin-1-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(4-methyl-1-piperazinyl)-
ethoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(1-methylpiperidin-4-yl)-
ethoxy]quinoIine-3-carbonitrile; and
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(4-propyl-1-piperazinyl)-
propoxy]-3-quinolinecarbonitrile; and pharmaceutically acceptable salts thereof.
Also provided is a process for the preparation of compounds of formula 1 wherein X is
CH and all other groups are as defined above, comprising:
(a) reacting a quinoline of formula

wherein R' is as defined herein, with an alcohol of formula

wherein R and n are as defined above, optionally in the presence of a base, e.g.
sodium hydride or sodium,

or (b) reacting a quinoline of formula

wherein R, R and n are as defined above and Y is chlorine or bromine,
with an aniline of formula

optionally in a suitable base, e.g. sodium hydride or pyridine hydrochloride,
or (c) cyclizing a compound of formula

to the desired quinoline, preferably under dehydrating conditions e.g. using
phosphorous oxychloride in acetonitrile, butyronitrile, toluene or xylene with alcohols
or amine bases as catalysts, at a suitable temperature e.g. 80-110°C, as described in
US 06/496,191.
The compounds of the invention are prepared as illustrated below. The compounds
of this invention were prepared from: (a) commercially available starting materials (b)
known starting materials which can be prepared as described in literature procedures
or (c) new intermediates described in the schemes and experimental procedures
herein.

Reactions are performed in a solvent appropriate to the reagents and materials
employed and suitable for the transformation being effected. It is understood by
those skilled in the art of organic synthesis that the various functionalities present on
the molecule must be consistent with the chemical transformations proposed. When
not specified, order of synthetic steps, choice of protecting groups and deprotection
conditions will be readily apparent to those skilled in the art. In addition, in some
instances, substituents on the starting materials may be incompatible with certain
reaction conditions. Restrictions pertinent to given substituents will be apparent to
one skilled in the art. Reactions were run under inert atmospheres where
appropriate.
Compounds of Formula I were prepared as described in Scheme 1. Compounds of
Formula 1 wherein R' is Me, X is N and n is 3 are readily obtained by treatment of
7-(3-chloropropoxy)-4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-3-quinoline-
carbonitrile, 1, with N-alkylpiperazine such as N-methylpiperazine, N-ethylpiperazine,
or N-propylpiperazine in the presence of sodium iodide either neat or in a solvent
such as ethylene glycol dimethyl ether. The preparation of these compounds has
been reported in the literature, [Boschelli, D. H., et. al., J. Med. Chem., 44, 3965
(2001)].
Analogously compounds of Formula 1 wherein R' is Me, X is N and n is 2 are readily
obtained by treatment of 7-(2-chloroethoxy)-4-[(2,4-dichloro-5-methoxyphenyl)-
amino]-6-methoxy-3-quinolinecarbonitrile, 2, with N-methyl or N-ethylpiperazine in
the presence of sodium iodide either neat or in a solvent such as ethylene glycol
dimethyl ether. The preparation of these compounds has been reported in the
literature, [Ye, F. et. al.,221th National Meeting of the American Chemical Society,
San Diego, California (April, 2001)].

Scheme 1

Alternatively compounds of Formula I can be prepared via a 7-fluoro-3-
quinolinecarbonitrile intermediate. Preparation of this key intermediate is shown in
Scheme 2. Anilines of formula 3 can be reacted with diethyl(ethoxymethylene)-
malonate either neat or in the presence of a cosolvent such as toluene, at
temperatures ranging from 60 to 120°C. Subsequent thermal cyclization, preferably in
a solvent system such as 3 : 1 mixture of diphenyl ether and biphenyl at elevated
temperature, such as 260°C, provides compounds of formula 4. Hydrolysis of the
ester group under preferably basic conditions, such as sodium hydroxide in an
alcoholic solvent such as ethanol, at elevated temperatures results in compounds of
formula 5. Conversion of the acid group to the primary amide can be accomplished
by treatment with an activating agent such as 1,1-carbonyldiimidazole followed by the
addition of either ammonia gas or preferably an aqueous solution of ammonium
hydroxide. Dehydration of the primary amide group of compounds of formula 6 with
a reagent such as cyanuric chloride in a solvent such as N.N-dimethylformamide
provides compounds of formula 7. Alternatively, anilines of formula 3 can be treated
with ethyl (ethoxymethylene)cyanoacetate either neat or in the presence of a
cosolvent such as toluene, at temperatures ranging from 60 to 120°C. Subsequent
thermal cyclization, preferably in a solvent system such as 3: 1 mixture of diphenyl
ether and biphenyl at elevated temperature, such as 260°C, provides compounds of
formula 7. Reaction of 7 with a chlorinating agent such as phosphorous oxychloride
gives compounds of formula 8.Treatment of compounds of formula 8 with 2,4-

dichloro-5-methoxyaniline in the presence of pyridine hydrochloride provides the key
7-fluro intermediates 9.

An alternate route to the compound of formula 8 where R' is Et is shown in Scheme
3. Using the conditions of Scheme 2, 4-benzyloxy-3~fluoroaniline is converted into the
compound of formula 10. Removal of the benzyl group with thioanisole and
trifluoroacetic acid provides the 6-hydroxy derivative of formula 11. Treatment of 11
with triphenyl phosphine, diethylazodicarboxylate and ethanol gives 'he compound of
formula 8 wherein R' is ethyl.


As shown in Scheme 4 reaction of compounds of formula 9 with an alcohol of formula
12 in the presence of a base such as sodium or sodium hydride provides the
compounds of the invention of Formula I. This reaction can be run in the presence of
a cosolvent such as dimethylformamide or dimethyl sulfoxide at optimal temperatures
of 120°C to 140°C.

Compounds of the present invention were evaluated in several standard
pharmacological tests that showed that compounds of the present invention inhibit
Src kinase and are useful for the prevention of vascular permeability.
Src Kinase Assay
Inhibitors of Src (partially purified enzyme preparation purchased from Upstate
Biotechnologies, Lake Placid, NY) tyrosine kinase activity are analyzed in an ELISA
format. The Boehringer Mannheim Tyrosine Kinase Assay Kit (Roche Diagnostics,

Basel, Switzerland) with a cdc2 substrate peptide containing Tyr15 is used for the
assay. Horseradish Peroxidase (HRP)-conjugated anti-phosphotyrosine is used to
detect phosphorylated peptide via a color reaction.
Reaction conditions: Five microliter aliquots of each compound prepared fresh at the
time of the assay are added as a solution in 10mM HEPES pH 7.5, 10% DMSO to
the reaction well. Thirty-five microliters of reaction mix containing Src, buffer and
peptide/bovine serum albumin mix are added to the compound wells and incubated
at 30°C for 10 minutes (reaction buffer: 50mM TrisHCI pH 7.5, 10mM MgCI2, 0.1 mM
EGTA, 0.5mM Na3VO4). The reaction is started by addition of 10 microliters of ATP
(500pM), incubated at 30°C for 1 hour, and stopped by addition of 20 microliters of
0.5M EDTA. The reaction mixture with the phosphorylated peptide is then transferred
to a streptavidin-coated microtiter plate and allowed to bind for 20 minutes. Unbound
peptide and reaction mixture is decanted and the plate is washed with PBS six times.
HRP-conjugated phosphotyrosine antibody supplied in the kit is incubated with the
plate for one hour, then decanted. The plate is again washed with PBS six times.
Substrate is added and absorbance at 405 nm is measured.
Alternatively, the assay performed essentially as described except a Delfia format
(Perkin-Elmer) is used and Europium-conjugated phosphotyrosine antibody was
used instead of HRP-conjugated phosphotyrosine antibody, Pierce Superblock was
used in place of bovine serum albumin and 6 washes were employed after the kinase
reaction and antibody binding. Europium fluorescence was used to monitor the
extent of reaction.
Activity is determined as % inhibition as calculated by the formula:
(1 - Abs/Abs(max)) x 100 = % inhibition. Where multiple concentrations of the test
agent are used, an IC50 (concentration which gives 50% inhibition) can be
determined. As shown in Table 1, compounds of the invention inhibit src kinase in
vitro.

