Note: Descriptions are shown in the official language in which they were submitted.
CA 02568609 2006-11-28
HALOETHYL UREA COMPOUNDS AND THEIR USE TO ATTENUATE, INHIBIT OR
PREVENT NON-CANCEROUS PATHOGENIC CELLULAR PROLIFERATION AND
DISEASES ASSOCIATED THEREWITH
FIELD OF INVENTION
The present invention pertains to the field of therapies for non-cancerous
pathogenic cellular
-proliferation. In particular the invention pertains to therapeutically active
halo urea derivatives
and their use to attenuate, inhibit or prevent non-cancerous cellular
proliferation and diseases and
disorders associated therewith.
BACKGROUND OF THE INVENTION
Non-cancerous pathogenic cellular proliferation has wide-ranging clinical
implications. Such
proliferation can be due to an abnormal cell proliferation or
hyperproliferation, which is defined
by an abnormally high rate of cell division, resulting in a rapid
proliferation of the cells. Non-
cancerous hyperproliferating cells typically resemble normal cells and
function like normal cells:
In contrast, malignant hyperproliferating cells are typically anaplastic and
are capable of
invasion and metastasis.
The pathogenesis of diseases involving non-cancerous cellular proliferation
often include cellular
migration and inflammation. For example, angiogenesis associated with disease
states, involves
migration of endothelial cells in addition to endothelial cell proliferation.
Moreover,
irmnunological mechanisms play a role in the pathogenesis of a number of
diseases associated
with cellular proliferation and, therefore, such disease states are associated
with inflammation.
For example, it has been suggested that immunologic mechanisms play a role in
the pathogenesis
of psoriasis. Specifically, it has been suggested that cytokines produced
during immune
activation and other inflammatory processes may lead to epidermal hyperplasia
observed in
psoriasis (Gottlieb J Invest Dermatol. 95(5):1SS-19S (1990)).
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A number of diseases and disorders are associated with such non-cancerous
pathogenic cellular
proliferation including, but not limited to, psoriasis, blood vessel
proliferative disorders, fibrotic
disorders, arteriosclerotic disorders, restenosis, neointimal hyperplasia,
endometriosis and
lymphoproliferative disorders.
One example of non-cancerous pathogenic cellular proliferation is psoriasis,
which is a common
skin disease resulting in thickening of the epidermis and the presence of red
scaly plaques.
Hyperproliferation of keratinocytes is a key feature of psoriasis along with
epidermal
inflammation and reduced differentiation of keratinocytes. Multiple mechanisms
have been
invoked to explain the keratinocyte hyperproliferation that characterizes
psoriasis, but no single
mechanism has been definitively implicated. Activation of epidermal growth
factor receptors,
alterations in protein kinase C signal transduction pathways, and the
attendant changes in
intracellular calcium metabolism may play a role in psoriatic epidermal
hyperplasia. Disordered
cellular immunity has also been implicated in the pathogenesis of psoriasis.
However, the exact
mechanisms of keratinocyte hyperproliferation and epidermal inflammation
remain unclear and,
because of the multifactorial nature of psoriasis, it is difficult to predict
whether pharmacologic
manipulation of complex signal transduction pathways, growth factor receptors,
or cellular
immune functions will attenuate the hyperproliferation of keratinocytes.
Topical treatments, either alone or as an adjunct to other therapies for those
patients with
moderate to severe psoriasis, are widely used in the treatment of psoriasis.
Topical therapies
include, for example, coal tar preparation (1-5% by weight), topical steroids,
anthralin cream
(1%) and synthetic retinoid tazarotene. Various side-effects are associated
with the use of such
therapies, for example, coal tar has a bad odour and stains clothing; long
term use of fluorinated
corticosteroids (which are more effective than hydrocortisone) may lead to
striae, telangiectasis
and ecchythmosis, and anthralin cream or synthetic retinoid tazarotene are
often irritating. Other
topical agents such as calcipotriene and vitamin D analogues (vitamin D3 or
calcipotriol) may
also provide temporary relief, while keratolytics such as salicylic acid can
help in removing the
thick scales from the psoriatic plaques.
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Natural sunlight may be beneficial for treatment of psoriasis, and this has
led to the use of UV
radiation therapy (see, for example, U.S. Patent Nos. 4,153,572 and
4,558,700). UVB radiation
(280-320 nm) of affected areas is one of the most common treatments for
moderate to severe
psoriasis, with its efficacy enhanced by coating the skin with a tar
containing emollient prior to
the radiation.
The use of oral corticosteroids, which have an immunosuppressive effect on
cytotoxic T
lymphocytes, is also common. Other frequently used oral therapeutic agents
include
methotrexate, cyclosporin, hydroxyurea and acitretin. As with topical
treatments, various side-
effects are associated with prolonged use of these compounds, for example,
methotrexate can
lead to leulcopenia and cumulative hepatic toxicity, cyclosporin may result in
hypertension and
nephrotoxicity, hydroxyurea use is limited by hematologic side effects and
patients using
acitretin may experience extreme dryness of mucous membranes, an increase in
arthralgias and
increased blood triglycerides.
A second example of non-cancerous pathogenic cellular proliferation are
fibrotic disorders.
Fibrotic disorders result in whole or in part from the proliferation of
fibroblasts. Fibrotic
disorders include but are not limited to fibrosis, cirrhosis and
arteriosclerotic conditions.
Another example of non-cancerous pathogenic cellular proliferation is
unregulated angiogenesis,
and/or angiogenesis involved in the pathogenesis of disease. Under normal
physiological
conditions, humans or animals undergo angiogenesis only in very specific
restricted situations,
for example, during wound healing, fetal and embryonal development and
formation of the
corpus luteum, endometrium and placenta. Persistent, unregulated angiogenesis,
however,
occurs in a multiplicity of disease states and supports the pathological
damage seen in these
disease states.
For example, while cornea and cartilage are avascular in healthy situations,
several diseases
involving these tissues are complicated by the massive arrival of new blood
vessels. Eye
angiogenic diseases include neovascular glaucoma, retrolental fibroplasia,
macular degeneration
and neovascularization of corneal grafts. Joint angiogenic diseases include
rheumatoid arthritis
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and arthrosis. Psoriasis also exhibits hypervascularization at the surface of
the skin. Solid
tumour growth is also critically dependent upon the formation of new blood
vessels to progress
locally and spread throughout the body. Regulation of cell replication and/or
differentiation is
important in the maintenance of existing blood vessels, for example, acute and
chronic
pathological processes, such as atherosclerosis, post-angioplastic restenosis
and hypertension,
involve the proliferation of different cellular components of mature blood
vessels (endothelial
cells, smooth muscle cells, myocytes and fibroblasts). '
Several kinds of compounds have been proposed for the prevention or inhibition
of angiogenesis
involved in disease states, for example, protamine (see Taylor et al., (1982)
Nature 297:307),
heparin (see Folkman et al., Science 221:719 (1983) and U.S. Patent Nos.
5,001,116 and
4,994,443) and certain steroids, such as tetrahydrocortisol, which lack gluco
and mineral
corticoid activity. The toxicity of protamine, however, limits its practical
use as a therapeutic.
Other factors found endogenously in animals, such as a 4 kDa glycoprotein from
bovine vitreous
humour and a cartilage derived factor, have been used to inhibit angiogenesis.
Cellular factors
such as interferon inhibit angiogenesis. For example, interferon-a or human
interferon-(3 has
been shown to inhibit tumour-induced angiogenesis in mouse dermis stimulated
by human
neoplastic cells. Interferon-(3 is also a potent inhibitor of angiogenesis
induced by allogeneic
spleen cells (Sidlcy et al., (1987) Cancer Research 47:5155-5161). The use of
human
recombinant interferon-a (alpha/A) in the treatment of pulmonary
hemangiomatosis, an
angiogenesis-induced disease, has been reported (White et al., (1989) New
England J. Med.
320:1197-1200).
Urea-based compounds have been described for diverse indications, including as
herbicides
(U.S. Patent No. 3,885,954), as prophylactics against gastrointestinal and
cardiovascular
disorders (U.S. Patent No. 4,707,478), as anti-parasitic agents (IJ.S. Patent
No. 4,707,478), as
anti-athersclerotic agents (I1.S. Patent No. 4,623,662), as treatments for
gastrointestinal,
spasmolytic and ulcerogeiuc disorders (U.S. Patent No. 4,304,786) and as anti-
cancer agents (for
example, U.S. Patent Nos. 3,968,249; 4,973,675 and 4,803,223). A class of 1-
aryl-3-(2-
chloroethyl)urea derivatives have been described as anti-cancer agents (U.S.
Patent Nos.
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5,530,026 and 5,750,547, and International Patent Application WO 00/61546) and
as (3-tubulin
inhibitors (International Patent Application WO 01/447504).
This background information is provided for the purpose of making known
information believed
by the applicant to be of possible relevance to the present invention. No
admission is necessarily
intended, nor should be construed, that any of the preceding information
constitutes prior art
against the present invention.
SUMMARY OF THE INVENTION
An object of the present invention is to provide haloethyl urea compounds and
their use as anti-
proliferative agent in the attenuation, inhibition, or prevention of non-
cancerous cellular
proliferation. In accordance with one aspect of the present invention there is
provided a method
of attenuating, inhibiting, or preventing non-cancerous cellular proliferation
comprising
contacting cells with an effective amount of a compound having structural
formula (I):
R~2
B-N N CH2X I
H H
or a pharmaceutically acceptable salt thereof, wherein:
XisF,Cl,BrorI;
R1 and R2 are each independently selected from the group of H, -R, -halo, -OR,
-SR, -NRR, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -C(O)NRR,
-C(S)NRR, -C(O)NR(SR), -C(S)NR(SR), -CH(CN)2, -CH[C(O)R]2, -CH[C(S)R]2,
-CH[C(O)ORJ2, -CH[C(S)OR]2, -CH[C(O)SR]2, -CH[C(S)SR]a, -NRC(O)R, -NRC(O)OR,
or Rl
and R2 when taken together form =O, =S or a C3-C6 spiro group;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine;
wherein:
CA 02568609 2006-11-28
B is substituted with one or more substituents selected from the group of (CI-
Cls) alkyl,
(Ca-Cis) alkenyl, (C2-C16) alkynyl, aryl, -O-(C1-C16) alkyl, -O-(C2-C16)
alkenyl, -O-(C2-Cls)
alkynyl, -O-aryl, -O-CH2-aryl, -S-(C1-C16)alkyl, -S-(C2-C16) alkenyl, -S-(C2-
C16) alkynyl, -S-
aryl, -S-CHZ-aryl, (C3-C8) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8)
cycloalkyl, -halo, -NRR,
-ONRR -N02, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -
SC(O)R, -SC(S)R, -OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -
C(O)NR(SR), -C(S)NR(SR), -CH(CN)2, -CH[C(O)R]2, -CH[C(S)R]Z, -CH[C(O)OR]2, -
CH[C(S)OR]2, -CH[C(O)SR]Z, -CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -
S(O)ZOR, -S(O)NRR, -S(O)ONRR;
wherein:
each R is independently selected from -H, (CI-C16) alkyl, substituted (Ci-C16)
alkyl, (C2-
C16) alkenyl, substituted (CZ-C16) alkenyl, (CZ-C16) alkynyl, substituted (CZ-
C16) alkynyl, (C3-C8)
cycloalkyl, substituted (C3-C$) cycloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl are optionally substituted with one or
more substituents
independently selected from the group of -halo, trihalomethyl, -R', -OR', -
SR', -NR'R', -NOz, -
CN, -OC(O)R', -OC(S)R', -SC(O)R, -SC(S)R -C(O)R', -C(S)R', -C(O)OR', -C(S)OR',
-C(O)SR',
-C(S)SR', -C(O)NR'R', -C(S)NR'R', -NR'C(O)R' and -NR'C(O)OR ;
the cycloalkyl is optionally substituted with one or more substituents
independently selected
from the group of R', -halo, OR', -SR', -NRR', -ONRR', -NO2, -CN, -C(O)R', -
C(S)R', -
OC(O)R', -SC(O)R', -SC(S)R', -OC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR',
-
C(O)NRR', -C(S)NRR', -C(O)NR'(OR7, -C(S)NR' -(OR7, -C(O)NR'(SR7, -C(S)NR'(SR~,
-
CH(CN)2, -CH[C(O)R72, -CH[C(S)R~2, -CH[C(O)OR']2, -CH[C(S)OR']2, -
CH[C(O)SR']2, -
CH[C(S)SR~a, -NR'C(O)R', -NR'C(O)OR', -S(O)-R', -S(O)OR', -S(O)zOR', -
S(O)NRR', -
S(O)ONRR', and
each R' is independently selected from the group of -H, (C1-C16) alkyl,
substituted (C1-Cls)
alkyl, (C~-C16) alkenyl, substituted (CZ-C16) alkenyl, (C2-C16) alkynyl,
substituted (C2-C16)
alkynyl, (C3-C8) cycloalkyl, substituted (C3-C8) cyloallcyl, aryl or
substituted aryl.
In accordance witli~another aspect of the invention compounds of formula (I)
are provided for
use as anti-proliferative agent in the attenuation, inhibition, or prevention
of non-cancerous
cellular proliferation.for use as a therapeutic agent in the treatment of a
disease or disorder,
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CA 02568609 2006-11-28
wherein pathogenesis of said disease or disorder is associated with non-
cancerous pathogenic
cellular proliferation.
BRIEF DESCRIPTION OF THE FIGURES
Figure 1 presents a graph illustrating the ability of compound 1 to inhibit
human foreskin
fibroblast cell proliferation (* Number of untreated cells were arbitrary
assign to 100%).
Figure 2 presents a graph illustrating the ability of compound 1 to inhibit
human umbilical vein
endothelial cell (HLJVEC) proliferation (* Number of untreated cells were
arbitrary assign to
100%).
Figure 3 presents graphs illustrating the ability of compound 1 (A); compound
5 (B); 1-(4-t-
butyl-phenyl)-3-ethyl urea (tBEU) (C) aand cDDP to inhibit human umbilical
vein endothelial
cell (HUVEC) proliferation.
Figure 4 presents a graph illustrating the ability of compounds 1, 2 and 3 to
affect the inhibition
of IL-2 production of PMA-stimulated Jurkat cells. The graph further
illustrates that the
compounds do not stimulate IL-2 production in non-stimulated (control) Jurkat
cells.
Figures SA and SB present graphs illustrating the ability of compounds 1, 2, 3
and 88 to affect
the inhibition of IL-2 production of PMA-stimulated Jurkat cells. The graphs
further illustrate
that the compounds do not stimulate IL-2 production in non-stimulated
(control) Jurkat cells.
Figure 6a-c present graphs illustrating the ability of compounds of the
invention to inhibit IL-2
production of PMA/PHA stimulated human lympho-mono cells and the cytotoxicity
of the
compounds in PMA/PHA stimulated lympho-mono cells.
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Figure 7 presents a graph illustrating the ability of compound 1 to inhibit
growth of
immortalized human keratinocytes (HaCat cells) (*Number of untreated cells
were arbitrary
assign to 100% growth).
Figures 8A-H present graphs illustrating the effect of various compounds of
the invention and
control compounds on immortalized human keratinocytes (HaGat cells). 8A
compound 1; 8B
compound 2; 8C compound 3; 8D compound 17; 8E 5-FU; 8F colchicine; 8G taxol,
and 8H
vinblastine.
Figure 9 presents graphs illustrating the cytotoxic activity of compounds 1
(9A), 2 (9B), 3 (9C),
22 (9D) and tB-CEU (9E) on HaCat cells.
Figure 10 presents the results of a Matrigel plug assay using compound 5.
Figure 11 presents the results of a Matrigel plug assay using compound 1,
compound 1,
compound 5 or tBEU.
Figure 12 presents graphs illustrating the effects of compound 1 (A) and
compound 5 (B) on cell
migration.
Figure 13 presents graphs illustrating the effects of compounds 1 (A); 5 (B)
and tBEU (C) on
endothelial cell migration.
Figure 14 presents a histogram depicting the effect of compound 1 on tumour
growth in the
chick CAM assay.
Figure 15 presents graphs depicting the effect of compounds 1 (A); 5 (B); tBEU
(C) and cDDP
(D) of the invention on tumour growth in the CAM assay.
Figure 16 presents graphs depicting the effect of compounds 1 (A); 5 (B); 2
(C) and 22 (D) of
the invention on tumour growth in the CAM assay.