Anchorage Independent Src-transformed Fibroblast Proliferation Assay
Rat2 fibroblasts stably transformed with a plasmid containing a CMV promotor
controlled v-Src/Hu c-Src fusion gene in which the catalytic domain of human c-Src
was inserted in place of the v-Src catalytic domain in the v-Src gene as follows:
Cloning and plasmid constructions. The Prague C v-Src gene from pSrcHis
(Wendler and Boschelli, Oncogene 4: 231-236; 1989) was excised with Ncol and
BamHI, treated with T4 DNA polymerase, and cloned into the Rl site of pTRE
(Clontech) that had been rendered flush by treatment with T4 DNA polymerase. The
PrC v-Src::hu c-Src fusion was created by replacing the Bgl2-Xbal fragment
encoding the carboxyl terminal ~ 250 amino acids of v-Src with the Bgl2-Xbal
fragment containing the v-Src::huc-Src fusion fragment (below). A partial clone of
human c-Src was amplified from a breast cDNA library (InVitrogen) using the
oligonucleotide pair
5'-CGCCTGGCCAACGTCTGCCCCACGTCCAAGCCGCAGACTCAGGGCCTG-S'
(SEQ ID NO: 1) and
5'-CCAACACACAAGCAGGGAGCAGCTGGGCCTGCAGGTACTCGAAGGTGGGC-
3' (SEQ ID NO: 2) and cloned into pCRScript (Stratagene). The catalytic domain of
human c-Src in this clone was amplified with these oligonucleotides (fuses v-src
nucleotide 734 to human c-Src nucleotide 742 and human c-Src nucleotide 1551 to
v-src nucleotide 1543 in the v-Src and human c-Src ORFs). Two v-Src sequences
were amplified by PCR (198 base pair v-src 5' fragment:
5'-GTGCCTATTGCCTCTCCGTTTCTGAC-3' (SEQ ID NO: 3) (primer 1) to
5'-ACGTGGGGCAGACGTTGGCCAGGCG-3') (SEQ ID NO: 4) (252 base pair
3' v-src fragment, 5'-CAGCTGCTCCCTGCTTGTGTGTTGG-3' (SEQ ID NO: 5)
(residues 1543-1567 in v-src ORF) to
5'-ATGAATTCTCTAGAGGMGACGCCATCATATTCCAAGCAG-3' (SEQ ID NO: 6)
residues 1769-1794 from v-src ATG with Xbal and EcoRI restriction sites added
(primer 4)). Primers 1 and 4 were used to generate a three-fragment PCR
amplification and fusion of the v-Src::human c-Src fusion fragment and the 5' and 3'
fragments amplified from the Prague C v-Src gene and 3'untranslated region from
Rous sarcoma virus. This reaction creates an in-frame v-Src::human c-Src gene
fusion (amino acid residue V244 of v-Src to C248 of human c-Src on the amino

terminal side and A517 of human c-Src to Q515 of v-Src). This gene fusion fragment
encodes the carboxyl terminal one-third of the v-Src SH2 domain and SH2-catalytic
domain linker fused to the human c-Src catalytic domain flanked by the v-Src
carboxyl-terminal tail. A naturally occurring Bgl2 site near the 5' end of the fusion
fragment and the engineered Xbal site at the 3' end of the fragment were used to
excise fragment for creation of the full-length v-Src::human c-Src fusion gene as
described above. The integrity of the constructs was confirmed by DNA sequencing.
Similar methods were used to clone this gene into other expression plasmids such as
pIRES (Clontech) for use in these studies.
These transformed Rat2 fibroblasts are used for the measurement of src dependent
suspension growth.
Ultra-low cluster plates (Corning Costar, Acton, MA) are seeded with 10,000 cells per
well on Day 1. Alternatively, Ultra-low cluster plates (Costar 3474) treated with
Sigmacote (Sigma, St. Louis, MO), rinsed with 70% ethanol, after drying in the hood,
are seeded with 5000 cells. Compound is added in serial two-fold dilutions from 10
micromolar to 0.009 micromolar on Day 2 and MTS reagent (Promega, Madison, Wl)
is added on Day 5 (100 microliters of MTS/medium mix +100 microliters of medium
already on the cells and the absorbance is measured at 490nm. The results are
analyzed as follows to yield an IC50 for proliferation (micromolar units) as follows:
%inhibition = (Abs490 nm sample - b!ank)/(Abs490 nm no cmpd control - blank) X
100%. As shown in Table 1, compounds of the present invention inhibit src
dependent cell proliferation.


IP administration of Example 1 provides neuroprotection in transient model of
focal ischemia
Example 1 was tested in a rat model of transient focal ischemia. Wistar rats were
subjected to a 90 min occlusion of the middle cerebral artery (MCA) using an
intraluminal suture approach as described by Longa et al., Stroke 1989, 20:84
followed by reperfusion for 48 hours. Eighty-five minutes after the initial onset of
ischemia, animals received compound of Example 1(1.5, 5, 15, or 45 mg/kg ip).

Following reperfusion, the animals were evaluated over a 48 hour period for
neurological function deficit and weight loss/gain. Infarct size was measured
following sacrifice at 48 hours post MCA occlusion. Example 1 at doses of 5 and 45
mg/kg significantly improved recovery from stroke-induced neurological deficits.
Reductions in the volume of infarcted brain tissue were observed at most doses of
Example 1 but statistical significance was achieved only at the 45 mg/kg ip dose.
Improvement in body weight recovery was observed in animals treated with
Example 1.
IV administration of Example 1 provides neuroprotection in transient model of
focal ischemia
Wistar rats were subjected to a 90 min occlusion of the middle cerebral artery (MCA)
using an intraluminal suture approach as described by Longa et a!., Stroke 1989,
20:84 followed by reperfusion for 48 hours. Thirty minutes after MCA occlusion, an
intravenous formulation of Example 1 in 20 mM citrate/0.85% saline, pH 3 was
administered at doses of 3, 10 and 30 mg/kg (iv). Following reperfusion, the animals
were evaluated over a 48 hour period for neurological function deficit and weight
loss/gain. Brain tissue infarction volume by were reduced by 22%, 53% and 42%,
respectively. Post-stroke weight loss was also significantly reduced. In addition, as
shown in Table 2, stroke-induced neurological deficits were significantly reduced at
all three doses. Thus, compounds of the present invention provide neuroprotection
following focal ischemia.


Therapeutic window
In this model of transient focal ischemia three studies were conducted to examine
therapeutic window. Wistar rats were subjected to 90 minute occlusion of the MCA
followed by reperfusion as described above. A single bolus of 10mg/kg Example 1
was administered at 30 minutes, 90 minutes, 3 hours, 4 hours, 5 hours and 6 hours
post stroke. Volume of infarcted tissue was measured by histological staining. Brain
tissue infarction was statistically reduced (as a % of vehicle treated) with a single
10mg/kg dose of Example 1 administered between 30 min and 4 hours after the
ischemic injury. Statistically significant protection from neurological deficits (as a
percent of vehicle treated) was achieved with a single 10mg/kg dose of Example 1
administered up to 5 hours post-stroke, and statistically significant protection from
ischemia-induced weight loss (as a % of vehicle treated) was achieved with a single
10mg/kg dose of Example 1 up to 5 hours post stroke. Thus, compounds of the
present invention exhibit a superior therapeutic window compared to presently
available treatments.
Post Ischemic vascular permeability
Wistar rats were subjected to a 90 min occlusion of the middle cerebral artery (MCA)
using an intraluminal suture approach as described by Longa et a/., Stroke 1989,
20:84 followed by reperfusion for 24 hours. Compound of Example 1 was
administered as a single IV bolus at 30 minutes after onset of ischemia at 3,10 and
30 mg/kg (iv). Two hours before sacrifice animals received an IV injection of 2%
Evans Blue in saline. Brains were perfused with saline and the striatum dissected.
Evans Blue was extracted and quantified by spectrofluorometer based on external
standards. Vascular permeability in the ischemic striatum was reduced as evidenced
by a 60% decrease of Evans Blue extravasation. Thus, compounds of the present
invention reduce vascular permeability associated with ischemic injury.
Permanent Focal Ischemia
Example 1 was also evaluated in two rat models of permanent focal ischemia. In a
model of extreme severity (intraluminal suture occlusion of internal carotid artery) and
a relatively short outcome (28 hours) little or no effect was shown.

In a model producing extensive infarction to sensorimotor cortex with quantitative
assessment of neurological deficits for 21 days post-stroke, compound of Example 1
provided significant improvement in the neurological outcome after stroke. Wistar
rats ( n=5 per group) were subjected to focal ischemic stroke model that results in a
extensive ischemia to the sensorimotor cortex substantially as described by Chen et
al., (Stroke 17:738, 1986). Example 1 or vehicle was administered as an IV bolus at
10 mg/kg at 90 minutes post-induction of stroke, 4 hours later, and at 24 and 28
hours later (total dose 40 mg/kg). Animals were evaluated for sensorimotor deficits
(postural reflex, visual and tactile forelimb placement and hindlimb placement tests)
on days 1, 2, 4, 7, 9, 11, 14, 16, 18 and 21 after induction of ischemia. Results were
evaluated by Generalized Regression Model to determine statistical significance of
the differences between slopes and to compare final neurological outcome after 21
days. By Day 21, there was statistically significant improvement in Behavioral Score
for subjects treated with Example 1 as compared to a control group. Thus,
compounds of the present invention provide long-term improvement of neurological
deficits.
Vascular permeability due to disease, injury, or other trauma, may occur in a variety
of tissues and organs including organs of the central nervous system, cardio-
pulmonary system, gastrointestinal system and renal system. Compounds of the
present invention are useful for inhibiting vascular permeability caused by disease,
injury, or other trauma. In particular, vascular permeability may be inhibited in
cerebral and spinal tissue following cerebrovascular events. Vascular permeability is
a major cause of vascular leakage and/or edema following a cerebrovascular event
and often leads to neurological disorders and disabilities. Cerebrovascular events
including, but not limited to transient and acute ischemic events, may be treated in
accordance with the present invention. Acute events include, but are not limited to,
stroke, head trauma, spinal trauma, general anoxia, hypoxia including fetal hypoxia,
hypoglycemia, hypotension as well as similar injuries seen during procedures from
embole, hyperfusion and hypoxia. Stroke includes, but is not limited to focal and
global ischemia, transient cerebral ischemic attacks, and other cerebral vascular
problems accompanied by cerebral ischemia. The instant invention would also be
useful in a range of cerebrovascular events including cerebral hemmorhage,
infarction due to embolism or thrombosis of the intra- or extra cranial arteries,

perinatal asphyxia, in cardiac arrest and status epilepticus, especially where blood
flow to the brain in halted for a period of time. Cerebrovascular events associated
with vascular leakage also include infections, including, but not limited to encephalitis
and meningitis associated with neuroinflammation, which, through vascular leakage
propagate injury to surrounding tissues. Systemic disease such as diabetes, multiple
sclerosis, kidney disease and atherosclerosis may also result in increased vascular
permeability. Compounds of the present invention are also useful for inhibiting
vascular permeability triggered by any local tissue/organ ischemic (hypoxic) event
outside of the central nervous system, including, but not limited to myocardial
ischemia and ischemic bowel disease.
Compounds of the present invention provide neuroprotection in a patient.
Neuroprotection, as used herein, refers to the protection of neural cells against cell
death or apoptosis. One measure of the extent of cell death or apoptosis is infarct
volume; the volume of necrotic or dead brain tissue. Imaging techniques and the
patient's clinical status can be used to assess infarct volume following an ischemic
event. Compounds of the present invention reduce infarct volume of a patient as
compared to typical infarct volume experienced in similar ischemic events in the
absence of agents of the present invention.
Compounds of the present invention prevent, reduce or inhibit neurodegeneration
and/or neurotoxicity associated with vascular permeability that result in symptoms
including, but not limited to, visual impairment, speech impairment, memory
impairment, cognitive impairment or dysfunction, and motor impairment including, but
not limited to, paralysis. Neurological deficits resulting from injury or disease
described above may be inhibited or prevented in accordance with the present
invention. Thus, the present invention provides methods of treating, preventing,
inhibiting or alleviating conditions associated with vascular leakage or permeability
listed above in a mammal, preferably in a human, the methods comprising providing
a pharmaceutically effective amount, and in particular a vascular permeability
inhibiting amount, of a compound of this invention to the mammal, and in particular a
human patient, in need thereof.

Also encompassed by the present invention are pharmaceutical compositions for
treating or modulating vascular permeability comprising at least one compound of
Formula I, mixtures thereof, and or pharmaceutical salts thereof, and a
pharmaceuticaHy acceptable carrier therefore. Such compositions are prepared in
accordance with acceptable pharmaceutical procedures, such as described in
Remingtons Pharmaceutical Sciences, 17th edition, ed. Alfonoso R. Gennaro, Mack
Publishing Company, Easton, PA (1985). PharmaceuticaHy acceptable carriers are
those that are compatible with the other ingredients in the formulation and biologically
acceptable.
Liquid carriers may be used in preparing solutions, suspensions, emulsions, syrups
and elixirs including intravenous solutions. The active ingredient of this invention can
be dissolved or suspended in a pharmaceuticaily acceptable liquid carrier such as
water, organic solvent, or a mixture of both. The liquid carrier can contain other
suitable pharmaceutical additives such as solubilizers, emulsifiers, buffers,
preservatives, sweeteners, flavoring agents, suspending agents, thickening agents,
colors, viscosity regulators, stabilizers, osmo-regulators, antioxidants and
antifoaming agents.
Suitable examples of liquid carriers for oral, intravenous and parenteral
administration include water (particularly containing additives as above e.g. cellulose
derivatives, preferably sodium carboxymethyl cellulose solution), saline, dextrose
solutions, dextrose-saline and dextrose-water solutions, alcohols (including
monohydric alcohols and polyhydric alcohols e.g. glycols) and their derivatives.
Liquid carriers are used in sterile form for parenteral and intravenous administration.
PH of liquid formulations may be adjusted in some cases by the addition of HCI,
sodium hydroxide, and phosphoric acid. Preferably compositions of the present
invention are liquid pharmaceutical compositions which are sterile solutions or
suspensions in an iso-osmotic, physiologically compatible buffered system.
Liquid pharmaceutical compositions of the present invention can be administered by,
for example, intramuscular, intraperitoneal, intravenous, or subcutaneous injection.
Pharmaceutical compositions of the present invention are preferably administered to
a patient by intraperitoneal or intravenous injection. Most preferably, the composition