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Figure 17 depicts the microtubule depolymerization and cytoskeleton disruption
in HUVECs
induced by compounds of the invention.
DETAILED DESCRIPTION OF THE INVENTION
The invention provides for the use of haloethyl urea derivatives of Formula I
for the attenuation,
inhibition or prevention of non-cancerous pathogenic cellular proliferation
and diseases
associated therewith.
Defitzitioias
The terms and abbreviations used in the instant examples have their normal
meanings unless
otherwise designated. For example "°C" refers to degrees Celsius; "N"
refers to normal or
normality; "mmol" and "mM" refer to millimole or millimoles; "~.M" refers to
micromole or
micromoles; "g" refers to gram or grams; "mL" means milliliter or milliliters;
"M" refers to
molar or molarity; "p-" refers to para, "MS" refers to mass spectrometry; "IR"
refers to infrared
spectroscopy; and "NMR" refers to nuclear magnetic resonance spectroscopy.
The terms are defined as follows:
The term "halogen" refers to fluorine, bromine, chlorine, and iodine atoms.
The term "hydroxyl" refers to the group -OH.
The term "thiol" or "mercapto" refers to the group -SH, and -S(O)o_2.
The term "lower alkyl" refers to a straight chain or branched, or cyclic,
alkyl group of 1 to 16
carbon atoms. This term is further exemplified by such groups as methyl,
ethyl, n-propyl, i-
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propyl, ya-butyl, t-butyl, 1-butyl (or 2-methylpropyl), cyclopropylmethyl, i-
amyl, n-amyl, hexyl
and the like.
The term "substituted lower alkyl" refers to lower alkyl as just described
including one or more
groups such as hydroxyl, thiol, alkylthiol, halogen, alkoxy, amino, amido,
carboxyl, cycloalkyl,
substituted cycloalkyl, heterocycle, cycloheteroalkyl, substituted
cycloheteroalkyl, acyl,
carboxyl, aryl, substituted aryl, aryloxy, hetaryl, substituted hetaryl,
aralkyl, heteroaralkyl, alkyl
alkenyl, alkyl alkynyl, alkyl cycloalkyl, alkyl cycloheteroalkyl, cyano. These
groups may be
attached to any carbon atom of the lower alkyl moiety.
The term "lower alkenyl" refers to a straight chain or branched hydrocarbon of
2 to 16 caxbon
atoms having at least one carbon to carbon double bond.
The term "substituted lower alkenyl" refers to lower alkenyl as just described
including one or
more groups such as hydroxyl, thiol, alkylthiol, halogen, alkoxy, amino,
amido, carboxyl,
cycloalkyl, substituted cycloalkyl, heterocycle, cycloheteroalkyl, substituted
cycloheteroalkyl,
acyl, carboxyl, aryl, substituted aryl, aryloxy, hetaryl, substituted hetaryl,
aralkyl, heteroaralkyl,
alkyl, alkenyl, alkynyl, alkyl alkenyl, alkyl alkynyl, alkyl cycloalkyl, alkyl
cycloheteroalkyl,
cyano. These groups may be attached to any carbon atom to produce a stable
compound.
The term "lower alkynyl" refers to a straight chain or branched hydrocarbon of
2 to 16 carbon
atoms having at least one carbon to carbon triple bond.
The term "substituted lower alkynyl" refers to lower allcynyl as just
described including one or
more groups such as hydroxyl, thiol, alkylthiol, halogen, alkoxy, amino,
amido, carboxyl,
cycloalkyl, substituted cycloalkyl, heterocycle, cycloheteroalkyl, substituted
cycloheteroalkyl,
acyl, carboxyl, aryl, substituted aryl, aryloxy, hetaryl, substituted hetaryl,
aralkyl, heteroaralkyl,
alkyl, allcenyl, alkynyl, alkyl. alkenyl, alkyl alkynyl, alkyl cycloalkyl,
alkyl cycloheteroalkyl,
cyano. These groups may be attached to any carbon atom to produce a stable
compound.
to
CA 02568609 2006-11-28
The term "alkyl alkenyl" refers to a group -R-CR'=CR"'R"", where R is lower
alkyl, or
substituted lower alkyl, R', R"', R"" are each independently selected from
hydrogen, halogen,
lower alkyl, substituted lower alkyl, acyl, aryl, substituted aryl, hetaryl,
or substituted hetaryl as
defined below.
The term "alkyl alkynyl" refers to a group -R-C-_-CR' where R is lower alkyl
or substituted lower
alkyl, R' is hydrogen, lower alkyl, substituted lower alkyl, acyl, aryl,
substituted aryl, hetaryl, or
substituted hetaryl as defined below.
The term "alkoxy" refers to the group -OR, where R is lower alkyl, substituted
lower alkyl, acyl,
aryl, substituted aryl, aralkyl, substituted aralkyl, heteroalkyl,
heteroarylalkyl, cycloalkyl,
substituted cycloalkyl, cycloheteroalkyl, or substituted cycloheteroalkyl as
defined below.
The term "alkylthio" denotes the group -SR, -S(O)"_1-z -R, where R is lower
alkyl, substituted
lower alkyl, aryl, substituted aryl aralkyl or substituted aralkyl as defined
below.
The term "acyl" refers to groups -C(O)R, where R is hydrogen, lower alkyl
substituted lower
alkyl, aryl, substituted aryl.
The term "aryloxy" refers to groups -OAr, where Ar is an aryl, substituted
aryl, heteroaryl, or
substituted heteroaryl group as defined below.
The term "amino" refers to the group NRR', where R and R' may independently be
hydrogen,
lower alkyl, substituted lower alkyl, aryl, substituted aryl, hetaryl,
cycloalkyl, or substituted
hetaryl as defined below or acyl.
The term "amido" or "amide" refers to the group -C(O)NRR', where R and R' may
independently
be hydrogen, lower alkyl, substituted lower alkyl, aryl, substituted aryl,
hetaryl, substituted
hetaryl as defined below.
The term "carboxyl" refers to the group -C(O)OR, where R may independently be
hydrogen,
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lower alkyl, substituted lower alkyl, aryl, substituted aryl, hetaryl,
substituted hetaryl and the like
as defined.
The terms "aryl" or "Ar" refer to an aromatic carbocyclic group having at
least one aromatic ring
(e.g., phenyl or biphenyl) or multiple condensed rings in which at least one
ring is aromatic,
(e.g., 1,2,3,4-tetrahydronaphthyl, naphthyl, anthryl, or phenanthryl, 9-
fluorenyl etc.).
The term "substituted aryl" refers to aryl optionally substituted with one or
more functional
groups, e.g., halogen, hydroxyl, thiol, lower alkyl, substituted lower alkyl,
trifluoromethyl,
alkenyl, alkenyl, alkylalkenyl, alkyl alkynyl, alkoxy, alkylthio, acyl,
aryloxy, amino, amido,
carboxyl, aryl, substituted aryl, heterocycle, heteroaryl, substituted
heterocycle, heteroalkyl,
cycloalkyl, substituted cycloalkyl, alkylcycloallcyl, alkylcycloheteroalkyl,
nitro, sulfamido or
cyano.
The term "heterocycle" refers to a saturated, unsaturated, or aromatic
carbocyclic group having a
single ring (e.g., morpholino, pyridyl or furyl) or multiple condensed rings
(e.g., naphthpyridyl,
quinoxalyl, quinolinyl, indolizinyl, indanyl or benzo[b)thienyl) and having at
least one hetero
atom, such as N, O or S, within the ring.
The term "substituted heterocycle" refers to heterocycle optionally
substituted with, halogen,
hydroxyl, thiol, lower alkyl, substituted lower alkyl, trifluoromethyl,
alkenyl, alkenyl,
alkylalkenyl, alkyl allcynyl, alkoxy, alkylthio, acyl, aryloxy, amino, amido,
carboxyl, aryl,
substituted aryl, heterocycle, heteroaryl, substituted heterocycle,
heteroalkyl, cycloalkyl,
substituted cycloalkyl, alkylcycloalkyl, alkylcycloheteroalkyl, nitro,
sulfamido or cyano and the
like.
The terms "heteroaryl" or "hetar" refer to a heterocycle in which at least one
heterocyclic ring is
aromatic.
The term "substituted heteroaryl" refers to a heterocycle optionally mono or
poly substituted
with one or more functional groups, e.g., halogen, hydroxyl, thiol, lower
alkyl, substituted lower
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alkyl, trifluoromethyl, alkenyl, alkenyl, alkylalkenyl, alkyl alkynyl, alkoxy,
alkylthio, acyl,
aryloxy, amino, amido, carboxyl, aryl, substituted aryl, heterocycle,
heteroaryl, substituted
heterocycle, heteroalkyl, cycloalkyl, substituted cycloalkyl, alkylcycloalkyl,
alkylcycloheteroalkyl, vitro, sulfamido or cyano and the like.
The term "aralkyl" refers to the group -R-Ar where Ar is an aryl group and R
is lower alkyl or
substituted lower alkyl group. Aryl groups can optionally be unsubstituted or
substituted with,
e.g., halogen, lower alkyl, alkoxy, alkyl thio, trifluoromethyl, amino, amido,
carboxyl, hydroxyl,
aryl, aryloxy, heterocycle, hetaryl, substituted hetaryl, vitro, cyano,
alkylthio, thiol, sulfamido
and the like.
The term "heteroalkyl" refers to the group -R-Het where Het is a heterocycle
group and R is a
lower alkyl group. Heteroalkyl groups can optionally be tmsubstituted or
substituted with e.g.,
halogen, lower alkyl, lower alkoxy, lower alkylthio, trifluoromethyl, amino,
amido, carboxyl,
hydroxyl, aryl, aryloxy, heterocycle, hetaryl, substituted hetaryl, vitro,
cyano, alkylthio, thiol,
sulfamido and the like.
The term "heteroarylalkyl" refers to the group -R-HetAr where HetAr is an
heteroaryl group and
R lower alkyl or substituted loweralkyl. Heteroarylalkyl groups can optionally
be unsubstituted
or substituted with, e.g., halogen, lower alkyl, substituted lower alkyl,
alkoxy, alkylthio, aryl,
aryloxy, heterocycle, hetaryl, substituted hetaryl, vitro, cyano, alkylthio,
thiol, sulfamido and the
like.
The term "cycloalkyl" refers to a cyclic or polycyclic alkyl group containing
3 to 15 carbon. For
polycyclic groups, these may be multiple condensed rings in which one of the
distal rings may be
aromatic (e.g. tetrahydronaphthalene, etc.).
The term "substituted cycloalkyl" refers to a cycloalkyl group comprising one
or more
substituents with, e.g halogen, hydroxyl, thiol, lower alkyl, substituted
lower alkyl,
trifluoromethyl, alkenyl, alkenyl, alkylallcenyl, alkyl alkynyl, alkoxy,
alkylthio, acyl, aryloxy,
amino, amido, carboxyl, aryl, substituted aryl, heterocycle, heteroaryl,
substituted heterocycle,
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heteroalkyl, cycloalkyl, substituted cycloalkyl, alkylcycloalkyl,
alkylcycloheteroalkyl, vitro,
sulfamido or cyano and the like.
The term "cycloheteroalkyl" refers to a cycloalkyl group wherein one or more
of the ring carbon
atoms is replaced with a heteroatom (e.g., N, O, S or P).
The term "substituted cycloheteroalkyl" refers to a cycloheteroalkyl group as
herein defined
which contains one or more substituents, such as halogen, lower alkyl, lower
alkoxy, lower
alkylthio, trifluoromethyl, amino, amido, carboxyl, hydroxyl, aryl, aryloxy,
heterocycle, hetaryl,
substituted hetaryl, vitro, cyano, alkylthio, thiol, sulfamido and the like.
The term "alkyl cycloalkyl" refers to the group -R-cycloalkyl where cycloalkyl
is a cycloalkyl
group and R is a lower alkyl or substituted lower alkyl. Cycloalkyl groups can
optionally be
unsubstituted or substituted with e.g. halogen, lower alkyl, lower alkoxy,
lower alkylthio,
trifluoromethyl, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle,
hetaryl,
substituted hetaryl, vitro, cyano, alkylthio, thiol, sulfamido and the like.
The terms "treatment," "therapy" and the like refer to improvement in the
recipient's status as
well as prophylaxis. The improvement can be subjective or objective and
related to features such
as symptoms or signs of the disease or condition being treated. Prevention of
deterioration of the
recipient's status is also encompassed by the term.
The term "non-cancerous pathogenic cellular proliferation" refers to excessive
or abnormal cell
proliferation or hyperproliferation. "Non-cancerous pathogenic cellular
proliferation" may
include proliferation of a wide variety of cell types, including but not
limited to keratinocytes
and endothelial cells. "Non-cancerous pathogenic cellular proliferation"
includes unregulated
angiogenesis which may or may not be associated with cancer.
The term "inhibit" or "inhibition" as it refers to non-cancerous pathogenic
cellular proliferation
includes the arrest, prevention or decrease in non-cancerous pathogenic
cellular proliferation,
both temporary and long-term.
14
CA 02568609 2006-11-28
The term "ameliorate" or "amelioration" includes the arrest, prevention,
decrease, or
improvement in one or more the symptoms, signs, and features of the disease
being treated, both
temporary and long-term.
The compounds according to the instant invention include compounds of the
following general
formula:
O R~ R2
I
B-N N CH2X
H H
or a pharmaceutically acceptable salt thereof, wherein:
XisF,Cl,BrorI;
R1 and R2 are each independently selected from the group of H, -R, -halo, -OR,
-SR, -NRR, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -C(O)NRR,
-C(S)NRR, -C(O)NR(SR), -C(S)NR(SR), -CH(CN)z, -CH[C(O)R]a, -CH[C(S)R]Z,
-CH[C(O)OR]2, -CH[C(S)OR]Z, -CH[C(O)SR]a, -CH[C(S)SR]Z, -NRC(O)R, -NRC(O)OR,
or Rl
and R2 when taken together form =O, =S or a C3-C6 spiro group;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine;
wherein:
B is substituted with one or more substituents selected from the group of (Ci-
C16) alkyl, (CZ-C16)
alkenyl, (C2-C16) alkynyl, aryl, -O-(C1-C16) alkyl, -O-(CZ-C16) alkenyl, -O-
(C2-C16) alkynyl, -O-
aryl, -O-CHZ-aryl, -S-(C1-C16)alkyl, -S-(CZ-C16) alkenyl, -S-(Cz-C16) alkynyl,
-S-aryl, -S-CH2_
aryl, (C3-C$) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8) cycloalkyl, -halo,
-NRR, -ONRR -
NOZ, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -
SC(O)R,
-SC(S)R, -OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR),
-C(S)NR(SR), -CH(CN)2, -CH[C(O)R]2, -CH[C(S)R]2, -CH[C(O)OR]Z, -CH[C(S)OR]Z,
-CH[C(O)SR]2, -CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)ZOR, -
S(O)NRR, -S(O)ONRR;
wherein:
CA 02568609 2006-11-28
each R is independently selected from -H, (C1-C16) alkyl, substituted (C1-C16)
alkyl, (Cz-
C16) alkenyl, substituted (Cz-C16) alkenyl, (Ca-C16) alkynyl, substituted (CZ-
C16) alkynyl, (C3-C8)
cycloalkyl, substituted (C3-C8) cycloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl are optionally substituted with one or
more substituents
independently selected from the group of -halo, trihalomethyl, -R', -OR', -
SR', -NR'R', -N02, -
CN, -OC(O)R', -OC(S)R', -SC(O)R, -SC(S)R -C(O)R', -C(S)R', -C(O)OR', -C(S)OR',
-C(O)SR',
-C(S)SR', -C(O)NR'R', -C(S)NR'R', -NR'C(O)R' and -NR'C(O)OR ;
the cycloalkyl is optionally substituted with one or more substituents
independently selected
from the group of R', -halo, OR', -SR', -NRR', -ONRR', -NOZ, -CN, -C(O)R', -
C(S)R', -
OC(O)R', -SC(O)R', -SC(S)R', -OC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR',
-
C(O)NRR', -C(S)NRR', -C(O)NR'(OR~, -C(S)NR' -(ORS, -C(O)NR'(SR~, -C(S)NR'(SR~,
-
CH(CN)2, -CH[C(O)R~2, -CH[C(S)R'JZ, -CH[C(O)OR']2, -CH[C(S)OR~2, -CH[C(O)SR72,
-
CH[C(S)SR']2, -NR'C(O)R', -NR'C(O)OR', -S(O)-R', -S(O)OR', -S(O)ZOR', -
S(O)NRR', -
S(O)ONRR', and
each R' is independently selected from the group of -H, (C1-C16) alkyl,
substituted (Cl-
C16) alkyl, (CZ-C1~) alkenyl, substituted (CZ-C16) alkenyl, (Ca-C16) alkynyl,
substituted (C2-C16)
alkynyl, (C3-Cg) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or
substituted aryl.