is administered intravenously such as by intravenous bolus injection, intravenous i.v.
drip, repeated slow bolus administration or infusion.
Oral administration may be either liquid or solid composition form. The compounds of
this invention may also be administered orally or parentally, neat or in combination
with conventional pharmaceutical carriers. Applicable solid carriers can include one
or more substances which may also act as flavoring agents, lubricants, solubilizers,
suspending agents, fillers, glidants, compression aids, binders or tablet-disintegrating
agents or an encapsulating material. In powders, the carrier is a finely divided solid,
which is in admixture with the finely divided active ingredient.. In tablets, the active
ingredient is mixed with a carrier having the necessary compression properties in
suitable proportions and compacted in the shape and size desired. The powders and
tablets preferably contain up to 99% of the active ingredient. Suitable solid carriers
include, for example, calcium phosphate, magnesium stearate, talc, sugars, lactose,
dextrin, starch, gelatin, cellulose, methyl cellulose, sodium carboxymethyl cellulose,
polyvinylpyrrolidine, low melting waxes and ion exchange resins.
Preferably the pharmaceutical composition is in unit dosage form, e.g. as tablets,
capsules, powders, solutions, suspensions, emulsions, granules, suppositories,
ampule, or bolus. In such form, the composition is sub-divided in unit dose
containing appropriate quantities of the active ingredient; the unit dosage forms can
be packaged compositions, for example packeted powders, lyophilyzed powder or
cake in ampoules or vials, or vials, ampoules, prefilled syringes or sachets containing
liquids. The unit dosage form can be, for example, capsule or tablet itself, or it can
be the appropriate number of any such compositions in package form.
The dose provided to a patient will vary depending upon what is being administered,
the purpose of the administration, such as prophylaxis or therapy, and the state of
the patient, the manner of administration, and the like. A "therapeutically effective
amount" is an amount sufficient to cure or ameliorate symptoms of a disease or
injury. Generally, a single dose (or dosage form) will contain from about 1 mg/kg to
about 30 mg/kg, and more preferably from about 1 mg/kg to about 10 mg/kg of
compound of the present invention. It is expected that some patients will receive
multiple doses. The dosage to be used in the treatment of a specific case must be

subjectively determined by the attending physician. The variables involved include
the specific condition and the size, age and response pattern of the patient.
The present invention provides advantages over previously known treatments for
stroke and other conditions associated with vascular permeability. In particular, while
it is preferable to treat patients as soon as possible after an ischemic injury,
compounds of the present invention may be effective in preventing
neurodegeneration and development of neurological deficits in some patients when
administered even up to about 18-24 hours after ischemic injury. Furthermore,
treatment may continue and improvement in a patient's prognosis may result from
continuous or repeated administration of compound of the present invention for up to
about 72 hours or longer following ischemic injury.
One embodiment of the invention involves the administration of the compound
between about 6 to 24 hours after the ischemic event. A further embodiment
involves the administration of the compound between about 18 to 24 hours after the
ischemic event.
Provide as used herein means either directly administering a compound or
composition of the present invention, or administering a prodrug, derivative or analog
which will form an equivalent amount of the active compound or substance within the
body.
The present invention includes prodrugs of compounds of Formula I. "Prodrug", as
used herein means a compound which is convertible in vivo by metabolic means
(e.g. by hydrolysis) to a compound of Formula I. Various forms of prodrugs are
known in the art, for example, as discussed in Bundgaard, (ed.), Design of Prodrugs,
Elsevier (1985); Widder, et al. (ed.), Methods in Enzymology, vol. 4, Academic Press
(1985); Krogsgaard-Larsen, et al., (ed). "Design and Application of Prodrugs,
Textbook of Drug Design and Development, Chapter 5, 113-191 (1991), Bundgaard,
et al., Journal of Drug Deliver Reviews, 8:1-38(1992), Bundgaard, J. of
Pharmaceutical Sciences, 77:285 et seq. (1988); and Higuchi and Stella (eds.)
Prodrugs as Novel Drug Delivery Systems, American Chemical Society (1975).

This invention will be more fully described in conjunction with the following specific
examples which are not to be construed as limiting the scope of this invention.
Reference Example 1
Ethyl 7-fluoro-6-methoxy-4-oxo-1,4-dihvdro-3-quinolinecarboxvlate
A mixture of 3-fluoro-4-methoxyaniline (3.00 g, 21.26 mmol) and diethyl ethoxy-
methylene malonate (4.59 g, 21.26 mmol) was heated at 110°C for 1 hour then
cooled to room temperature. Hexane was added and the solids were collected by
filtration. This material was suspended in 45 mL of a 3 : 1 mixture of diphenyl ether:
biphenyl and the mixture was heated at reflux for 2 hours to provide a brown solution.
The reaction mixture was cooled to room temperature and hexane was added. The
resultant solid was collected by filtration washing with hexane to provide 2.62 g of
ethyl 7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quinolinecarboxylate as a white solid,
mp >300°C.
MS 265.9 (M+H)+
Analysis for C13H12FNO4
Calcd: C, 58.87; H, 4.56; N, 5.28.
Found: C, 58.66; H, 4.16; N, 5.14.
Reference Example 2
7-Fluoro-6-methoxy-4-oxo-1,4-dihvdro-3-quinolinecarboxvlic acid
A mixture of ethyl 7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quinolinecarboxylate (2.2
g, 8.30 mmol) and 13.2 mL of 1 N sodium hydroxide and 40 mL of ethanol was
heated at reflux for 3 hours then cooled to room temperature. Water was added and
the mixture was acidified with acetic acid. The resultant solid was collected by
filtration washing with water to provide 1.90 g of 7-f!uoro-6-methoxy-4-oxo-1,4,-
dihydro-3-quinolinecarboxylic acid as a white solid, mp 265-267°C.
MS 238.1 (M+H)+
Analysis for C11H8FNO4 - 1.2 H2O
Calcd: C, 51.04; H, 4.03; N, 5.41.
Found: C, 50.98; H, 3.95; N, 5.33.

Reference Example 3
7-Fluoro-6-methoxy-4-oxo-1,4-dihvdro-3-quinolinecarboxamide
A mixture of y-fluoro-e-methoxy-4-oxo-1,4-dihydro-3-quinotinecarboxylic acid (1.0 g,
4.21 mmol) and 1,1''-carbonyldiimidazole (1.51 g, 9.28 mmol) in 14 mL of N,N-
dimethylformamide was heated at 65°C for 2 hours then cooled to room temperature
and poured into 200 mL of aqueous ammonium hydroxide on an ice water bath. The
solution was allowed to stir at room temperature overnight and then concentrated to
a small volume. Ice cold water was added followed by acidification with acetic acid.
The resultant solid was collected by filtration washing with water to provide 821 mg of
7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quinolinecarboxamide as a white solid, mp
>300°C.
MS 236.8 (M+H)+
Analysis for C11H9FN2O3 - 0.2 H2O
Calcd: C, 55.09; H, 3.94; N, 11.68.
Found: C, 55.00; H, 3.63; N, 11.49.
Reference Example 4
7-Fluoro-6-methoxy-4-oxo-1,4-dihvdro-3-quinolinecarbonitrile
A mixture of 7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quinolinecarboxamide (700 mg,
3.0 mmol) and cyanuric chloride (341 mg, 1.65 mmol) in 15 mL of N,N-dimethyl-
formamide was heated at 65°C for 6 hours then cooled to room temperature and an
additional 206 mg of cyanuric chloride was added. The mixture was heated at 65°C
for 4 hours then stirred overnight at room temperature. The reaction mixture was
poured into ice water and neutralized with saturated sodium bicarbonate. The solids
were collected by filtration washing with water and hexane to provide 610 mg of
crude product. Purification by flash column chromatography eluting with a gradient of
3% methanol in dichloromethane to 10% methanol in dichloromethane, provided 272
mg of 7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quinolinecarbonitrile, mp 147-149°C.
MS 216.8 (M-H)-
Analysis for C11H7FN2O2- 0.1 dichloromethane
Calcd: C, 58.80; H, 3.19; N, 12.36.
Found: C, 59.06; H, 2.96; N, 11.97.

Alternative route to Reference Example 4
7-Fluoro-6-methoxv-4-oxo-1,4-dihvdro-3-quinolinecarbonitrile
A mixture of 3-fIuoro-4-methoxyaniline (15.31 g, 108 mmol) and ethyl(ethoxy-
methy!ene)cyanoacetate (18.36 g, 108 mmol) in toluene was heated at 100-110DC for
4.5 hours then cooled to room temperature. A 1 : 1 mixture of hexane and ethyl
acetate was added and the mixture was cooled on an ice bath. The solids were
collected washing with hexane to provide a first crop of 26.10 g and a second crop of
1.24 g. A 2.0 g portion of this material was added to 18 mL of a 3 : 1 mixture of
diphenyl ether: biphenyl that was heated to reflux. This mixture was heated at reflux
for 4 hours then cooled and poured into hexane. The solids were collected by
filtration and washed with ethyl acetate and hexane to provide 624 mg of 7-fluoro-6-
methoxy-4-oxo-1,4,-dihydro-3-quinolinecarbonitrile as a brown solid. The filtrate was
concentrated, the residue was dissolved in ethyl acetate and hexane was added.
The resultant solid was collected by filtration to give 1.07 g of 7-fluoro-6-methoxy-4-
oxo-1,4-dihydro-3-quinolinecarbonitri!e as a yellow solid.
Reference Example 5
4-Chloro-7-fluoro-6-methoxy-3-quinolinecarbonitrile
A mixture of 7-fluoro-6-methoxy-4-oxo-1,4-dihydro-3-quino!inecarbonitrile (1.0 g, 4.59
mmol) and 14 g of phosphorous oxychloride was heated at reflux for 30 minutes
then concentrated in vacuo. The residue was partitioned between aqueous sodium
bicarbonate and ethyl acetate. The organic layer was dried over magnesium sulfate,
filtered and concentrated on to silica gel. Purification by flash column
chromatography eluting with a gradient of 1 : 5 ethyl acetate : hexane to 1 :1 ethyl
acetate : hexane, provided 631 mg of 4-chloro-7-fluoro-6-methoxy-3-quinolinecarbo-
nitrile, mp 160-162°C.
MS 236.9 (M+H)+
Analysis for C11H6CIFN2O
Calcd: C, 55.83; H, 2.56; N, 11.84.
Found: C, 55.66; H, 2.84; N, 11.91.

Reference Example 6
4-[(2,4-Pichloro-5-methoxvphenvl)amino1-7-fluoro-6-methoxv-3-quinolinecarbonitrile
A mixture of 4-chloro-7-fluoro-6-methoxy-3-quinoIinecarbonitrile (4.12 g , 18 mmol)
2,4-dichloro-5-methoxyaniIine (4.56 g, 24 mmol) (Theodoridis, G.; Pestic. Sci. 1990,
30, 259) and pyridine hydrochloride (2.31 g, 19.9 mmol) in 45 mL of 2-ethoxyethanol
was heated at 120°C for 3.hours then cooled to room temperature. The reaction
mixture was added to aqueous sodium bicarbonate and stirred for 20 minutes. The
solids were collected by filtration to provide 4.89 g of 4-[(2,4-dichloro-5-methoxy-
phenyl)amino]-7-fluoro-6-methoxy-3-quinolinecarbonitrile, mp >260°C.
HRMS theory 392.03634; found 392.03556 (M+H)+
Analysis for C-18H12CI2FN3O2 2.0 H2O
Calcd: C, 50.48; H, 3.77; N, 9.81.
Found: C, 50.41; H, 2.82; N, 9.78.
Reference Example 7
6-Benzyloxy-7-fluoro -4-oxo-1,4-dihvdro-3-quinolinecarbonitrile
A mixture of 4-benzyloxy-3-f!uoroaniline (6.06 g, 27.9 mmol) (US 5,622,967) and
ethyl (ethoxymethylene)cyanoacetate (5.08 g, 30.0 mmol) was heated at 120°C for
45 minutes then cooled to room temperature. This solid was added in portions to a
3 : 1 mixture of dipheny! ether: biphenyl at 245°C. This mixture was heated at 245°C
for 3 hours then cooled and the solids were collected by filtration, washing with
hexane and diethyl ether to provide 2.60 g of 6-benzyloxy-7-fluoro-4-oxo-1, 4-
dihydro-3-quinolinecarbonitrile, mp >250°C.
MS 293.1 (M-H)-
Reference Example 8
6-Benzyloxy-4-chloro-7-fluoro-3-quinolinecarbonitrile
A mixture of 6-benzyloxy-7-fluoro-4-oxo-1,4-dihydro-3-quinolinecarbonitrile (645 mg,
2.19 mmol) and 10 mL of phosphorous oxychloride was heated at 115 °C for 1.5
hours then concentrated in vacuo. The residue was treated with ice cold aqueous
ammonium hydroxide and the resultant solid was collected by filtration. Purification
by flash column chromatography eluting with a gradient of 1% ethyl acetate in

hexane to 6% ethyl acetate in hexane, provided 284 mg of 6-benzyloxy-4-chloro-7-
fluoro-3-quinolinecarbonitrile, mp 159-160°C.
MS 313.13 (M+H)+
Analysis for C17H10ClFN20
Calcd: C, 65.15; H, 3.06; N, 8.82.
Found: C, 65.29; H, 3.22; N, 8.96.
Reference Example 9
4-Chloro-7-fluoro-6-hydroxy-3-quinolinecarbonitrile
A mixture of 6-benzyloxy-4-chloro-7-fluoro-3-quinolinecarbonitrile (733 mg, 2.34
mmol) and 1 mL of thioanisole in 12 mL of trifluoroacetic acid was heated at reflux
for 9 hours then concentrated in vacuo. The residue was treated with ice water and
then basified to pH 9-10 by the addition of aqueous ammonium hydroxide. The
resultant solid was collected by filtration and washed with diethyl ether. The filtrate
was extracted with 10% methanol in ethyl acetate. The organic layer was dried over
sodium sulfate, filtered and concentrated in vacuo. The residue was combined with
the solid obtained initially, and this material was dissolved in 5% methanol in ethyl
acetate and absorbed onto silica gel. Purification by flash column chromatography
eluting with a gradient of hexane to increasing amounts of ethyl acetate in hexane to
5% methanol in ethyl acetate provided 260 mg of 4-chloro-7-fluoro-6-hydroxy-3-
quinolinecarbonitrile, mp >250°C.
MS 220.9 (M-H)-
Analysis for C10H4CIFN2O
Calcd: C, 53.96; H, 1.81; N, 12.58.
Found: C, 54.23; H, 2.02; N, 12.06.
Reference Example 10
4-Chloro-6-ethoxy-7-fluoro-3-quinolinecarbonitrile
To a 0°C mixture of 4-chloro-7-fluoro-6-hydroxy-3-quinolinecarbonitrile (185 mg, 0.83
mmol), triphenylphosphine (392 mg, 1.49 mmol) and ethanol (153 mg, 3.32 mmol)
in 15 mL of tetrahydrofuran was added diethylazodicarboxylate (260 mg, 1.80
mmol). The reaction mixture was kept at 0ºC for 45 minutes then stirred at room
temperature overnight. The reaction mixture was concentrated in vacuo and purified