In one embodiment, the compound of the invention is one in which R1 and R2 are
each
independently selected from H, (C~-C6) alkyl and (C1-C6) alkoxy.
In a specific embodiment in the compound of formula (I) B is phenyl,
substituted with one or
more substituents indepnednetly selected from the list as shown above.
W another embodiment in the compound of formula (I), B is phenyl substituted
with halo, -CN,
-C(O)R, -C(O)OR, -OC(O)R, -C(O)NRR, -OR, (C1-C16) alkyl, (CZ-C16) alkenyl, (CZ-
Cls)
alkynyl, wherein said alkyl, alkenyl and alkynyl are optionally substituted
with -CN, -C(O)R', -
C(O)OR', -OC(O)R', -C(O)NR'R', -OR', wherein R and R' are as defined above.
In another embodiment the substituents of the compounds of formula (I) are as
follows:
XisF,Cl,BrorI;
16
CA 02568609 2006-11-28
R1 and R2 are as defined above, and
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine; and
substituted with at least one substituent selected from, (C1-C~6) alkyl, (C2-
C16) alkenyl, (Ca-C16)
alkynyl, -O-(Cl-C16) alkyl, -O-(C2-C16) alkenyl, -O-(CZ-C16) alkynyl, aryl,
substituted aryl, -O-
aryl, -O-CHZ-aryl, -S-(C1-C16) alkyl, -S-(CZ-C16) alkenyl, -S-(CZ-C16)
alkynyl, -S-aryl, -S-CHZ-
aryl, (C3-C8) cycloalkyl, -O-(C3-C$) cycloalkyl, -S-(C3-C8) cycloalkyl, -ONRR,
-C(O)R, -C(S)R
-C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -SC(S)R, -OC(S)R, -
C(O)NRR, -
C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -C(S)NR(SR), -CH(CN)2,
-CH[C(O)R]z, -CH[C(S)R]Z, -CH[C(O)OR]2, -CH[C(S)OR]Z, -CH[C(O)SR]a, -
CH[C(S)SR]2,
-NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)20R, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -NO2,
-NR'R', -O-alkyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R ;
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -NOZ, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R';
said -O-alkenyl, -O-alkynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-cycloalkyl
are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NOz, -
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above.
In another embodiment the substituents of the compounds of formula (I) are as
follows:
XisF,Cl,BrorI;
Rl and R2 are as defined above, and
B is phenyl substituted with at least one substituent selected from, (CI-C16)
alkyl, (C2-C16)
allcenyl, (C2-C16) alkynyl, -O-(C~-C16) alkyl, -O-(Cz-C16) alkenyl, -O-(CZ-
C16) alkynyl, aryl,
substituted aryl, -O-aryl, -O-CHZ-aryl, -S-(C1-C16) alkyl, -S-(Ca-C16)
alkenyl, -S-(C2-C16)
alkynyl, -S-aryl, -S-CH2-aryl, (C3-C$) cycloalkyl, -O-(C3-C8) cycloallcyl, -S-
(C3-C8) cycloalkyl, -
ONRR, -C(O)R, -C(S)R -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -
SC(S)R,
1~
CA 02568609 2006-11-28
-oc(s)R, -c(o)NRR, -c(s)NRR, -c(o)NR(oR), -c(s)NR(oR), -c(o)NR(sR), -
c(s)NR(sR), -
CH(CN)z, -CH[C(O)R]z, -CH[C(S)R]z, -CH[C(O)OR]z, -CH[C(S)OR]z, -CH[C(O)SR]z, -
CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)zOR, -S(O)NRR, -
S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -NOz,
-NR'R', -O-alkyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R';
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -NOz, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R ;
said -O-alkenyl, -O-alkynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-cycloalkyl
are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NOz, -
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above.
In another embodiment the substituents of the of the compounds of formula (I)
are as follows:
XisF,Cl,BrorI;
R1 and R2 are as defined above, and
B is phenyl substituted with at least one group selected from, (C1-C16) alkyl,
(Cz-C16) alkenyl,
(Cz-C16) alkynyl, -~-(CI-Cis) alkyl, -O-(Cz-C16) alkenyl, -O-(Cz-C16) alkynyl,
-aryl, -O-aryl, -O-
CHz-aryl, -OC(O)R, -C(O)R, -C(O)OR, -C(O)NR'R', -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one group selected from halo, -CN, -O-
alkyl, -O-alkenyl,
-O-allcynyl, -O-aryl, -OC(O)R', -C(O)R', and -C(O)NR'R':
said alkenyl, alkynyl, -O-alkyl, -O-alkenyl and -O-alkynyl are each
independently substituted .
with at least one group selected from halo, -CN, -OH, -OR', -O-aryl, -OC(O)R',
-C(O)R', -
C(O)OR' and -C(O)NR'R', and
R and R' are as defined above.
18
CA 02568609 2006-11-28
In another embodiment, the substituents of the of the compounds of formula (I)
are as follows:
XisFCl,BrorI;
Rl and R2 are each independently selected from the group of H, (C1-C6) alkyl,
(C1-C6) hydroxy
alkyl, or R1 and R2 when taken together form =O;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine; and
substituted with at least one substituent selected from (CI-C16) alkyl, (C2-
C16) alkenyl, (CZ-Cls)
alkynyl, -O-(C1-C16) alkyl, -O-(CZ-C16) alkenyl, -O-(C2-C16) alkynyl, aryl,
substituted aryl, -O-
aryl, -O-CH2-aryl, -S-(C1-C16) alkyl, -S-(Cz-C16) alkenyl, -S-(C2-C16)
alkynyl, -S-aryl, -S-CH2-
aryl, (C3-C$) cycloalkyl, -O-(C3-C$) cycloalkyl, -S-(C3-C8) cycloalkyl, -ONRR,
-C(O)R, -C(S)R
-C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -SC(S)R, -OC(S)R, -
C(O)NRR, -
C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -C(S)NR(SR), -CH(CN)2,
-CH[C(O)R]2, -CH[C(S)R]2, -CH[C(O)OR]Z, -CH[C(S)OR]2, -CH[C(O)SR]Z, -
CH[C(S)SR]2,
-NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)20R, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -NO2,
-NR'R', -O-alkyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R ;
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -NOZ, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R';
said -O-alkenyl, -O-allcynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-
cycloalkyl are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NOZ, -
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above.
In another embodiment, the compounds of formula (I) include the compounds of
formula (II):
19
CA 02568609 2006-11-28
O
B-N~N~CH X
2
H H
or a pharmaceutically acceptable salt thereof, wherein:
XisFCl,BrorI;
B is an aryl group selected from phenyl, indaaie, fluorene, indazole, indole,
and pyridine;
wherein:
B is substituted with one or more substituents selected from the group of (Ci-
Clg) alkyl,
(CZ-C16) alkenyl, (C2-C16) alkynyl, aryl, -O-(C1-C16) alkyl, -O-(C2-C16)
alkenyl, -O-(C2-Cls)
alkynyl, -O-aryl, -O-CH2-aryl, -S-(C1-C16)alkyl, -S-(CZ-C16) alkenyl, -S-(Ca-
C16) alkynyl, -S-
aryl, -S-CHZ-aryl, (C3-C8) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8)
cycloalkyl, -halo, -NRR,
-ONRR -NOZ, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -
SC(O)R, -SC(S)R, -OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -
C(O)NR(SR), -C(S)NR(SR), -CH(CN)2, -CH[C(O)R]~, -CH[C(S)R]a, -CH[C(O)OR]a, -
CH[C(S)OR]Z, -CH[C(O)SR]2, -CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -
S(O)20R, -S(O)NRR, -S(O)ONRR;
wherein:
each R is independently selected from -H, (C1-C16) alkyl, substituted (Cl-C16)
alkyl, (Ca-
C16) alkenyl, substituted (C2-C16) alkenyl, (CZ-C16) alkynyl, substituted (C2-
C16) alkynyl, (C3-C8)
cycloalkyl, substituted (C3-C8) cycloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl are optionally substituted with one or
more
substituents independently selected from the group of -halo, trihalomethyl, -
R', -OR', -SR', -
NR'R', -N02, -CN, -OC(O)R', -OC(S)R', -SC(O)R, -SC(S)R -C(O)R', -C(S)R', -
C(O)OR', -
C(S)OR', -C(O)SR', -C(S)SR', -C(O)NR'R', -C(S)NR'R', -NR'C(O)R' and -
NR'C(O)OR;
the cycloallcyl is optionally substituted with one or more substituents
independently
selected from the group of R', -halo, OR', -SR', -NRR', -ONRR', -NOZ, -CN, -
C(O)R', -C(S)R',
-OC(O)R', -SC(O)R', -SC(S)R', -OC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -
C(S)SR', -
C(O)NRR', -C(S)NRR', -C(O)NR'(OR~, -C(S)NR' -(ORS, -C(O)NR'(SR~, -C(S)NR'(SR~,
_
CH(CN)a, -CH[C(O)R']2, -CH[C(S)R~2, -CH[C(O)OR~2, -CH[C(S)OR']a, -CH[C(O)SR~2,
-
CA 02568609 2006-11-28
CH[C(S)SR']2, -NR'C(O)R', -NR'C(O)OR', -S(O)-R', -S(O)OR', -S(O)20R', -
S(O)NRR', -
S(O)ONRR', and
each R' is independently selected from the group of -H, (C1-C16) alkyl,
substituted (C1-C16)
alkyl, (C2-C16) alkenyl, substituted (CZ-C16) alkenyl, (C2-C16) alkynyl,
substituted (C2-C16)
alkynyl, (C3-Cg) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or
substituted aryl.
In a specific embodiment in the compound of formula (II) B is phenyl,
substituted with one or
more substituents indepnednetly selected from the list as shown above.
W another embodiment, in the compounds of formula II, X is Cl or Br.
In another embodiment the substituents of the compounds of formula (II) are as
follows:
XisF,Cl,BrorI;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine; and
substituted with at least one substituent selected from (C1-Cls) alkyl, (CZ-
C16) alkenyl, (CZ-Cls)
alkynyl, -O-(C1-C16) alkyl, -O-(CZ-C16) alkenyl, -O-(C2-C16) alkynyl, aryl,
substituted aryl, -O-
aryl, -O-CHZ-aryl, -S-(CI-C16) alkyl, -S-(CZ-C16) alkenyl, -S-(C2-C16)
alkynyl, -S-aryl, -S-CH2-
aryl, (C3-C8) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8) cycloalkyl, -ONRR,
-C(O)R, -C(S)R
-C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -SC(S)R, -OC(S)R, -
C(O)NRR, -
C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -C(S)NR(SR), -CH(CN)2,
-CH[C(O)R]2, -CH[C(S)R]2, -CH[C(O)OR]Z, -CH[C(S)OR]2, -CH[C(O)SR]2, -
CH[C(S)SR]2,
-NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)ZOR, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -NOz,
-NR'R', -O-alkyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R';
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -NOa, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R';
said -O-alkenyl, -O-alkynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-cycloalkyl
are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NO2, -
21
CA 02568609 2006-11-28
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above
In another embodiment the substituents of the compounds of formula (I) are as
follows:
X is F, Cl, Br or I;
B is phenyl substituted with at least one substituent selected from (C1-C16)
alkyl, (C2-C16)
alkenyl, (CZ-C16) alkynyl, -O-(Cl-C16) alkyl, -O-(CZ-C~6) alkenyl, -O-(CZ-C16)
alkynyl, aryl,
substituted aryl, -O-aryl, -O-CHZ-aryl, -S-(C1-C16) alkyl, -S-(C2-C16)
alkenyl, -S-(CZ-Cls)
alkynyl, -S-aryl, -S-CHZ-aryl, (C3-C$) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-
(C3-C8) cycloalkyl, -
ONRR, -C(O)R, -C(S)R -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -
SC(S)R,
-OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -
C(S)NR(SR), -
CH(CN)z, -CH[C(O)R]Z, -CH[C(S)R]2, -CH[C(O)OR]2, -CH[C(S)OR]2, -CH[C(O)SR]2, -
CH[C(S)SR]Z, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)ZOR, -S(O)NRR, -
S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -N02,
-NR'R', -O-allcyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R';
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -NOZ, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R ;
said -O-alkenyl, -O-alkynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-cycloalkyl
are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NOa, -
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above.
In another embodiment the substituents of the of the compounds of formula (I)
are as follows:
XisF,Cl,BrorI;
22
CA 02568609 2006-11-28
B is phenyl substituted with at least one group selected from (C1-C16) alkyl,
(C2-C16) alkenyl,
(C2-C16) alkynyl, -O-(C1-C16) alkyl, -O-(Cz-C16) alkenyl, -O-(Cz-C16) alkynyl,
-aryl, -O-aryl, -O-
CH2-aryl, -OC(O)R, -C(O)R, -C(O)OR, -C(O)NR'R', -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one group selected from halo, -CN, -O-
alkyl, -O-alkenyl,
-O-alkynyl, -O-aryl, -OC(O)R', -C(O)R', and -C(O)NR'R':
said alkenyl, alkynyl, -O-alkyl, -O-alkenyl and -O-alkynyl are each
independently substituted
with at least one group selected from halo, -CN, -OH, -OR', -O-aryl, -OC(O)R',
-C(O)R', -
C(O)OR' and -C(O)NR'R', and
R and R' are as defined above.
In another embodiment the substituents of the of the compounds of formula (II)
are as follows:
X is F, Cl, Br or I; and
B is substituted with at least one group selected from aryl, -O-aryl, -O-CHz-
aryl and halo.
In another embodiment the substituents of the of the compounds of formula (II)
are as follows:
X is F, Cl, Br or I;
B is substituted with at least one substituent selected from the group of (C1-
C16) alkyl, (C1-Cls)
alkynyl or -O-alkyl;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of -CN, -O-alkyl, -
OC(O)R', -C(O)R', , -C(O)NR'R' or halo;
said alkyny and -O-alkyl are are substituted with at least one substituent
selected from -CN, -
OH, -O-alkyl, -OC(O)R', -C(O)R', -C(O)OR', -C(O)NR'R' or halo; and
R' is as defined above.
In another embodiment the substituents of the of the compounds of formula (II)
are as follows:
X is F, Cl, Br or I;
B is substituted with at least one group selected from -NRC(O)R, -NRC(O)OR, -
S(O)-R,
23
CA 02568609 2006-11-28
-S(O)OR, -S(O)ZOR, -S(O)NRR, -S(O)ONRR, -C(O)R, -C(O)OR, -OC(O)R, -C(O)NRR;
wherein R is as defined above.