by flash column chromatography eluting with a gradient of 1% ethyl acetate in
hexane to 5% ethyl acetate in hexane to provide 4-chloro-6-ethoxy-7-fluoro-3-
quinolinecarbonitrile, mp 165-166°C.
MS 251.0 (M+H)+
Analysis for C12H8CIFN2O
Calcd: C, 57.50; H, 3.22; N, 11.18.
Found: C, 57.24; H, 3.41; N, 11.09.
Reference Example 11
4-[(2,4-Dichloro-5-methoxvphenvl)amino]-6-ethoxy-7-fluoro-3-quinolinecarbonitrile
Following the procedure of Reference Example 6, a mixture of 4-chloro-6-ethoxy-7-
fluoro-3-quinolinecarbonitrile (197 mg, 0.78 mmol), 2,4-dichloro-5-methoxyani!ine
(220 mg, 1.14 mmol) and pyridine hydrochloride (120 mg, 1.04 mmol) provided, after
flash column chromatography eluting with a gradient of dichloromethane to 1%
methanol in dichloromethane, 183 mg of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-
ethoxy-7-fluoro-3-quinolinecarbonitrile, mp 184-186°C.
MS 406.0 (M+H)
Analysis for C19H14CI2FN3O2 0.5 H2O
Calcd: C, 54.96; H, 3.64; N, 10.12.
Found: C, 54.99; H, 3.59; N, 10.05.
Example 1
4-[(2,4-Dichloro-5-methoxvphenyl)aminol- 6-methoxy-7-f3-(4-methyl-1-
piperazinvl)propoxy]-3-quinolinecarbonitrile
A mixture of 7-[3-chloropropoxy]-4-[(2,4-dichloro-5-methoxyphenyl)arnino]-6-
methoxy-3-quinolinecarbonitrile (656 mg, 1.40 mmol) and sodium iodide (210 mg,
1.40 mmol) in 4 mL of N-methylpiperazine was heated at 80°C for 20 h. The reaction
mixture was concentrated in vacuo and partitioned between ethyl acetate and
saturated aqueous sodium bicarbonate. The organic layer was washed with brine,
dried over sodium sulfate, filtered and concentrated in vacuo. The residue was
purified by column chromatography eluting with 30% methanol in dichloromethane.
The fractions containing product were collected and concentrated in vacuo. Diethyl
ether was added to the residue and the light pink solid was collected by filtration to

provide 560 mg (75%) of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-
(4-methyl-1-piperazinyl)propoxy]-3-quinolinecarbonitrile: mp 116-120°C; MS (ES) m/z
530.2, 532.2 (M+1).
Example 2
4-[(2,4-Dichloro-5-methoxvphenvl)amino1- 7-[3-(4-ethyl-1 -piperazinyl)propoxy]- 6-
methoxv-3-quinolinecarbonitrile
A mixture of 7-[3-chloropropoxy]-4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-
methoxy-3-quinolinecarbonitrile (3.50g, 7.50 mmol), sodium iodide (1.12 g, 7.50
mmol) and 4.8 mL of N-ethylpiperazine in 5 mL of ethylene glycol dimethyl ether was
heated at 95°C for 20 h. The reaction mixture was concentrated in vacuo and
partitioned between ethyl acetate and saturated aqueous sodium bicarbonate. The
organic layer was washed with saturated aqueous sodium bicarbonate, followed by
brine, dried over sodium sulfate, filtered and concentrated in vacuo. Diethyl etherwas
added to the residue and the white solid was collected by filtration to provide 1.80 g
(44%) of 4-[(2,4-dichIoro-5-methoxyphenyl)amino]-7-[3-(4-ethyl-1-piperazinyl)-
propoxy]-6-methoxy-3-quinolinecarbonitrile: mp 102-104°C; MS (ES) m/z 544.3,
546.4 (M+1).
Example 3
4-[(2,4-Dichloro-5-methoxvphenyl)aminol-6-methoxy-7-[2-(4-methyl-1-
piperazinyl)ethoxy]-3-quinolinecarbonitrile
Prepared according to the method used for the preparation of Example 1 by the
reaction of 7-[2-chloroethoxy]-4-[(2,4-dichIoro-5-methoxyphenyl)amino]-6-methoxy-3-
quinolinecarbonitrile and N-methylpiperazine: mp 165-167°C; MS (ES) m/z 516.0,
518.2 (M+1).
Example 4
4-[(2,4-Dichloro-5-methoxvphenyl)amino]-7-[2-(4-ethyl-1-piperazinyl)ethoxy]-6-
methoxy-3-quinolinecarbonitrile
Prepared according to the method used for the preparation of Example 1 by the
reaction of 7-[2-chloroethoxy]-4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-3-

quinolinecarbonitrile and N-ethylpiperazine: mp 101-105°C; MS (ES) m/z 530.4,
532.4 (M+1).
Example 5
4-[(2,4-Dichloro-5-methoxyphenyl)aminol-6-methoxy-7-[(1-methylpiperidin-4-
yl)methoxy]-3-quinolinecarbonitrile
To a solution of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-7-[luoro-6-methoxy-3-
quinolinecarbonitrile (600 mg, 1.53 mmol) and i-methylpiperidine-4-methanol (395
mg, 3.06 mmol) in 10 mL of N, N-dimethylformamide at 135°C was added sodium
hydride (362 mg, 9.06 mmol) in portions. After 45 minutes the reaction mixture was
poured into saturated sodium bicarbonate. After stirring for 15 minutes the solid was
collected by filtration. The residue was purified by flash column chromatography,
eluting with a gradient of 5% methanol in dichloromethane to 25% methanol in
dichloromethane. Trituration with diethyl ether provided 396 mg of 4-[(2,4-dichloro-5-
methoxyphenyl)amino]-6-methoxy-7-(1-methylpiperidin-4-yl)methoxyJ-3-quinoline-
carbonitrile, mp 200-202°C.
MS 501.3 (M+H)+
Analysis for C25H26CI2N4O3 - 0.8H2O
Calcd: C, 58.21; H, 5.39; N, 10.86.
Found: C, 58.19; H, 5.23; N, 10.67.
Example 6
4-r(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[2-(1-methylpiperidin-4-
yl)ethoxy]-3-quinolinecarbonitrile
A mixture of sodium hydride (128 mg, 3.2 mmol) and 1-methyl-4-piperidineethanol
(180 mg, 1.25 mmol) [ EP 0581538] in 5 mL of N, N-dimethylformamide was heated
at 110°C for 1 hour. 4-[(2,4-Dichloro-5-methoxyphenyl)amino]-7-fluoro-6-methoxy-3-
quinolinecarbonitrile (200 mg, 0.51 mmol) was added and the reaction mixture was
heated at 135°C for 5 hours. Over the next 4 hours an additional 128 mg of sodium
hydride was added to the reaction mixture at 130°C. The reaction mixture was
partitioned between ethyl acetate and water. The organic layer was dried over
sodium sulfate, filtered and concentrated in vacuo. The residue was purified by
preparative thin layer chromatography, eluting with 20% methanol in