In another embodiment, the compounds of formula (I) include the compounds of
formula (III):
O O
B-N- 'N' 'CH X III
2
H H
or a pharmaceutically acceptable salt thereof, wherein:
XisF,Cl,BrorI;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine;
wherein:
B is substituted with one or more substituents selected from the group of (C1-
C16) alkyl, (CZ-Cls)
alkenyl, (C2-C16) alkynyl, aryl, -O-(C1-C16) alkyl, -O-(CZ-C16) alkenyl, -O-
(C2-C16) alkynyl, -O-
aryl, -O-CHZ-aryl, -S-(C1-C16)alkyl, -S-(C2-C16) alkenyl, -S-(C2-C16) alkynyl,
-S-aryl, -S-CH2-
aryl, (C3-C8) cycloalkyl, -O-(C3-C$) cycloalkyl, -S-(C3-C8) cycloalkyl, -halo,
-NRR, -ONRR -
NOa, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -
SC(O)R,
-SC(S)R, -OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR),
-C(S)NR(SR), -CH(CN)z, -CH[C(O)R]2, -CH[C(S)R]2, -CH[C(O)OR]Z, -CH[C(S)OR]2,
-CH[C(O)SR]2, -CH[C(S)SR]Z, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)20R,
-S(O)NRR, -S(O)ONRR;
wherein:
each R is independently selected from -H, (C1-C16) alkyl, substituted (C1-C16)
alkyl, (C2-
C16) alkenyl, substituted (CZ-C16) alkenyl, (CZ-C16) allcynyl, substituted (CZ-
C16) alkynyl, (C3-C$)
cycloalkyl, substituted (C3-Cg) cycloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl are optionally substituted with one or
more substituents
independently selected from the group of -halo, trihalomethyl, -R', -OR', -
SR', -NR'R', -N02, -
CN, -OC(O)R', -OC(S)R', -SC(O)R, -SC(S)R -C(O)R', -C(S)R', -C(O)OR', -C(S)OR',
-C(O)SR',
-C(S)SR', -C(O)NR'R', -C(S)NR'R', -NR'C(O)R' and -NR'C(O)OR;
24
CA 02568609 2006-11-28
the cycloalkyl is optionally substituted with one or more substituents
independently selected
from the group of R', -halo, OR', -SR', -NRR', -ONRR', -NOa, -CN, -C(O)R', -
C(S)R', -
OC(O)R', -SC(O)R', -SC(S)R', -OC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR',
-
C(O)NRR', -C(S)NRR', -C(O)NR'(OR~, -C(S)NR' -(ORS, -C(O)NR'(SR~, -C(S)NR'(SR7,
-
CH(CN)2, -CH[C(O)R']2, -CH[C(S)R~a, -CH[C(O)OR~2, -CH[C(S)OR']2; -CH[C(O)SR~2,
-
CH[C(S)SR~2, -NR'C(O)R', -NR'C(O)OR', -S(O)-R', -S(O)OR', -S(O)20R', -
S(O)NRR', -
S(O)ONRR', and
each R' is independently selected from the group of -H, (C1-C16) alkyl,
substituted (C1-C16)
alkyl, (CZ-C16) alkenyl, substituted (C2-C16) alkenyl, (C2-C16) alkynyl,
substituted (CZ-C16)
alkynyl, (C3-C8) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or
substituted aryl.
In a specific embodiment in the compound of formula (III) B is phenyl,
substituted with one or
more substituents indepnednetly selected from the list as shown above.
In another specific embodiment, the substitutents of formula (I) are as
follows:
O R~ R~
B-N- 'N' 'CH X
2
H H
XisFCl,BrorI;
R1 and R2 are each independently selected from the group of H, (C1-C6) alkyl,
(C1-C6)
hydroxy alkyl, (C1-C6) alkoxy, (C3-C~) cycloalkyl, halo substituted (C1-C6)
alkyl, halo di-
substituted (C1-C6) alkyl, halo tri-substituted (C1-C6) alkyl and halo;
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine;
wherein:
B is substituted with one or more substituents selected from the group of (C1-
C16) alkyl, (C2-C16)
alkenyl, (C2-C16) alkynyl, aryl, -O-(C1-C16) alkyl, -O-(Cz-C16) alkenyl, -O-
(Cz-C16) alkynyl, -O-
aryl, -O-CH2-aryl, -S-(C~-C16)alkyl, -S-(C2-C16) alkenyl, -S-(C2-Ci6) alkynyl,
-S-aryl, -S-CH2-
aryl, (C3-C8) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8) cycloalkyl, -halo,
-NRR, -ONRR,
CA 02568609 2006-11-28
-N02, -CN, -C(O)R, -C(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -
SC(O)R,
-SC(S)R, -OC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR),
-C(S)NR(SR), -CH(CN)2, -CH[C(O)R]a, -CH[C(S)R]Z, -CH[C(O)OR]2, -CH[C(S)OR]Z,
-CH[C(O)SR]2, -CH[C(S)SR]a, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)aOR,
-S(O)NRR, -S(O)ONRR;
wherein:
each R is independently selected from -H, (C1-C16) alkyl, substituted (C1-C16)
alkyl, (C2-
C16) alkenyl, substituted (C2-C16) alkenyl, (Ca-C16) alkynyl, substituted (CZ-
C16) alkynyl, (C3-C8)
cycloalkyl, substituted (C3-C8) cycloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl are optionally substituted with one or
more substituents
independently selected from the group of -halo, trihalomethyl, -R', -OR', -
SR', -NR'R', -N02, -
CN, -OC(O)R', -OC(S)R', -SC(O)R, -SC(S)R -C(O)R', -C(S)R', -C(O)OR', -C(S)OR',
-C(O)SR',
-C(S)SR', -C(O)NR'R', -C(S)NR'R', -NR'C(O)R' and -NR'C(O)OR';
the cycloalkyl is optionally substituted with one or more substituents
independently selected
from the group of R', -halo, OR', -SR', -NRR', -ONRR', -NOa, -CN, -C(O)R', -
C(S)R', -
OC(O)R', -SC(O)R', -SC(S)R', -OC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR',
-
C(O)NRR', -C(S)NRR', -C(O)NR'(OR~, -C(S)NR' -(ORS, -C(O)NR'(SR~, -C(S)NR'(SR7,
-
CH(CN)2, -CH[C(O)R']2, -CH[C(S)R']2, -CH[C(O)OR']2, -CH[C(S)OR']2, -
CH[C(O)SR']2, -
CH[C(S)SR']2, -NR'C(O)R', -NR'C(O)OR', -S(O)-R', -S(O)OR', -S(O)ZOR', -
S(O)NRR', -
S(O)ONRR', and
each R' is independently selected from the group of -H, (CI-C16) alkyl,
substituted (Cl-C16) alkyl,
(CZ-C16) allcenyl, substituted (CZ-C16) alkenyl, (CZ-C16) alkynyl, substituted
(C2-C16) alkynyl,
(C3-C8) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or substituted aryl.
In another specific embodiment the substituents of the compounds of formula
(I) are as follows:
XisF,Cl,BrorI;
Rl and R2 are each independently selected from the group of H, (C1-C6) alkyl,
(C1-C6) hydroxy
alkyl, (CI-C6) alkoxy, (C3-C~) cycloalkyl, halo substituted (C1-C6) alkyl,
halo di-substituted (C1-
C6) alkyl, halo tri-substituted (C1-C6) alkyl and halo;
26
CA 02568609 2006-11-28
B is an aryl group selected from phenyl, indane, fluorene, indazole, indole,
and pyridine; and
substituted with at least one substituent selected from (C1-C16) alkyl, (C2-
C16) alkenyl, (CZ-C16)
alkynyl, -O-(C1-C16) alkyl, -O-(C2-C16) alkenyl, -O-(C2-C16) alkynyl, aryl,
substituted aryl, -O-
aryl, -O-CHZ-aryl, -S-(C1-C16) alkyl, -S-(C2-Clg) alkenyl, -S-(CZ-C16)
alkynyl, -S-aryl, -S-CH2-
aryl, (C3-C$) cycloalkyl, -O-(C3-C8) cycloalkyl, -S-(C3-C8) cycloalkyl, -ONRR,
-C(O)R, -C(S)R
-C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R, -SC(S)R, -OC(S)R, -
C(O)NRR, -
C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -C(S)NR(SR), -CH(CN)2,
-CH[C(O)R]2, -CH[C(S)R]Z, -CH[C(O)OR]2, -CH[C(S)OR]2, -CH[C(O)SR]2, -
CH[C(S)SR]z,
-NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)ZOR, -S(O)NRR, -S(O)ONRR;
wherein:
said alkyl is substituted with at least one substituent selected from the
group of halo, -CN, -N02,
-NR'R', -O-allcyl, -O-alkenyl, -O-alkynyl, -O-aryl, -OC(O)R', -OC(S)R', -
C(O)R', -C(S)R', -
C(O)NR'R' and -C(S)NR'R ;
said alkenyl, alkylnyl, -O-alkyl, -S-alkyl, are each independently substituted
with at least one
group selected from halo, -CN, -N02, -NR'R', -OH, -OR', -O-aryl, -OC(O)R', -
OC(S)R', -C(O)R',
-C(S)R', -C(O)OR', -C(O)NR'R' and -C(S)NR'R ;
said -O-alkenyl, -O-alkynyl, -S-alkenyl, -S-alkynyl, cycloalkyl, -O-cycloalkyl
are are each
optionally and independently substituted with at least one group selected from
halo, -CN, -NOZ, -
NR'R', -OH, -OR', -O-aryl, -OC(O)R', -OC(S)R', -C(O)R', -C(S)R', -C(O)OR', -
C(O)NR'R' and -
C(S)NR'R', and
wherein R, R' and aryl are as defined above and their substituents are as
defined above.
In another embodiment, the substituents of the compounds of formula (I) are as
follows:
XisF,Cl,BrorI;
Rl and R2 are each independently selected from the group of H, (C1-C6) alkyl,
(C1-C6) hydroxy
alkyl, (C1-C6) alkoxy, (C3-C~) cycloalkyl, halo substituted (C1-C6) alkyl,
halo di-substituted (C1-
Cg) alkyl, halo tri-substituted (C1-C6) alkyl and halo,
B is phenyl substituted with at least one group selected from (C1-C16) alkyl,
(C2-C16) alkenyl,
(Cz-C16) alkynyl, -O-(C1-C16) alkyl, -O-(C2-C16) alkenyl, -O-(CZ-C16) alkynyl,
-aryl, -O-aryl, -O-
CH2-aryl, -OC(O)R, -C(O)R, -C(O)OR, -C(O)NR'R', -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -S(O)NRR, -S(O)ONRR;
27
CA 02568609 2006-11-28
wherein:
said alkyl is substituted with at least one group selected from halo, -CN, -O-
alkyl, -O-alkenyl,
-O-alkynyl, -O-aryl, -OC(O)R', -C(O)R', and -C(O)NR'R':
said alkenyl, alkynyl, -O-alkyl, -O-alkenyl and -O-alkynyl are each
independently substituted
with at least one group selected from halo, -CN, -OH, -OR', -O-aryl, -OC(O)R',
-C(O)R', -
C(O)OR' and -C(O)NR'R', and
R and R' are as defined above.
In an another embodiment, the compounds of formula (I) include the compounds
of formula
(1V):
O R~R2
N~N~CH
~X
H H
R3
or a pharmaceutically acceptable salt thereof, wherein:
R3 is selected from the group of H, R, -halo, OR, -SR, -NRR, -ONRR, -NOZ, -CN,
-C(O)R, -C(S)R,-OC(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R,
-SC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -
C(S)NR(SR),
-CH(CN)2, -CH[C(O)R]2, -CH[C(S)R]2, -CH[C(O)OR]2, -CH[C(S)OR]2, -CH[C(O)SR]2,
-CH[C(S)SR]Z, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)ZOR, -S(O)NRR,
-S (O)ONRR;
each R is independently selected from -H, (C1-C16) alkyl, substituted (C1-C16)
alkyl, (C2-
C16) alkenyl, substituted (C2-C16) alkenyl, (Ca-C16) alkynyl, substituted (C2-
C16) alkynyl, (C3-C$)
cycloalkyl, substituted (C3-C$) cyloalkyl, aryl or substituted aryl;
the alkyl, alkenyl, alkynyl and aryl substituents are each independently
selected from the
group of -halo, trihalomethyl, -R', -OR', -SR', -NR'R', -N02, -CN, -OC(O)R', -
OC(S)R', -
SC(O)R', -SC(S)R' -C(O)R', -C(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR', -
C(O)NR'R',
-C(S)NR'R', -NR'C(O)R', -NR'C(O)OR' and aryl;
28
CA 02568609 2006-11-28
the cycloalkyl substituents are each independently selected from the group of
H, R, -halo,
OR, -SR, -NRR, -ONRR -N02, -CN, -C(O)R, -C(S)R, -OC(O)R, -SC(O)R, -SC(S)R, -
OC(S)R, -
C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -
C(S)NR(OR), -
C(O)NR(SR), -C(S)NR(SR), -CH(CI~Z, -CH[C(O)R]Z, -CH[C(S)R]2, -CH[C(O)OR]a, -
CH[C(S)OR]2, -CH[C(O)SR]2, -CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -
S(O)20R, -S(O)NRR, -S(O)ONRR', and
each R' is independently selected from the group -H, (C1-C16) alkyl,
substituted (C1-Cls)
alkyl, (CZ-C16) alkenyl, substituted (CZ-C16) alkenyl, (CZ-C16) alkynyl,
substituted (CZ-C16)
alkynyl, (C3-C8) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or
substituted aryl.
In another embodiment in the compoundof formula (IV), R3 is selected from
halo, -CN,
-C(O)R, -C(O)OR, -OC(O)R, -C(O)NRR, -OR, (C~-C16) alkyl, (Ca-C16) alkenyl, (C2-
C16)
alkynyl, wherein said alkyl, alkenyl and alkynyl are optionally substituted
with -CN, -C(O)R', -
C(O)OR',
-OC(O)R', -C(O)NR'R', -OR', wherein R and R' are as defined above.
In an another embodiment, the compounds of formula (I) include the compounds
of formula (V):
R~ R2
Rs ~ ~ N' 'N. 'CH
H H 2X
or a pharmaceutically acceptable salt thereof, wherein:
R3 is selected from the group of H, R, -halo, OR, -SR, -NRR, -ONRR, -NOa~ -CN,
-C(O)R, -C(S)R,-OC(S)R, -C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -OC(O)R, -SC(O)R,
-SC(S)R, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -C(S)NR(OR), -C(O)NR(SR), -
C(S)NR(SR),
-CH(CN7z, -CH[C(O)R]2, -CH[C(S)R]Z, -CH[C(O)OR]2, -CH[C(S)OR]a, -CH[C(O)SR]2,
-CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -S(O)OR, -S(O)20R, -S(O)NRR,
-S(O)ONRR;
29
CA 02568609 2006-11-28
each R is independently selected from -H, (C1-Cls) alkyl, substituted (C1-C16)
alkyl, (C2-
Cis) alkenyl, substituted (C2-C16) alkenyl, (Ca-C16) alkynyl, substituted (C~-
C16) alkynyl, (C3-C$)
cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or substituted aryl;
the aryl substituents are each independently selected from the group of -halo,
trihalomethyl, -R', -OR', -SR', -NR'R', -N02, -CN, -C(O)R', -C(S)R', -OC(O)R',
-OC(S)R',
-SC(O)R', -SC(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR', -C(O)NR'R', -
C(S)NR'R',
-NR'C(O)R', -NR'C(O)OR ;
the alkyl, alkenyl and alkynyl substituents are each independently selected
from the group of
-halo, trihalomethyl, -R', -OR', -SR', -NR'R', -NO2, -CN, -OC(O)R', -OC(S)R', -
SC(O)R',
-SC(S)R' -C(O)R', -C(S)R', -C(O)OR', -C(S)OR', -C(O)SR', -C(S)SR', -C(O)NR'R',
-C(S)NR'R', -NR'C(O)R', -NR'C(O)OR' and aryl;
the cycloallcyl substituents are each independently selected from the group of
H, R, -halo,
OR, -SR, -NRR, -ONRR -NO2, -CN, -C(O)R, -C(S)R, -OC(O)R, -SC(O)R, -SC(S)R, -
OC(S)R, -
C(O)OR, -C(S)OR, -C(O)SR, -C(S)SR, -C(O)NRR, -C(S)NRR, -C(O)NR(OR), -
C(S)NR(OR), -
C(O)NR(SR), -C(S)NR(SR), -CH(CN)Z, -CH[C(O)R]2, -CH[C(S)R]Z, -CH[C(O)OR]a, -
CH[C(S)OR]a, -CH[C(O)SR]2, -CH[C(S)SR]2, -NRC(O)R, -NRC(O)OR, -S(O)-R, -
S(O)OR, -
S(O)ZOR, -S(O)NRR, -S(O)ONRR', and
each R' is independently selected from the group -H, (C1-C16) alkyl,
substituted (C1-Cls)
alkyl, (C2-C16) alkenyl, substituted (C2-C16) allcenyl, (Ca-C16) alkynyl,
substituted (Ca-Cls)
alkynyl, (C3-C8) cycloalkyl, substituted (C3-C8) cyloalkyl, aryl or
substituted aryl.
In another embodiment in the compoundof formula (V), R3 is selected from halo,
-CN,
-C(O)R, -C(O)OR, -OC(O)R, -C(O)NRR, -OR, (C1-C16) alkyl, (CZ-C16) alkenyl, (C2-
Ci6)
alkynyl, wherein said alkyl, allcenyl and alkynyl are optionally substituted
with -CN, -C(O)R', -
C(O)OR',
-OC(O)R', -C(O)NR'R', -OR', wherein R and R' are as defined above.
In an illustrative embodiment, the compounds according to formula (1~ include
those listed
below:
4-iodo-1-[3-(2-chloroethyl)ureido]benzene;
4-tert-butyl-1-[3-(2-chloroethyl)ureido]benzene;
CA 02568609 2006-11-28
4-sec-butyl-1-[3-(2-chloroethyl)ureido]benzene;
4-isopropyl-1-[3-(2-chloroethyl)ureido]benzene;
4-(p-[3-(2-chloroethyl)ureido) phenyl)butanol;
4-pentyl-1-[3-(2-chloroethyl)ureido] benzene;
4-hexyl-1-[3-(2-chloroethyl)ureido] benzene;
4-cyclohexyl-1-[3-(2-chloroethyl)ureido] benzene;
4-heptyl-1-[3-(2-chloroethyl)ureido] benzene;
4-octyl-1-[3-(2-chloroethyl)ureido) benzene;
4-decyl-1-[3-(2-chloroethyl)ureido) benzene;
4-dodecyl-1-[3-(2-chloroethyl)ureido] benzene;
3-ethoxy-1-[3-(2-chloroethyl)ureido] benzene;
4-pentoxy-1-[3-(2-chloroethyl)ureido] benzene;
4-hexyloxy-1-[3-(2-chloroethyl)ureido] benzene;
2,4,6-trimethyl-1-[3-(2-chloroethyl)ureido] benzene;
2-ethyl-1-[3-(2-chloroethyl)ureido] benzene;
2,4-diethyl-1-[3-(2-chloroethyl)ureido] benzene;
2-propyl-1-[3-(2-chloroethyl)ureido] benzene;
2,6-diisopropyl-1-[3-(2-chloroethyl)ureido] benzene;
2-isopropyl-6-methyl-1-[3-(2-chloroethyl)ureido] benzene;
2,5-ditert butyl-1-[3-(2-chloroethyl)ureido] benzene;
2-methoxy-5-methyl-1-[3-(2-chloroethyl)ureido] benzene;
2-methoxy-6-methyl-1-[3-(2-chloroethyl)ureido] benzene;
2-methyl-5-methoxy-1-[3-(2-chloroethyl)ureido] benzene;
2-methyl-4-methoxy-1-[3-(2-chloroethyl)ureido] benzene;
2-ethoxy-1-[3-(2-chloroethyl)ureido] benzene;
4-ethoxy-1-(3-(2-chloroethyl)ureido] benzene;
4-butoxy-1-(3-(2-chloroethyl)ureido] benzene;
1,4-di[3-(2-chloroethyl)ureido] benzene;
2-cyano-1-[3-(2-chloroethyl)ureido] benzene;
4-(3-hydroxypropyl)-1-[3-(2-chloroethyl)ureido] benzene;
4-(2-hydroxybutyl)-1-[3-(2-chloroethyl)ureido] benzene;
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4-(4-(2-butyrate ethanoic acid))-1-[3-(2-chloroethyl)ureido] benzene;
4-(3-(2-propylate ethanoic acid))-1-[3-(2-chloroethyl)ureido] benzene;
4-ethylthio-1-[3-(2-chloroethyl)ureido] benzene;
2-[3-(2-chloroethyl)ureido] fluorene;
5-[3-(2-chloroethyl)ureido] indane;
6-[3-(2-chloroethyl)ureido] indazole;
5-[3-(2-chloroethyl)ureido] 4,6-dimethyl pyridine;
5-[3-(2-chloroethyl)ureido] indole;
1-(4-tert-Butyl-phenyl)-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-ethyl)-3-(4-cyclohexyl-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(4-heptyl-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pentyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(4-methoxy-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(4-iodo-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(4-phenoxy-phenyl)-urea;
1-(4-B enzyloxy-phenyl)-3 -(2-chloro-ethyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(2-methoxy-ethyl)-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(4-ethoxy-butyl)-phenyl]-urea;
1-Biphenyl-4-yl-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-acetyl)-3-(4-iodo-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(4-fluoro-butyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pent-1-ynyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(4-hydroxy-phenyl)-urea;
Acetic acid 5- f 4-[3-(2-chloro-ethyl)-ureido]-phenyl}-pentyl ester;
N- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-acetamide;
N-Butyl-4-(5-chloro-1,3-dimethyl-2-oxo-pentyl)-benzenesulfonamide;
1-(2-Chloro-ethyl)-3-[3 -( 1-hydroxy-ethyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3 -( 1-hydroxy-ethyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(2-heptyl-phenyl)-urea;
1-(2-Chloro-acetyl)-3-[3-(5-hydroxy-pentyl)-phenyl]-urea;
R-1-(4-tert-Butyl-phenyl)-3-(2-chloro-1-methyl-ethyl)-urea;
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S-1-(4-tart-Butyl-phenyl)-3-(2-chloro-1-methyl-ethyl)-urea;
1-(4-tent-Butyl-phenyl)-3-(2-chloro-1,1-dimethyl-ethyl)-urea;
1-(2-Bromo-ethyl)-3-(3-iodo-phenyl)-urea;
3-[3-(2-Bromo-ethyl)-ureido]-benzoic acid ethyl ester.
In another illustrative embodiment, the compounds according to formula (I)
include those listed
below:
4-iodo-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-ethyl)-3-(4-iodo-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(4-phenoxy-phenyl)-urea;
1-(4-Benzyloxy-phenyl)-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(2-methoxy-ethyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[4-(4-ethoxy-butyl)-phenyl]-urea;
1-Biphenyl-4-yl-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-acetyl)-3-(4-iodo-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(4-fluoro-butyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pent-1-ynyl)-phenyl]-urea;
Acetic acid 5-~4-[3-(2-chloro-ethyl)-ureido]-phenyl}-pentyl ester;
n- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl]-acetamide;
n-Butyl-4-(5-chloro-1,3-dimethyl-2-oxo-pentyl)-benzenesulfonamide;
1-(2-Chloro-ethyl)-3-[3-( 1-hydroxy-ethyl)-phenyl]-urea;?
1-(2-Bromo-ethyl)-3-(3-iodo-phenyl)-urea;
3-[3-(2-Bromo-ethyl)-ureido]-benzoic acid ethyl ester;
1-(4-n-hexyl-phenyl)-3-(2-chloro-1-methyl-ethyl)urea;
1-(4-iodo-phenyl)-3-(2-bromo-1-methyl-ethyl)urea;
(R) 1-(4-iodo-phenyl)-3-(2-bromo-1-methyl-ethyl)urea;
(S) 1-(4-iodo-phenyl)-3 -(2-bromo-1-methyl-ethyl)urea.
In another illustrative embodiment, the compounds according to formula (I)
include those listed
below:
1-(4-tart-butylphenyl)-3 -(2-chloroethyl)urea;
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CA 02568609 2006-11-28
1-(2-chloroethyl)-3-(4-cyclohexylphenyl)urea;
1-(2-chloroethyl)-3-(4-hepthylphenyl)urea;
1-(2-chloroethyl)-3-(4-iodophenyl)urea;
1-(2-chloroethyl)-3-(4-phenoxyphenyl)urea;
1-(4-benzyloxyphenyl)-3-(2-chloroethyl)urea;
1-(biphenyl-4-yl)-3-(2-chloroethyl)urea;
1-(2-chloroethyl)-3-(4-hydroxyphenyl)urea;
N f 3-[3-(2-chloroethyl)ureido]phenyl} acetamide;
N Butyl-3-[3-(2-chloroethyl)ureido]benzenesulfonamide;
1-(2-chloroethyl)-3-[3-(1-hydroxyethyl)phenyl]urea;
1-(2-chloroethyl)3-[4-(2-methoxyethyl)phenyl)urea;
1-(2-chloroethyl)-3-[4-(4-ethoxybutyl)phenyl)urea;
1-(2-chloroethyl)-3-[4-(4-fluorobutyl)phenyl]urea;
1-(2-chloro ethyl)-3-[3 -(5-hydroxypent-1-ynyl)phenyl)urea;
1-(2-chloroethyl)-3-[3-(5-hydroxypentyl)phenyl)urea;
Acetic acid 5-{4-[3-(2-chloroethyl)ureido]phenyl}pentyl ester;
6-~3-[3-(2-chloroethyl)ureido]phenoxy)hexanoic acid ethyl ester;
1-(2-chloroethyl)-2-(2-heptylphenyl)urea;
1-(2-Chloroacetyl)-3-(4-iodophenyl)urea;
1-(2-chloroacetyl)-3-[3-(5-hydroxypentyl)phenyl]urea;
(R)-1-(2-chloro-1-methylethyl)-3 -(4-iodophenyl)urea;
(xS)-1-(2-chloro-1-methylethyl)-3-(4-iodophenyl)urea;
(R)-1-(4-tent-Butylphenyl)-3 -(2-chloro-1-methylethyl)urea;
(S~-1-(4-tent-Butylphenyl)-3-(2-chloro-1-methylethyl)urea;
1-(4-teat-Butylphenyl)-3-(2-chloro-1,1-dimethylethyl)urea;
1-(2-Bromoethyl)-3-(3-iodophenyl)urea;
4-test-Butylphenyl(4,5-dihydrooxazol-2-yl)amine;
In another illustrative embodiment, the compounds according to formula (17
include those listed
below:
1-(2-Chloro-ethyl)-3-m-tolyl-urea;
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CA 02568609 2006-11-28
1-(2-Chloro-ethyl)-3-(3-ethyl-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(3-methoxy-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(4-hydroxy-butyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[4-(3-hydroxy-propyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(3-iodo-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[4-(5-hydroxy-pentyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pent-1-ynyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pentyl)-phenyl]-urea;
3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid ethyl ester;
1-(2-Chloro-ethyl)-3-[3-(5-methoxy-pentyl)-phenyl]-urea;
f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl-acetic acid;
1-(2-Bromo-ethyl)-3-[3-(5-hydroxy-pentyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(3-heptyl-phenyl)-urea;
1-(2-Chloro-ethyl)-3-[3-(6-hydroxy-hexyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(4-hydroxy-butyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-[3-(3-hydroxy-propyl)-phenyl]-urea;
5-~3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-pentanoic acid amide;
1-(3-Bromo-phenyl)-3-(2-chloro-ethyl)-urea;
1-(2-Chloro-ethyl)-3-(3-chloro-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(3-hydroxy-phenyl)-urea;
Acetic acid 3-[3-(2-chloro-ethyl)-ureido]-phenyl ester;
1-(2-Chloro-ethyl)-3-(3-hydroxymethyl-phenyl)-urea;
Acetic acid 5- f 3-[3-(2-chloro-ethyl)-ureido]-phenyl}-pentyl ester;
Acetic acid 4-~3-[3-(2-chloro-ethyl)-ureido]-phenyl]-butyl ester;
Acetic acid 3-[3-(2-chloro-ethyl)-ureido]-benzyl ester;
Acetic acid 3- f 3-[3-(2-chloro-ethyl)-ureido]-phenyl}-propyl ester;
1-(2-Chloro-ethyl)-3-[3-(2-hydroxy-ethyl)-phenyl]-urea;
Acetic acid 2- f 3-[3-(2-chloro-ethyl)-ureido]-phenyl]-ethyl ester;
Acetic acid 6-{3-[3-(2-chloro-ethyl)-ureido)-phenyl}-hexyl ester;
Pentanedioic acid mono-{3-[3-(2-chloro-ethyl)-ureido]-phenyl] ester;
1-(2-Chloro-ethyl)-3-[3-(7-hydroxy-heptyl)-phenyl]-urea;
CA 02568609 2006-11-28
1-(2-Chloro-ethyl)-3-(3-cyano-phenyl)-urea;
3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid;
1-(2-Chloro-ethyl)-3-[3-(3-methoxy-propyl)-phenyl]-urea;
1-(3-pentyl-phenyl)-3-(2-chloro-ethyl)-urea;
5-{3-[3-(2-Chloro-ethyl)-ureidoJ-phenyl)-pentanoic acid ethyl ester;
5-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl]-pentanoic acid;
5-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-pentanoic acid methyl ester;
1-(2-Chloro-ethyl)-3-(3-hexyl-phenyl)-urea;
1-(2-Chloro-ethyl)-3-(3-hexyl-phenyl)-urea;
6-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl]-hexanoic acid ethyl ester;
6-{3-[3-(2-Chloro-ethyl)-ureido)-phenyl}-hexanoic acid;
6-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl]-hexanoic acid methyl ester;
3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid methyl ester;
1-(2-Chloro-ethyl)-3-[3-(4-hydroxy-but-1-ynyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3 -[3 -(3-hydroxy-prop-1-ynyl)-phenyl]-urea;
1-(2-Chloro-ethyl)-3-(3-cyanomethyl-phenyl)-urea;
2- {3-[3-(2-Chloro-ethyl)-ureido]-phenyl-acetamide;
3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-acetic acid ethyl ester;
Acetic acid 3-{3-[3-(2-chloro-ethyl)-ureido]-phenyl]-propyl ester.
Compounds of Formula I wherein X is Br or I may undergo rearrangement to
provide a
rearrangement product. Such rearrangement products are considered to be within
the scope of the
present invention. Thus, the present invention contemplates the compounds of
Formula I wherein
X is Br or I as a form of pro-drugs, for which both the pro-drug form and the
rearrangement
product may have activity in inhibiting proliferation of cells.
As noted supra, this invention includes the pharmaceutically acceptable salts
of the compounds
defined by Formula I, II, III, IV or V. A compound of this invention can
possess a sufficiently
acidic, a sufficiently basic, or both functional groups, and accordingly react
with any of a number
of organic and inorganic bases, and inorganic and organic acids, to form a
pharmaceutically
acceptable salt.
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CA 02568609 2006-11-28
The term "pharmaceutically acceptable salt," as used herein, refers to salts
of the compounds of
the above formulae, which are substantially non-toxic to living organisms.
Typical
pharmaceutically acceptable salts include those salts prepared by reaction of
the compounds of
the present invention with a pharmaceutically acceptable mineral or organic
acid or an organic or
inorganic base. Such salts are known as acid addition and base addition salts.
Acids commonly employed to form acid addition salts are inorganic acids such
as hydrochloric
acid, hydrobromic acid, hydriodic acid, sulfuric acid, phosphoric acid, and
the like, and organic
acids such as p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-
bromophenylsulfonic
acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid,
and the like. Examples of
such pharmaceutically acceptable salts are the sulfate, pyrosulfate,
bisulfate, sulfite, bisulfate,
phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate,
pyrophosphate,
chloride, bromide, iodide, acetate, propionate, decanoate, caprylate,
acrylate, formate,
hydrochloride, dihydrochloride, isobutyrate, caproate, heptanoate, propiolate,
oxalate, malonate,
succinate, suberate, sebacate, fumarate, maleate, butyne-1,4-dioate, hexyne-
1,6-dioate, benzoate,
chlorobenzoate, methylbenzoate, hydroxyberazoate, methoxyberazoate, phthalate,
xylenesulfonate,
phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, y-
hydroxybutyrate, glycolate,
tartrate, methanesulfonate, propanesulfonate, naphthalene-1-sulfonate,
napththalene-2-sulfonate,
mandelate and the like.
In one embodiment of the invention, the pharmaceutically acceptable acid
addition salts are those
formed with mineral acids such as hydrochloric acid and hydrobromic acid, and
those formed
with organic acids such as malefic acid and methanesulfonic acid.
Salts of amine groups may also comprise quarternary ammonium salts wherein the
amino
nitrogen carries a suitable organic group such as an alkyl, allcenyl, alkynyl,
or aralkyl moiety.
Base addition salts include those derived from inorganic bases, such as
ammonium or alkali or
alkaline earth metal hydroxides, carbonates, bicarbonates, and the like. Such
bases useful in
preparing the salts of this invention thus include sodium hydroxide, potassium
hydroxide,
ammonium hydroxide, potassium carbonate, sodium carbonate, sodium bicarbonate,
potassium
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CA 02568609 2006-11-28
bicarbonate, calcium hydroxide, calcium carbonate, and the like. In one
embodiment, the base
addition salt is a potassium or sodium salt.