dichloromethane to provide 105 mg of 4-[(2,4-dichloro-5-methoxyphenyl)amino]- 6-
methoxy-7-[(2-(1 -methylpiperidin-4-yl)ethoxy]-3-quinolinecarbonitrile, mp 190-191°C.
MS 515.19 (M+H)+
Analysis for C26H28CI2N4O3 -1.0 H2O
Calcd: C, 58.53; H, 5.67; N, 10.50.
Found: C, 58.65; H, 5.57; N, 10.34.
Examples 7 and 8 are obtained analogously by the method of Example 5 and the
corresponding alcohol.
Example 7
4-[(2,4-Dichloro-5-methoxyphenyl)annino]-6-methoxy-7-[3-(1-methvlpiperidin-4-
yl)propoxy]quinoline-3-carbonitriie
MP 144-145°C; Mass spec. 529.2 (ES +)
Example 8
4-[(2,4-Dichloro-5-methoxyphenvnaminol-7-[(1-ethvlpiperidin-4-yl)methoxy]-6-
methoxyquinoline-3-carbonitrile
MP 192-195°C; Mass spec. 515.2 (ES +)
Example 9
4-r(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-methylpiperazin-1-
yl)propoxy]quinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoline-
carbonitrile (200 mg, 0.49 mmol), 3-(4-methyl-piperazin-1-yl)propanol (155 mg, 0.98
mmol) (WO 20047212) and sodium hydride (196 mg, 4.6 mmol) in 5 mL of N,N-
dimethylformamide was heated at 125°C for 3 hours. The reaction mixture was
poured into saturated sodium bicarbonate and stirred for 1 hour. The aqueous
solution was extracted with 10% methanol in dichloromethane. The organic layer was
washed with brine, dried over magnesium sulfate and concentrated in vacuo. The
residue was purified by preparative thin layer chromatography, eluting with 15%
methanol in dichloromethane. Trituration with hexane provided 116 mg of 4-[(2,4-

dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-methylpiperazin-1-yl)propoxy]-
quinoline-3-carbonitrile as a light brown solid, mp 137-138°C.
MS 542.0 (M-H)-
Analysis for C27H31l2N5O3 - 0.6 H2O
Calcd: C, 58.40; H, 5.84; N, 12.61.
Found: C, 58.31; H, 5.71; N, 12.43.
Example 10
4-[(2.4-Dichloro-5-methoxyphenyl)aminol- 6-ethoxy-7-[(1-niethylpiperidin-4-
yl)methoxy]quinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoline-
carbonitrile (200 mg, 0.49 mmol), 1-methylpiperidine-4-methanol- (188 mg, 0.98
mmol) (WO 20047212) and sodium hydride (196 mg, 4.6 mmol) in 5 mL of N,N-
dirnethylformamide was heated at 125°C for 3 hours. The reaction mixture was
poured into saturated sodium bicarbonate and stirred for 1 hour. The solid was
collected by filtration, washed with water and dried in vacuo. The solid was purified
by preparative thin layer chromatography, eluting with 15% methanol in
dichloromethane. Trituration with diethyl ether provided 67 mg of 4-[(2,4-dichloro-5-
methoxyphenyl)amino]-6-ethoxy-7-[(1-methylpiperidin-4-yl)methoxy]quino!ine-3-
carbonitrile as a light brown solid, mp 182-186°C.
MS 513.0 (M-H)-
Analysis for C26H28Cl2N4O3 -1.4 H2O
Calcd: C, 57.76; H, 5.74; N, 10.36.
Found: C, 57.65; H, 5.43; N, 10.15.
Example 11
4-[(2,4-Dichloro-5-methoxyphenyl)aminol-6-ethoxy-7-[3-(4-ethylpiperazin-1-
yl)propoxy]quinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoline-
carbonitrile (200 mg, 0.49 mmol) and 3-(4-ethyl-piperazin-1-yl)propanol (241 mg,
0.98 mmol) in 5 mL of N, N-dimethylformamide was heated at 125°C for 5 min.
Sodium hydride (60%) (98 mg, 2.45 mmol) was added and the mixture was heated
at 125°C for 1 hour. Additional sodium hydride (98 mg, 2.45 mmol) was added and

the mixture was heated at 125°C for 2 hours. The reaction mixture was cooled to
room temperature and poured into saturated sodium bicarbonate and stirred for 1
hour. The aqueous solution was extracted with 10% methanol in dichloromethane.
The organic layer was dried over sodium sulfate and concentrated in vacuo. The
residue was purified by preparative thin layer chromatography, developing with 12%
methanol in dichloromethane. Trituration with diethyl ether and hexane provided 146
mg of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-ethylpiperazin-1 -y|)-
propoxy]quinoline-3-carbonitrile as a light brown solid, mp 127-130°C.
MS 558.3 (M+H)+
Analysis for C28H33Cl2N5O3 - 1.5 H2O
Calcd: C, 57.44; H, 6.20; N, 11.96.
Found: C, 57.44; H, 6.24; N, 11.79.
Example 12
4-[(2,4-Dichloro-5-methoxyphenyl)aminol-6-ethoxy-7-[3-(1-methylpiperidin-4-
yl)propoxy]quinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoline-
carbonitrile (200 mg, 0.49 mmol) and 3-(1-methyl-4-piperidinyl)propanol (154 mg,
0.98 mmol) ) in 5 mL of N, N-dimethylformarnide was heated at 125°C for 5 min.
Sodium hydride (60%) (98 mg, 2.45 mmol) was added and the mixture was heated
at 125°C for 1 hour. Additional sodium hydride (98 mg, 2.45 mmol) was added and
the mixture was heated at 125°C for 2 hours. The reaction mixture was cooled to
room temperature and poured into saturated sodium bicarbonate and stirred for 1
hour. The precipitate was collected, washed with water and dried in vacuo. The
residue was purified by preparative thin layer chromatography, developing with 15%
methanol in dichloromethane. Trituration with diethyl ether provided 146 mg of 4-
[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(1-methylpiperidin-4-
yl)propoxy]quinoline-3-carbonitrile as an off-white solid, mp 148-151°C.
MS 543.2 (M+H)+
Analysis for C28H32Cl2N4O3 - 1.8 H2O
Calcd: C, 58.39; H, 6.23; N, 9.73.
Found: C, 58.40; H, 6.16; N, 9.64.

Example 13
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(4-methvl-1-
piperazinyl)ethoxy]quinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoIine-
carbonitrile (200 mg, 0.49 mmoi) and 2-(4-methyl-1-piperazinyl)ethanol (141 mg, 0.98
mmol)) in 5 mL of N, N-dimethylformamide was heated to 100°C. Sodium hydride
(60%) (196 mg, 4.9 mmol) was added in portions and the mixture was heated at
125ºC for 3 hours. The reaction mixture was cooled to room temperature and treated
with 25 mL of water. The mixture was stirred for 2 hours. The precipitate was
collected, washed with water and dried in vacuo. The residue was purified by flash
column chromatography, eluting with a gradient of 5% methanol in dichloromethane
to 15% methanol in dichloromethane. Trituration with diethyl ether provided 123 mg
of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(4-methyl-1-piperazinyl)-
ethoxy]quinoline-3-carbonitrile as an off-white solid, mp 141-143°C.
MS 530.2 (M+H)+
Analysis for C26H29CI2N5O3
Calcd: C, 58.87; H, 5.51; N, 13.20.
Found: C, 58.48; H, 5.45; N, 12.95.
Example 14
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(1-methylpiperidin-4-
yl)ethoxylquinoline-3-carbonitrile
A mixture of 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[luoro-3-quinoline-
carbonitrile (200 mg, 0.49 mmol) and 1-methyl-4-piperidinethanol (140 mg, 0.98
mmol) ) in 5 mL of N, N-dimethylformamide was heated to 100°C. Sodium hydride
(60%) (162 mg, 4.05 mmol) was added in portions and the mixture was heated at
125°C for 3 hours. The reaction mixture was cooled to room temperature and treated
with 25 mL of water. The precipitate was collected, washed with water and dried in
vacuo. The residue was purified by flash column chromatography, eluting with first
dichloromethane then a gradient of 5% methanol in dichloromethane to 30%
methanol in dichloromethane. Trituration with diethyl ether provided 121 mg of 4-
[(2,4-dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(1-methylpiperidin-4-yl)ethoxy]-
quinoline-3-carbonitrile as an off-white solid, mp 174-176°C.