It should be recognized that the particular counterion forming a part of any
salt of this invention
is usually not of a critical nature, as long as the salt as a whole is
pharmacologically acceptable
and as long as the counterion does not contribute undesired qualities to the
salt as a whole.
This invention further encompasses the pharmaceutically acceptable solvates of
the compounds
of Formula I, II, III, IV or V. Many of these compounds can combine with
solvents such as
water, methanol, ethanol and acetonitrile to form pharmaceutically acceptable
solvates such as
the corresponding hydrate, methanolate, ethanolate and acetonitrilate.
The compounds of the present invention have multiple asymmetric (chiral)
centers. As a
consequence of these chiral centers, the compounds of the present invention
occur as racemates,
mixtures of enantiomers and as individual enantiomers, as well as
diastereomers and mixtures of
diastereomers. All asymmetric forms, individual isomers and combinations
thereof, are within
the scope of the present invention.
The prefixes "R" and "S" are used herein as commonly used in organic chemistry
to denote the
absolute configuration of a chiral center, according to the Calm-Ingold-Prelog
system. The
stereochemical descriptor R (rectus) refers to that configuration of a chiral
center with a
clockwise relationship of groups tracing the path from highest to second-
lowest priorities when
viewed from the side opposite to that of the lowest priority group. The
stereochemical descriptor
S (sinister) refers to that configuration of a chiral center with a
counterclockwise relationship of
groups tracing the path from highest to second-lowest priority when viewed
from the side
opposite to the lowest priority group. The priority of groups is decided using
sequence rules as
described by Cahn et al., Aiagew. Chem., 7~, 413-447, 1966 and Prelog, V. and
Helmchen, G.;
Aragew. Claem. Irat. Ed. Esag., 21, 567-5~3, 192).
In addition to the R,S system used to designate the absolute configuration of
a chiral center, the
older D-z system is also used in this document to denote relative
configuration, especially with
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CA 02568609 2006-11-28
reference to amino acids and amino acid derivatives. In this system a Fischer
projection of the
compound is oriented so that carbon-1 of the parent chain is at the top. The
prefix "D" is used to
represent the relative configuration of the isomer in which the functional
(determining) group is
on the right side of the carbon atom at the chiral center and "L", that of the
isomer in which it is
on the left.
As would be expected, the stereochemistry of the Formula I II, III, IV and V
compounds may be
critical to their potency as agonists or antagonists. The relative
stereochemistry is preferably
established early during synthesis, which avoids stereoisomer separation
problems later in the
process. Subsequent synthetic steps then employ stereospecific procedures so
as to maintain the
preferred configuration.
Non-toxic metabolically labile esters and amides of compounds of Formula I II,
III, IV or V are
ester or amide derivatives that are hydrolyzed ih vivo to afford said
compounds of Formula I II,
III, IV or V and a pharmaceutically acceptable alcohol or amine. Examples of
metabolically
labile esters include esters formed with (C1-C6) alkanols in which the alkanol
moiety may be
optionally substituted by a (C1-C8) alkoxy group, for example, methanol,
ethanol, propanol and
methoxyethanol. Examples of metabolically labile amides include amides formed
with amines
such as methylamine.
The>"apeutic Uses of Compounds of Formula (I)
The compounds of Formula I can be used to attenuate, inhibit and/or prevent
non-cancerous
cellular proliferation in a patient in need of such therapy. The compounds of
the invention are
also for use in the treatment of diseases and disorders associated with non-
cancerous pathogenic
cellular proliferation, the pathogenesis of which, may include cell migration
and/or
inflammation. The compounds can be used alone or they can be used as part of a
mufti-drug
regimen in combination with known therapeutics.
In one embodiment, the compounds of Formula I can be used to attenuate,
inhibit or prevent non-
cancerou~ cell proliferation of keratinocytes. In another embodiment diseases
or disorders that
can be treated with the compounds of Formula I include those associated with
hyperproliferation
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CA 02568609 2006-11-28
and/or migration of keratinocytes and/or epidermal or epithelial cells.
Typically, such diseases
and disorders are dermatological and include psoriasis, eczema, lupus-
associated skin lesions,
dermatitides (such as seborrheic dermatitis and solar dermatitis), keratoses
(such as seborrheic
keratosis, senile keratosis, actinic keratosis, photo-induced keratosis, and
keratosis follicularis),
scars and prophylaxis against scar formation (including hypertrophic scars),
acne vulgaris,
keloids and prophylaxis against keloid formation, nevi, warts (including
verruca, condyloma or
condyloma acuminatum), human papilloma viral (HPV) infections (such as
venereal warts),
leukoplakia, lichen planus, and keratitis.
In one embodiment, the compounds of Formula I can be used to attenuate,
inhibit or prevent non-
cancerous cell proliferation of fibroblasts. In another embodiment, the
compounds of the
invention are also useful for the treatment of fibrotic disorders such as
arteriosclerotic conditions,
fibrosis and other medical complications of fibrosis, which result in whole or
in part from the
proliferation of fibroblasts. An arteriosclerotic condition refers to
classical atherosclerosis,
accelerated atherosclerosis, atherosclerotic lesions and other
arteriosclerotic conditions
characterized by undesirable endothelial and/or vascular smooth muscle cell
proliferation,
including vascular complications of diabetes. Proliferation of vasculax smooth
muscle cells
produces accelerated atherosclerosis, which is the main reason for failure of
heart transplants that
are not rejected. The compounds of Formula I can, therefore, be used to
inhibit such obstruction
and reduce the risk of, or even prevent, such failures.
In one embodiment, the compounds of Formula I can be used to attenuate,
inhibit or prevent non-
cancerous cell proliferation of endothelial and smooth muscle cells.
Proliferation of endothelial
and vascular smooth muscle cells is the main feature of neovascularization.
Abnormal
neovascularization has long been associated with solid tumour growth and
metastasis.
Accordingly, in one embodiment of the present invention, the compounds are
used to attenuate,
inhibit or prevent angiogenesis associated with cancer. Abnormal
neovascularization also plays
a pivotal role in a variety of other diseases and disorders. In another
embodiment, the compounds
of Formula I can be used in the treatment of such diseases and disorders,
which include,
rheumatoid arthritis, psoriatic arthritis, diabetic retinopathy, diabetic
glomerulosclerosis,
CA 02568609 2006-11-28
neovascular glaucoma, macular degeneration, Crohn's disease, endometriosis,
psoriasis and
atherosclerosis.
In another embodiment, the compounds of Formula I can be used to treat
vascular injury which
is associated with endothelial and vascular smooth muscle cell proliferation.
The injury can be
caused by a number of traumatic events or interventions, including vascular
surgery and balloon
angioplasty. Restenosis is the main complication of successful balloon
angioplasty of the
coronary arteries. Thus, by inhibiting unwanted endothelial and smooth muscle
cell proliferation,
the compounds of the present invention can be used to delay, or even avoid,
the onset of
restenosis.
In another embodiment, the compounds of the invention are also useful for the
treatment of
inflammatory dermatosis, rosacea, prerosacea. Another example of an
angiogenesis associated
disorder is Rosacea. Rosacea is a common, chronic, progressive inflammatory
dermatosis based
upon vascular instability. It primarily affects the central part of the face.
Rosacea is characterized
by facial flushing/blushing, facial erythema, papules, pustules, and
telangiectasia.
In one embodiment, the compounds of Formula I can be used to attenuate,
inhibit or prevent non-
cancerous cell proliferation of endothelial cells. In another embodiment, the
compounds of
Formula I can be used to treat angiogenesis-associated diseases and disorders.
As is known in
the art, endothelial cell proliferation and/or migration are key features of
neovascularisation and
angiogenesis. Thus, one embodiment of the invention contemplates the use of
the compounds in
the treatment of angiogenesis-associated diseases and disorders. As used
herein, the terms
angiogenesis-associated disease and angiogenesis-associated disorder refer to
a disease or
disorder which occurs as a consequence of, or which results in, increased
vascularization in a
tissue. Angiogenesis may be an actual cause of the disease or a condition
resulting from of the
disease. Examples of angiogenesis-associated diseases and disorders include,
but are not limited
to, immune disorders such as inflammation, chronic articular rheumatism and
psoriasis, disorders
associated with inappropriate or inopportune invasion of vessels such as
diabetic retinopathy,
neovascular glaucoma, restenosis, capillary proliferation in atherosclerotic
plaques and
osteoporosis, cancer, cancer associated disorders, such as solid tumors, solid
tumor metastases,
41
CA 02568609 2006-11-28
angiofibromas, retrolental fibroplasia, hemangiomas, Kaposi sarcoma ,
psoriasis, acne, rosacea,
warts, eczema, hemangiomas, lymphangiogenesis, Sturge-Weber syndrome,
neurofibromatosis,
tuberous sclerosis, chronic inflammatory disease, arthritis, chronic articular
rheumatism and
psoriasis. Skin disorders which have as a primary or secondary
characterisation, increased
vascularization, are considered to be angiogenesis-associated disorders for
purposes of the
present invention.
Thus, methods which inhibit angiogenesis in a diseased tissue ameliorates
symptoms of the
disease and, depending upon the disease, can contribute to cure of the
disease. The compounds of
the present invention are suitable for use in a variety of tissues that can be
invaded by blood
vessels upon angiogenic stimuli including skin, muscle, gut, connective
tissue, joints, bones and
the like.
In one embodiment the compounds of Formula I can be used to attenuate, inhibit
or prevent IL-2
production in order to ameliorate inflammation associated with angiogenesis-
associated diseases
and disorders, as well as inflammatory conditions not associated with
angiogenesis. The present
invention, therefore, further contemplates that the compounds can be used in
the treatment of
diseases such as, inflammatory arthritis (including rheumatoid arthritis),
psoriasis, rheumatism.
Efficacy of the Therapeutic Compounds
In accordance with the present invention, the therapeutic compounds of Formula
I are capable of
attenuating, inhibiting, or preventing non-cancerous cellular proliferation
ifa vivo. One skilled in
the art will appreciate that compounds within Formula I will demonstrate
different activities in
their ability to attenuate, inhibit, or prevent non-cancerous cellular
proliferation and to treat the
diseases associated with such proliferation, the pathogenesis of which, may
include the cellular
migration and/or inflammation. The ability of the compounds to attenuate,
inhibit, or prevent
non-cancerous cellular proliferation, cellular migration and/or inflammation
can be initially
determined in vitro if desired. The present invention thus contemplates a
preliminary in vitro
42
CA 02568609 2006-11-28
screening step to further characterize compounds suitable for incorporation
into the therapeutic
compositions. A number of standard tests to determine the ability of a
compound to attenuate,
inhibit, or prevent proliferation, cellular migration and/or inflammation are
known in the art and
can be employed to test the compounds of Formula I. Exemplary procedures are
described
herein.
Functional Assays
Candidate compounds of Formula I can be tested in vitro and ira vivo to
determine their activity
in inhibiting non-cancerous cellular proliferation and/or cell migration. The
compounds can also
be tested for their ability to inhibit production of pro-inflammatory
cytokines, such as IL-2.
1. In vitro Testirag
Cell Proliferation Assays
The compounds can be assayed initially for their ability to inhibit the growth
of cells, such as
endothelial cells or keratinocytes, (i. e. their cytotoxicity) in vitro using
standard techniques. In
general, cells of a specific test cell line are grown to an appropriate
density (e.g. approximately 1
x 104) and the candidate compound is added. After an appropriate incubation
time (for example
48 to 74 hours), the cell density is assessed. Methods of measuring cell
density are known in the
art, for example, the cell density can be assessed under a light inverted
microscope by measuring
the surface of the culture plate covered by the cell monolayer; or by using
the resazurin reduction
test (see Fields ~z. Lancaster (1993) Am. Biotechnol. Lab. 11:48-50; O'Brien
et al., (2000) Eur. J.
Bioclaena. 267:5421-5426 and U.S. Patent No. 5,501,959), the sulforhodamine
assay (Rubinstein
et al., (1990) J. Natl. Cancer Inst. 82:113-118) or the neutral red dye test
(Kitano et al., (1991)
Euro. J. Clin. Investg. 21:53-58; West et al., (1992) J. Investigative l7erm.
99:95-100).
Alternatively, the cells can be detached from the plate, for example, by
incubation with trypsin
and then counted in an hemocytometer. Percent inhibition of proliferation of
the cells is
calculated by comparison of the cell density in the treated culture with the
cell density in control
cultures, for example, cultures not pre-treated with the candidate compound
and/or those pre-
treated with a control compound (typically a known therapeutic). Cells may be
treated with a
43
CA 02568609 2006-11-28
mitogen prior to addition of the candidate compound to assess the ability of
the compounds to
inhibit proliferation of stimulated cells as opposed to unstimulated, or
quiescent cells.
DNA synthesis can be also assessed and used as an indication of cell
proliferation. For example,
by the uptake of [3H]thymidine. Typically cells are grown to an appropriate
density (generally to
confluence) at which point the growth medium is replaced with a medium that
renders the cells
quiescent (for example, DME 0.5% serum). The quiescent cells are exposed to a
mitogenic
stimulus, such as 10% serum, PDGF, bFGF, or other appropriate mitogen
according to the cell
line, at a suitable interval after the medium replacement. [3H]thymidine is
subsequently added to
the cells, and the cells are maintained at 37°C. After an appropriate
incubation time, the cells are
washed, the acid-precipitable radioactivity is extracted and the amount of
radioactivity
determined, for example, by using a scintillation counter.
Cell Mig~atiofz Assays
In general, the ability of a compound to inhibit migration of cells, such as
endothelial cells and
keratinocytes, can be assessed ifZ vitf o using standard cell migration
assays. Typically, such
assays are conducted in multi-well plates, the wells of the plate being
separated by a suitable
membrane into top and bottom sections. The membrane can be coated with an
appropriate
compound, the selection of which is dependent on the type of cell being
assessed and can be
readily determined by one skilled in the art. Examples include but are not
limited to fibronectin,
collagen, gelatine or Matrigel. An appropriate chemo-attractant, the selectin
of which is
dependent on the type of cell being assessed and can readily be determined by
one skilled in the
art. Examples include but are not limited to EGM-2, IL-8, a-FGF, (3-FGF, EGF
and the like, is
added to the bottom chamber as a chemo-attractant. An aliquot of the test
cells together with the
test compound is added to the upper chamber, typically various dilutions of
the test compound
are tested. After a suitable incubation time, the membrane is rinsed, fixed
and stained. The cells
on the upper side of the membrane are wiped off, and then randomly selected
fields on the
bottom side are counted. Cell (for example, keratinocyte or endothelial cell)
migration can also
be assessed iya vitro by monitoring the closure of a denuded area scratched in
a confluent
monolayer in the presence or absence of test compound (see Phan et al., Wound
Repair Regen
9(4):305-313).
44
CA 02568609 2006-11-28
The migration and differentiation of endothelial cells during angiogenesis can
also be studied in
vitro using the Matrigel tube formation assay (see Grant et al., (1992) J.
Cell. Physiol.153:614).
In general, cell culture plates are coated with a Matrigel solution and then
incubated at 37°C to
promote gelling. HWECs or other suitable cells are resuspended in growth media
and added to
each well. The candidate compound, positive control compounds) (for example,
aFGF and/or
bFGF) and/or media alone (as a second control) are also added to the wells at
this time. After an
appropriate incubation time, the plates are fixed and the length of the tubes
measured by
microscopy.
Examples of suitable cell lines to assess the anti-angiogenic properties of
candidate compounds
include, but are not limited to, human umbilical vein endothelial cells
(HUVECs), bovine aortic
endothelial cells (BAECs), human coronaxy artery endothelial cells (HCAECs)
and vascular
smooth muscle cells. HUVECs can be isolated from umbilical cords using
standard methods
(see, for example, Jaffe et al. (1973) J. Clifa. Ihvest. 52: 2745), or they
can be obtained from the
ATCC or various commercial sources, as can other suitable endothelial cell
lines. Suitable cell
lines to assess the anti-psoriasis properties of candidate compounds include,
but are not limited
to, keratinocytes (e.g. HaCat cells). The use of proliferating keratinocytes
in culture as a test
system for determining the utility of a compound for treating psoriasis is
well documented (see,
for example, Kitano et al., (1991) Euro. J. Clin. Investg. 21:53-58; West et
al., (1992) J.
Investigative Deem. 99:95-100).
Cytokine Assays
A number of assays, including but not limited to enzyme immunoassay systems
such as ELISA,
ELISPOT, are known in the art to measure cytokine concentrations in a sample.
Accordingly, in
one embodiment of the invention, the ability of a candidate compound to
inhibit IL-2 production
in cells is determined by treating a T cell culture with candidate compound
and quantifying IL-2
production using an IL-2 ELISA and comparing to control treated cultures.
Alternatively, IL-2
concentration can be examined in samples, including but not limited to serum,
isolated from
mammals treated with the compounds of the invention.
2. In vivo Testing
CA 02568609 2006-11-28
A number of assays are known in the art for testing the ability of candidate
compounds to inhibit
angiogenesis in vivo. For example, the ability of the candidate compounds to
inhibit endothelial,
cell migration can be determined using the chick chorioallantoic membrane
(CAM) assay,
Matrigel plug assay and/or corneal micropocket assay. In addition, the ability
of the
compositions to inhibit psorasis can be assessed using various marine models
of psorasis.
The CAM assay is a standard assay that is used to evaluate the ability of a
test compound to
inhibit the growth of blood vessels into various tissues, i.e. both
angiogenesis and
neovascularization (see Brooks et al., in Methods in Molecular Biology, Vol.
129, pp. 257-269
(2000), ed. A.R. Howlett, Humana Press Inc., Totowa, NJ; Ausprunk et al.,
(1975) Am. J.
Pathol., 79:597-618; Ossonski et al., (1980) CafZCer Res., 40:2300-2309).
Since the CAM assay
measures neovascularization of whole tissue, wherein chick embryo blood
vessels grow into the
chorioallantoic membrane (CAM) or into the tissue transplanted on the CAM, it
is a well-
recognised assay model for in vivo angiogenesis.
The Matrigel plug assay is also a standard method for evaluating the anti-
angiogenic properties
of compounds iya vivo (see, for example, Passaniti, et a1.,(1992) Lab. Invest.
67:519-528). In this
assay, a test compound is introduced into cold liquid Matrigel which, after
subcutaneous
injection into a suitable animal model, solidifies and permits penetration by
host cells and the
formation of new blood vessels. After a suitable period of time, the animal is
sacrificed and the
Matrigel plug is recovered, usually together with the adjacent subcutaneous
tissues. Assessment
of angiogenesis in the Matrigel plug is achieved either by measuring
haemoglobin or by scoring
selected regions of histological sections for vascular density, for example by
immunohistochemistry techniques identifying specific factors such as
hemagglutinin (HA),
CD31 (platelet endothelial cell adhesion molecule-1) or Factor VIII.
Modifications of this assay
have also been described (see, for example, Akhtar et al., (2002) Ahgiogenesis
5:75-80; Kragh et
al., (2003) IfZt J Oncol. 22:305-11).
The corneal micropocket assay is usually conducted in mice, rats or rabbits
and has been
described in detail by others (see D'Amato, et al., (1994) Proc. Natl, Acad.
Sci. USA, 91:4082-
4085; Koch et al., (1991) Agents Actiotas, 34:350-7; Kenyon, et al., (1996)
Invest. Ophthalmol.
Vis. Sci. 37:1625-1632). Briefly, pellets for implantation are prepared from
sterile hydron
46
CA 02568609 2006-11-28
polymer containing a suitable amount of the test compound. The pellets are
surgically implanted
into corneal stromal micropockets created at an appropriate distance medial to
the lateral corneal
limbus of the animal. Angiogenesis can be quantitated at various times after
pellet implantation
through the use of stereomicroscopy. Typically, the length of neovessels
generated from the
limbal vessel ring toward the centre of the cornea and the width of the
neovessels are measured.
The efficacy of the compounds of Formula I in the treatment of various
diseases and disorders
associated with angiogenesis can be tested initially in an appropriate animal
model. Accepted
animal models are available for a number of diseases (see, for example, Enna
et al., (Eds.)
Cuf°rent Protocols in Phaf°rnacology, J. Wiley ~ Sons, New
York, NY).
As indicated above, the compounds of the invention can be used in the
treatment of psoriasis. A
number of murine psoriasis models are known in the art and can be used
initially to assess the
ability of the candidate compounds to treat psoriasis. Murine psoriasis models
include immune
comprised mice injected with CD45Rb positive cells (U.S. Patent No 64,10,824).
Alternatively a
human psoriasis xenograft model (Sugai et al. (1998) J Dermatol Sci 17:85-92)
or transgenic
mouse models (Xia et al. Blood 102(1):161-168 can be used.
Additional Tests
In addition to the above tests, the compounds of the invention can be
submitted to other standard
tests, such as cytotoxicity tests, stability tests, bioavailability tests and
the like. As will be readily
apparent to one skilled in the art, the therapeutic compositions of the
invention will need to meet
certain criteria in order to be suitable for human use and to meet regulatory
requirements. Thus,
once a compound of the invention has been found to be suitable for animal
administration,
standard in vitro and in vivo tests can be conducted to determine information
about the
metabolism and pharmacokinetic (PK) of the compositions and combinations
(including data on
drug-drug interactions where appropriate) which can be used to design human
clinical trials.
Clinical Trials
One skilled in the art will appreciate that, following the demonstrated
effectiveness of the
compounds of the present invention in vitro and in animal models (i.e. pre-
clinical efficacy), the
47
CA 02568609 2006-11-28
safety profile of the compounds can be determined in at least two non-human
species and then
the compositions will progress into Clinical Trials in order to further
evaluate their efficacy in
the treatment of non-cancerous pathogenic cellular proliferation and the
diseases and disorders
associated therewith in order to obtain regulatory approval for therapeutic
use. As is known in
the art, clinical trials progress through phases of testing, which are
identified as Phases I, II, III,
and IV. ha vitro and in vivo information about the metabolism and
pharmacokinetic (PK) of the
compounds of the invention determined from pre-clinical studies facilitates
the design of initial
Phase I and Phase II clinical studies.
Phase I
These studies are conducted to investigate the safety, tolerability and PK of
the compounds and
to help design Phase II studies, for example, in terms of appropriate doses,
routes of
administration, administration protocols.
Phase II
Phase I studies allow the selection of safe dose levels for Phase II studies.
An important factor in
the protocol design of the Phase II studies is the adequate recruitment of the
patient population to
be studied based on stringent selection criteria defining the demographics
(age, race and sex) of
the study and the previous medical history of the patient. A protocol for
Phase II studies
typically specifies baseline data that can be used to characterise the
population, to evaluate the
success of randomization in achieving balance of important prognostic factors,
and to allow for
consideration of adjusted analyses.
Clinical biomarkers can be defined as follows (Atkinson A et al.' Clin.
Pharmacol. Ther. 69, ~9-
95 (2001):
Biological maYker (bioma~ker): a characteristic that is objectively measured
and evaluated as an
indicator of normal biological process, pathogenic process, or pharmacological
response to a
therapeutic intervention.
Clinical endpoint: a characteristic or variable that reflects how a patient
feels or functions, or
how long a patient survives.
48
CA 02568609 2006-11-28
Surrogate endpoint: biomarker intended to substitute for a clinical endpoint.
A clinical
investigator uses epidemiological, therapeutic, pathophysiological, or other
scientific evidence to
select a surrogate endpoint that is expected to predict benefit, harm or the
lack of benefit or
harm. The FDA defines a surrogate endpoint, or marker, as a laboratory
measurement or physical
sign that is used in therapeutic trials as a substitute for a clinically
meaningful endpoint that is a
direct measure of how a patient feels, functions or survive and is expected to
predict the effect of
the therapy.
Phase III
Phase III trials focus on determining how the compound compares to the
standard, or most
widely accepted, treatment. In Phase III trials, patients are randomly
assigned to one of two or
more "arms". In a trial with two arms, for example, one arm will receive the
standard treatment
(control group) and the other arm will be treated with the therapeutic
composition/combination
(investigational group).
Phase IV
Phase IV trials can be used to further evaluate the long-term safety and
effectiveness of the
compound. Phase IV trials are less common than Phase I, II and III trials and
would take place
after the therapeutic composition has been approved for standard use.
Preparation of Compounds of Formula I
In one embodiment of the present invention, compounds of Formula I wherein Rl
and R2 are
both H are prepared by the general method provided below.
R~ R2 ~ \~ /R2
B-NHZ+ O=C=N-CCHZCI -~ B-N N-CCHzCI
H H
VI VII I
49
CA 02568609 2006-11-28
An amine derivative of Formula IV wherein B is as defined for compound of
formula (I) is
reacted with an isocyanate of Formula V in anhydrous ether under nitrogen
atmosphere at room
temperature for 2 to 16 hours or until the disappearance on thin layer
chromatography (TLC) of
the starting amine of Formula II. The solid residue obtained is filtered and
dried in vacuo, the
filtrate is evaporated under reduced pressure, and the solid remaining therein
is also dried in
vacuo. Both solids are independently recrystallized with suitable organic
solvents and the final
crystals are pooled after evaluation of their purity by chromatography, IR, 1H
NMR and MS.
Where the crystallization is difficult, purification by column chromatography
on silica gel may
be required prior to the final crystallization. The compounds of Formula IV
are either
commercially available or can be prepared with the standard procedures known
to a worker
skilled in the relevant art.
In another embodiment of the invention, compounds of formula I, wherein B is a
substituted
phenyl, R~ and/or RZ are other than H and X is Cl, are prepared by the general
method provided
below.
H H H H
NH2 ~ N N N N
yOH c ~ ~ ~CI
O R1 AR2 ~ I R ~'R2
O 1
R R
1 1 R1
VI~~
a = Boc20, DMAP, CH2Cla; b = (R) or (S) NH2CRaR3CHZOH, CH2Cla; c = PPh3, CC14
/ CH2C12,
Formation of the urea moiety is achieved using the 4-dimethylaminopyridine-
catalyzed reaction
of the relevant 4-alkylaniline (VI) with di-tef°t-butyldicarbonate in
dichloromethane followed by
the trapping of the ira situ generated isocyanate with the appropriate (R)- or
(S)-2-aminoalcohol
(see Knolker, et al., Synlett (1996) 502-504 and Journal ofMedicinal
Chemistry, 46, 2003, 5055-
5063). This procedure ensures a racemization-free synthesis of urea under mild
conditions and
circumvents side reactions such as the formation of symmetrical disubstituted
urea (see I~nolker,
et al., Synlett (1997) 925-928). Subsequently, chloration of the chiral 2-
hydroxyethylureas (VII)
is achieved using triphenylphosphine in a mixture of carbon tetrachloride and
dichloromethane at
CA 02568609 2006-11-28
room temperature, affording the final enantiomerically pure (R)- or (~-(1-
alkyl-2-
chloro)ethylurea derivative.
Other methods of preparing compounds of Formula I are known and can be readily
employed by
one skilled in the art to obtain the compounds of the invention. Certain
compounds of Formula I
are also available commercially. Further exemplary methods of preparing the
compounds are
provided by the Examples. These methods are provided by means of example only
and are not
intended to limit the scope of the invention in any way.
Administration of Therapeutic Compounds and Pharamceutical Compositions
The present invention provides methods of attenuating, inhibiting or
preventing non-cancerous
pathogenic cellular proliferation in a mammal comprising administering an
effective amount of
one or more compounds of Formula I, or non-toxic metabolically-labile esters
or amides thereof,
or pharmaceutically acceptable salts thereof. Accordingly, the present
invention provides
methods for the treatment of diseases and disorders associated with non-
cancerous pathogenic
cellular proliferation, the pathogenesis of which, may include cell migration
and/or
inflammation. In one embodiment of the invention, methods for the treatment of
diseases and
disorders characterised by endothelial cell proliferation and/or migration,
keratinocyte
proliferation and/or migration, inflammation, or combinations thereof are
provided. In another
embodiment of the invention, methods for the amelioration of inflammation
associated with IL-2
production are provided.
The compounds of the present invention are typically formulated prior to
administration. The
present invention thus provides pharmaceutical compositions comprising one or
more
compounds of Formula I and a pharmaceutically acceptable caxrier, diluent, or
excipient. The
pharmaceutical compositions are prepared by known procedures using well-known
and readily
available ingredients.
Compounds of the general Formula I or pharmaceutical compositions comprising
the compounds
may be administered orally, topically, parenterally, by inhalation or spray,
or rectally in dosage
unit formulations containing conventional non-toxic pharmaceutically
acceptable carriers,
51
CA 02568609 2006-11-28
adjuvants and vehicles. In the usual course of therapy, the active compound is
incorporated into
an acceptable vehicle to form a composition for topical administration to the
affected area, such
as hydropohobic or hydrophilic creams or lotions, or into a form suitable for
oral, rectal or
parenteral administration, such as syrups, elixirs, tablets, troches,
lozenges, hard or soft capsules,
pills, suppositiories, oily or aqueous suspensions, dispersible powders or
granules, emulsions,
injectables, or solutions. The term parenteral as used herein includes
subcutaneous injections,
intradermal, intra-articular, intravenous, intramuscular, intravascular,
intrasternal injection or
infusion techniques.
Compositions intended for oral use may be prepared in either solid or fluid
unit dosage forms.
Fluid unit dosage form can be prepared according to procedures known in the
art for the
manufacture of pharmaceutical compositions and such compositions may contain
one or more
agents selected from the group consisting of sweetening agents, flavouring
agents, colouring
agents and preserving agents in order to provide pharmaceutically elegant and
palatable
preparations. An elixir is prepared by using a hydroalcoholic (e.g., ethanol)
vehicle with suitable
sweeteners such as sugar and saccharin, together with an aromatic flavoring
agent. Suspensions
can be prepared with an aqueous vehicle with the aid of a suspending agent
such as acacia,
tragacanth, methylcellulose and the like.
Solid formulations such as tablets contain the active ingredient in admixture
with non-toxic
pharmaceutically acceptable excipients that are suitable for the manufacture
of tablets. These
excipients may be for example, inert diluents, such as calcium carbonate,
sodium carbonate,
lactose, calcium phosphate or sodium phosphate: granulating and disintegrating
agents for
example, corn starch, or alginic acid: binding agents, for example starch,
gelatin or acacia, and
lubricating agents, for example magnesium stearate, stearic acid or talc and
other conventional
ingredients such as dicalcium phosphate, magnesium aluminum silicate, calcium
sulfate, starch,
lactose, methylcellulose, and functionally similar materials. The tablets may
be uncoated or they
may be coated by known techniques to delay disintegration and absorption in
the gastrointestinal
tract and thereby provide a sustained action over a longer period. For
example, a time delay
material such as glyceryl monostearate or glyceryl distearate may be employed.
52
CA 02568609 2006-11-28
Formulations for oral use may also be presented as hard gelatin capsules
wherein the active
ingredient is mixed with an inert solid diluent, for example, calcium
carbonate, calcium
phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient
is mixed with water
or an oil medium, for example peanut oil, liquid paraffin or olive oil. Soft
gelatin capsules are
prepared by machine encapsulation of a slurry of the compound with an
acceptable vegetable oih
light liquid petrolatum or other inert oil.
Aqueous suspensions contain active materials in admixture with excipients
suitable for the
manufacture of aqueous suspensions. Such excipients are suspending agents, for
example
sodium carboxylmethylcellulose, methyl cellulose, hydropropylmethylcellulose,
sodium alginate,
polyvinylpyrrolidone, gum tragacanth and gum acacia: dispersing or wetting
agents may be a
naturally-occurnng phosphatide, for example, lecithin, or condensation
products of an allcylene
oxide with fatty acids, for example polyoxyethylene stearate, or condensation
products of
ethylene oxide with long chain aliphatic alcohols, for example hepta-
decaethyleneoxycetanol, or
condensation products of ethylene oxide with partial esters derived from fatty
acids and a hexitol
such as polyoxyethylene sorbitol monooleate, or condensation products of
ethylene oxide with
partial esters derived from fatty acids and hexitol anhydrides, for example
polyethylene sorbitan
monooleate. The aqueous suspensions may also contain one or more
preservatives, for example
ethyl, or ra-propyl- p-hydroxy benzoate, one or more colouring agents, one or
more flavouring
agents or one or more sweetening agents, such as sucrose or saccharin.
Oily suspensions may be formulated by suspending the active ingredients in a
vegetable oil, for
example peanut oil, olive oil, sesame oil or coconut oil, or in a mineral oil
such as liquid paraffin.