MS 529.1 (M+H)+
Analysis for C27H30CI2N4O3
Calcd: C, 61.25; H, 5.71; N, 10.58.
Found: C, 61.40; H, 5.84; N, 10.35.
Example 15
4-[(2,4-Dichloro-5-methoxyphenyl)anninol-6-methoxy-7-[3-(4-propvl-1-
piperazinyl)propoxy]-3-quinolinecarbonitrile
Prepared according to the method used for the preparation of Example 1 by the
reaction of 7-[3-chtoroethoxy]-4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-3-
quinolinecarbonitrile and N-propylpiperazine: mp 97-101°C; MS (ES) m/z 558.2,
560.2(M+1).


WE CLAIM:
1. A method of providing neuroprotection in a patient following a
cerebrovascular ischemic event comprising providing a therapeutically effective
amount of a compound of the formula
wherein:
X is N, CH
n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutically
acceptable salts thereof; with the proviso that when n is 1, X is not N.
2. A method of inhibiting neurological deficits in a patient following a
cerebrovascular ischemic event comprising providing a therapeutically effective
amount of a compound of the formula
wherein:
X is N, CH
n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutically
acceptable salts thereof; with the proviso that when n is 1, X is not N.

3. A method of reducing infarct volumes in a patient following a cerebrovascular
ischemic event comprising administering a therapeutically effective amount of a
compound of the formula
wherein:
X is N, CH
n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutically
acceptable salts thereof; with the proviso that when n is 1, X is not N.
4. A method of inhibiting post-ischemic vascular permeability of cerebral blood
vessels in a patient suffering from a cerebrovascular event comprising administering
a therapeutically effective amount of a compound of the formula

wherein:
X is N, CH
n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutically
acceptable salts thereof; with the proviso that when n is 1, X is not N.
5. The method of any one of claims 1 to 4 wherein R' is methyl.

6. The method of any one of claims 1 to 5 wherein R is methyl or ethyl.
7. The method of any one of claims 1 to 6 wherein X is N.
8. The method of any one of claims 1 to 6 wherein X is CH.
9. The method of any one of claims 1 to 4 wherein the compound is:
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(4-methyl-1-piperazinyl)-
propoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 7-[3-(4-ethyl-1-piperazinyl)propoxy]- 6-
methoxy-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[2-(4-methyl-1-piperazinyl)-
ethoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-rnethoxyphenyl)amino]- 7-[2-(4-ethyl-1-piperazinyl)ethoxy]-6-
methoxy-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 6-methoxy-7-[(1-methylpiperidin-4-yl)-
methoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]- 6-methoxy-7-[2-(1-methylpiperidin-4-yl)-
ethbxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-7-[(1-ethylpiperidin-4-yl)methoxy]-6-
methoxyquinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-methylpiperazin-1-yl)-
propoxylquinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[(1-methylpiperidin-4-yl)-
methoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(4-ethylpiperazin-1-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(4-methyl-1-piperazinyl)-
ethoxy]quinoline-3-carbonitrile;

4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(1-methylpiperidin-4-yl)-
ethoxy]quinoline-3-carbonitrile; or
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(4-propyl-1-piperazinyl)-
propoxy]-3-quinolinecarbonitrile;
and pharmaceutically acceptable salts thereof.
10. The method of any one of claims 1 to 9 wherein compound is administered
between about 6 to about 24 hours after the ischemic event.
11. The method of any one of claims 1 to 10 wherein the therapeutically effective
amount is from about 1 mg/kg to about 30 mg/kg.
12. The method of any one of claims 1 to 11 comprising administering compound
of Formula I intravenously.
13. The method of any one of claims 1 to 12 wherein the patient is a human.
14. The method of any one of claims 1 to 13 wherein the ischemic event is
transient.

15. The method of any one of claims 1 to 13 wherein the ischemic event is acute.
16. The method of any one of claims 1 to 15 wherein the ischemic event is
stroke, head trauma, spinal trauma, general anoxia, or hypoxia.
17. The method of any one of claims 1 to 15 wherein the ischemic event occurs
during cranial hemmorhage, perinatal asphyxia, cardiac arrest or status epilepticus.

18.
wherein:
n is an integer from 1-3; and
R' and R are independently, alkyl of 1 to 3 carbon atoms, and pharmaceutical^
acceptable salts thereof.
19. A compound of Claim 18 wherein R' is methyl.
20. A compound of Claim 18 or Claim 19 wherein R is methyl or ethyl.
21. A compound which is:
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-methoxy-7-(1-methylpiperidin-4-yl)-
methoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxypheny!)amino]- 6-methoxy-7-[2-(1-methylpiperidin-4-yl)-
ethoxy]-3-quinolinecarbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyI)amino]-6-methoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-7-[(1-ethylpiperidin-4-yl)methoxy]-6-
methoxyquinoline-3-carbonitrile;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[(1-methylpiperidin-4-yl)-
methoxy]quinoline-3-carbonitriIe;
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[3-(1-methylpiperidin-4-yl)-
propoxy]quinoline-3-carbonitrile; or
4-[(2,4-Dichloro-5-methoxyphenyl)amino]-6-ethoxy-7-[2-(1-methylpiperidin-4-yl)-
ethoxy]quinoline-3-carbonitrile;
and pharmaceutically acceptable salts thereof.

22. A pharmaceutical composition comprising a compound as defined in any one
of claims 18 to 21 and a pharmaceutically acceptable carrier or excipient.
23. A pharmaceutical composition comprising a vascular permeability inhibiting
amount of a compound as defined in any one of claims 1 to 9 and a pharmaceutical
carrier or excipient.
24. The composition as claimed in Claim 21 in an intravenous dosage form.-
25. A process for the preparation of a compound as claimed in any one of claims
18 to 21 comprising one of the following:

wherein R'is as defined in Claim 18,
with an alcohol of formula

wherein R and n are as defined in Claim 18,
or (b) reacting a quinoline of formula

wherein R, R and n are as defined in Claim 18 and Y is a halogen,
with an aniline of formula


or

wherein R, R' and n are as defined in Claim 18, to give the desired quinoline.
26. Use of a compound as defined in any one of claims 1 to 9 in the preparation
of a medicament for the provision of neuroprotection in a patient following a
cerebrovascular ischemic event, the inhibition of neurological deficits in a patient
following a cerebrovascular ischemic event, the reduction of infarct volumes in a
patient following a cerebrovascular ischemic event or the inhibition of post-ischemic
vascular permeability of cerebral blood vessels in a patient suffering from a
cerebrovascular event.

Compounds of the Formula (I), wherein X is N, CH n
is an integer from 1-3; and R' and R are independently, alkyl of 1 to 3
carbon atoms, and pharmaceutically acceptable salts thereof, with the
proviso that when n is 1, X is not N; are useful for inhibiting vascular
permeability caused by disease, injury, or other trauma.