The oily suspensions may contain a thickening agent, for example beeswax, hard
paraffin or
cetyl alcohol. Sweetening agents such as those set forth above, and flavouring
agents may be
added to provide palatable oral preparations. These compositions may be
preserved by the
addition of an anti-oxidant such as ascorbic acid.
Dispersible powders and granules suitable for preparation of an aqueous
suspension by the -
addition of water provide the active ingredient in admixture with a dispersing
or wetting agent,
suspending agent and one or more preservatives. Suitable dispersing or wetting
agents and
53
CA 02568609 2006-11-28
suspending agents are exemplified by those already mentioned above. Additional
excipients, for
example sweetening, flavouring and colouring agents, may also be present.
Pharmaceutical compositions of the invention may also be in the form of oil-in-
water emulsions.
The oil phase may be a vegetable oil, for example olive oil or peanut oil, or
a mineral oil, for
example liquid paraffin or mixtures of these. Suitable emulsifying agents may
be naturally-
occurring gums, for example gum acacia or gum tragacanth, naturally-occurring
phosphatides,
for example soy bean, lecithin, and esters or partial esters derived from
fatty acids and hexitol,
anhydrides, for example sorbitan monooleate, and condensation products of the
said partial esters
with ethylene oxide, for example polyoxyethylene sorbitan monooleate. The
emulsions may also
contain sweetening and flavoring agents.
The pharmaceutical compositions may be in the form of a sterile injectable
aqueous or
oleaginous suspension. This suspension may be formulated according to known
art using those
suitable dispersing or wetting agents and suspending agents that have been
mentioned above.
The sterile injectable preparation may also be a sterile injectable solution
or a suspension in a
non-toxic parentally acceptable diluent or solvent, for example as a solution
in 1,3-butanediol.
Among the acceptable vehicles and solvents that may be employed are water,
Ringer's solution
and isotonic sodium chloride solution. In addition, sterile, fixed oils are
conventionally
employed as a solvent or suspending medium. For this purpose any bland fixed
oil may be
employed including synthetic mono- or diglycerides. In addition, fatty acids
such as oleic acid
find use in the preparation of injectables. Adjuvants such as local
anaesthetics, preservatives and
buffering agents can also be included in the injectable solution or
suspension.
The compounds) of the general Formula I may be administered, together or
separately, in the
form of suppositories for rectal administration of the drug. These
compositions can be prepared
by mixing the drug with a suitable non-irritating excipient which is solid at
ordinary
temperatures but liquid at the rectal temperature and will therefore melt in
the rectum to release
the drug. Such materials include cocoa butter and polyethylene glycols.
54
CA 02568609 2006-11-28
For ophthalmic applications, such as treatment of keratitis, the compounds can
be formulated
into solutions, suspensions, and ointments appropriate for use in the eye
(see, for example, Mitra
(ed.), (1993) Ophthalmic Drug Delivery Systems, Marcel Dekker, Inc., New York,
N.Y.;
Havener, (1983) Ocular Pharmacology, C.V. Mosby Co., St. Louis).
Other pharmaceutical compositions and methods of preparing pharmaceutical
compositions are
known in the art and are described, for example, in "Remington: The Science
and Practice of
Pharmacy" (formerly "Remingtons Plaarrnaceutical Sciences"); Gennaro, A.,
Lippincott,
Williams & Wilkins, Philidelphia, PA (2000).
In general, the route of administration of the compounds of Formula I is
topical (including
administration to the eye, scalp, and mucous membranes), oral, or parenteral.
Topical
administration is usually most effective for the treatment of skin lesions,
including lesions of the
scalp, lesions of the cornea (keratitis), and lesions of mucous membranes
where such direct
application is practical. Shampoo formulations can be advantageous for
treating scalp lesions,
such as seborrheic dermatitis and psoriasis of the scalp, and mouthwash and
oral paste
formulations can be advantageous for mucous membrane lesions, such as oral
lesions and
leukoplakia. Oral administration is an alternative for treatment of skin
lesions and other lesions
discussed above where direct topical application is not as practical, as well
as for other
applications. The present invention contemplates the administration of one or
more compounds
of Formula I either alone or in combination with other therapeutics.
The dosage to be administered is not subject to defined limits, but it will
usually be an effective
amount. It will usually be the equivalent, on a molar basis of the
pharmacologically active free
form produced from a dosage formulation upon the metabolic release of the
active free drug to
achieve its desired pharmacological and physiological effects. The
compositions may be
formulated in a unit dosage form. The term "unit dosage form" refers to
physically discrete units
suitable as unitary dosages for human subjects and other mammals, each unit
containing a
predetermined quantity of active material calculated to produce the desired
therapeutic effect, in
association with a suitable pharmaceutical excipient. Examples of ranges for
the compounds) in
CA 02568609 2006-11-28
each dosage unit are from about 0.05 to about 100 mg, or more usually, from
about 1.0 to about
30 mg.
Daily dosages of the compounds of the present invention will typically fall
within the range of
about 0.01 to about 100 mg/kg of body weight, in single or divided dose.
However, it will be
understood that the actual amount of the compounds) to be administered will be
determined by a
physician, in the light of the relevant circumstances, including the condition
to be treated, the
chosen route of administration, the actual compound administered, the age,
weight, and response
of the individual patient, and the severity of the patient's symptoms. The
above dosage range is
given by way of example only and is not intended to limit the scope of the
invention in any way.
In some instances dosage levels below the lower limit of the aforesaid range
may be more than
adequate, while in other cases still larger doses may be employed without
causing harmful side
effects, for example, by first dividing the larger dose into several smaller
doses for
administration throughout the day.
Pharmaceutical Kits
The present invention additionally provides for therapeutic kits containing
the therapeutic
combinations for use in the treatment of a subj ect in need of therapy for
attenuating non-cancer
cell hyperproliferation. Individual components of the kit would be packaged in
separate
containers and, associated with such containers, can be a notice in the form
prescribed by a
governmental agency regulating the manufacture, use or sale of pharmaceuticals
or biological
products, which notice reflects approval by the agency of manufacture, use or
sale for human
administration.
When the components of the kit are provided in one or more liquid solutions,
the liquid solution
can be an aqueous solution, for example a sterile aqueous solution. In this
case the container
means may itself be an inhalant, syringe, pipette, eye dropper, or other such
like apparatus, from
which the composition may be administered to a patient or applied to and mixed
with the other
components of the kit.
The components of the lcit may also be provided in dried or lyophilised form
and the kit can
additionally contain a suitable solvent for reconstitution of the lyophilised
components.
56
CA 02568609 2006-11-28
Irrespective of the number or type of containers, the kits of the invention
also may comprise an
instrument for assisting with the administration of the composition to a
patient. Such an
instrument may be an inhalant, syringe, pipette, forceps, measured spoon, eye
dropper or any
such medically approved delivery vehicle.
To gain a better understanding of the invention described herein, the
following examples are set
forth. It should be understood that these examples are for illustrative
purposes only. Therefore,
they should not limit the scope of this invention in any way.
EXAMPLES
The following abbreviations are used in the Examples: EtOAc, ethyl acetate;
THF,
tetrahydrofuran; EtOH, ethanol; TLC, thin layer chromatography; GC, gas
chromatography;
HPLC, high pressure liquid chromatography; m-CPBA, m-chloroperbenzoic acid;
Et20, diethyl
ether; DMSO, dimethyl sulfoxide; DBU, 1,8-diazabicyclo-[5.4.0]undec-7-ene,
MTBE, methyl
t-butyl ether; and FDMS, field desorption mass spectrometry.
EXAMPLE 1: Preparation of 1-(4-tart-butylphenyl)-3-(2-chloroethyl)urea [1]
0
t-Bu ~ ~ NH2 + OC=N-CH2CH2CI . t Bu ~ ~ NH"NH-CH2CH2CI
2-Chloroethyl isocyanate (1.15 equiv.) was slowly added dropwise to a
magnetically stirred and
cooled solution (ice bath) of freshly distilled 4-t-butylaniline (1 equiv.) in
dichloromethane (18
mL of solvent/g of aniline). The ice bath was then removed and the reaction
mixture was stirred
at room temperature for 20 h. After completion of the reaction, the solvent
was removed by
vacuum distillation to give a white solid, which was purified by
recrystallization from
dichloromethane/hexane to obtain 88% of 1-(4-tart-butylphenyl)-3-(2-
chloroethyl)urea.
NMR 1H (CDC13, 300 MHz) 8 7.28 (d, 2H, J = 8.5 Hz), 7,17 (d, 2H, J = 8.5 Hz),
3.52 (m, 4H),
1.27 (s, 9H).
57
CA 02568609 2006-11-28
EXAMPLE 2: Preparation of 1-(2-chloroethyl)-3-(4-cyclohexylphenyl)urea [2]
\ ~ N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-cyclohexyl
aniline was used instead of 4-t-butyl aniline. The final product was
recrystallized from
THF/hexane to obtain 82% yield.
1H NMR (CDCl3 + MeOD, 300 MHz) 8 7.10 (d, 2H, J = 8.4 Hz), 6.98 (d, 2H, J =
8.4 Hz), 3.48
(m, 2H), 3.38 (m, 2H), 2.31 (m, 1H), 1.6-1.7 (m, SH), 1.1-1.3 (m, SH).
EXAMPLE 3: Preparation of 1-(2-chloroethyl)-3-(4-hepthylphenyl)urea [3]
O
N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-heptylaniline
was used instead of 4-t-butyl aniline. The final product was recrystallized
from THF/hexane to
obtain 93% Yield.
1H NMR (CDC13, 300 MHz) b 7.15 (d, 2H, J= 8.4 Hz), 7.01 (d, 2H, J= 8.4 Hz),
3.53 (t, 2H, J=
5.2 Hz), 3.45 (t, 2H, J = 5.2 Hz), 2.46 (t, 2H, ,I = 7.7 Hz), 1.49 (m, 2H),
1.21 (m, 8H), 0.80 (t,
3H, J= 6.7 Hz).
EXAMPLE 4
/O
O
N~N~CI
H H
58
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EXAMPLE 5: Preparation of 1-(2-chloroethyl)-3-(4-iodophenyl)urea [5]
I ~ O
\ I ~ CI
N N~
H H
This compound was prepared according to the process of Example 1, except that
4-iodoaniline
was used instead of 4-t-butyl aniline. The final product was recrystallized
from THF/hexane.
Yield 60%
1H NMR (DMSO, 300 MHz) b 8.80 (s, 1H), 7.53 (d, 2H, J = 8.7 Hz), 7.24 (d, 2H,
J = 8.7 Hz),
6.45 (m, 1H), 3.64 (m, 2H), 3.39 (m, 2H).
EXAMPLE 6: Preparation of 1-(2-chloroethyl)-3-(4-phenoxyphenyl)urea [6]
~ \ o i~ ~o
N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-
phenoxyaniline was used instead of 4-t-butyl aniline. 1H NMR (CDC13, 300 MHz)
b 7.1-7.3 (m,
SH), 6.96 (m, 4H), 3.63 (m, 2H), 3.58 (m, 2H).
EXAMPLE 7: Preparation of 1-(4-benzyloxyphenyl)-3-(2-chloroethyl)urea [7]
i
\ I ° ~ o
\ I N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-
benzyloxyaniline was used instead of 4-t-butyl aniline.
59
CA 02568609 2006-11-28
EXAMPLE 8: Preparation of 1-(biphenyl-4-yl)-3-(2-chloroethyl)urea [8]
i
0
\ I N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-
biphenylamine was used instead of 4-t-butyl aniline. The final product was
recrystallized from
methanol/water. Yield 79%.
1H NMR (CDCl3, 300 MHz) ~ 7.44 (m, 4H), 7.2-7.35 (m, SH), 3.55 (m, 2H), 3.46
(m, 2H).
EXAMPLE 9: Preparation of 1-(2-chloroethyl)-3-(4-hydroxyphenyl)urea [9]
HO ~ O
N~N~CI
H H
This compound was prepared according to the process of Example 1, except that
4-
hydroxyaniline was used instead of 4-t-butyl aniline. The final product was
recrystallized from
THF/hexane. Yield 30%
1H NMR (CDCl3, 300 MHz) 8 7.04 (d, 2H, J= 8.7 Hz), 6.69 (d, 2H, J= 8.7 Hz),
3.53 (m, 2H),
3.44 (t, 2H, J= 5.5 Hz).
EXAMPLE 10: Preparation of N {3-[3-(2-chloroethyl)ureido]phenyl}acetamide [10]
\ O
N ~ N~N~CI
H H H
CA 02568609 2006-11-28
This compound was prepared according to the process of Example 1, except that
3'-
aminoacetanilide was used instead of 4-t-butyl aniline. The final product was
recrystallized from
ethyl acetate/methanol/hexane. Yield 46%.
1H NMR (DMSO, 300 MHz) 8 9.86 (br s, 1H), 8.67 (br s, 1H), 7.66 (br s, 1H),
7.12 (s, 4H), 6.35
(t, 1H, J= 5.7 Hz), 3.65 (t, 2H, J= 6.1 Hz), 3.41 (t, 2H, J= 6.1 Hz), 2.02 (s,
3H).
EXAMPLE 11: Preparation of N Sutyl-3-[3-(2-
chloroethyl)ureido]benzenesulfonamide [1l]
H ~ O
~N%S\ I ~ N~N~OI
O O H H
This compound was prepared according to the process of Example l, except that
3-amino-N-
butylbenzenesulfonmide was used instead of 4-t-butyl aniline. The final
product was
recrystallized from from ethanol/water. Yield 50%.
1H NMR (DMSO, 300 MHz) ~ 9.02 (s, 1H), 7.98 (s, 1H), 7.50 (m, 2H), 7.43 (t,
1H, J= 7.9 Hz),
7.30 (d, 1H, J = 7.9 Hz), 6.48 (t, 1H, J = 5.7 Hz), 3.67 (t, 2H, J = 6.0 Hz),
3.43 (q, 2H, J = 6.0
Hz), 2.72 (q, 2H, J= 6.5 Hz), 1.34 (m, 2H), 1.24 (m, 2H), 0.79 (t, 3H, J= 7.1
Hz).
EXAMPLE 12: Preparation of 1-(2-chloroethyl)-3-[3-(1-hydroxyethyl)phenyl]urea
[12]
0
~ ~ci
N- 'N
H H
OH
This compound was prepared according to the process of Example 1, except that
3-(1-
hydroxyethyl) aniline was used instead of 4-t-butyl aniline. The final product
was purified by
flash chromatography on silica gel (3/2 ethyl acetate/chloroform). Yield 63%
1H NMR (CDC13, 300 MHz) 8 7.1-7.3 (m, 4H), 4.85 (q, 1H, J = 6.6 Hz), 3.63 (m,
2H), 3.58 (m,
2H), 1.46 (d, 3H, J = 6.6 Hz).
61
CA 02568609 2006-11-28
EXAMPLE 13: Preparation of 1-(2-chloroethyl)3-[4-(2-methoxyethyl)phenyl)urea
[13]
O /
O
\ ~ ~ CI
N N~
H H
To a cold (ice bath) suspension of NaH 60% (483 mg, 12.1 mmol) in dry THF (20
mL) was
added a solution of 2-(4-nitrophenyl)ethanol (1.04 g, 6.25 mmol) in dry THF (5
mL). The
mixture was stirred at 0 °C for 10 minutes, then methyl iodide (0.50
mL, 8.0 mmol) was added.
The ice bath was removed and the solution was stirred at room temperature for
24 h. Excess of
NaH was quenched carefully with water and the solution was poured into brine.
The aqueous
phase was extracted with ether (3 times). The organic portions were washed
with brine, dried
over MgS04 and concentrated to give 1-(2-methoxyethyl)-4-nitrobenzene which
was purified by
flash chromatography on silica gel (ether/petroleum ether 1/1).
1-(2-Methoxyethyl)-4-nitrobenzene was dissolved in a mixture of ethanol (5 mL)
and water (0.5
mL) and conc. HCl (0.25 mL). Iron powder was added (140 mg) and the mixture
was refluxed
for 2 hours. The solid was removed by filtration on Celite. The solution was
neutralized with
NaOH 1M (to pH 8) and extracted with ethyl acetate (3 times). The organic
portions were
reunited, washed with brine, dried over KZC03 and concentrated to yield 4-(2-
methoxyethyl)aniline which was purified by flash chromatography (CH2Cl2).
4-(2-methoxyethyl)aniline was then reacted with 2-Chloroethyl isocyanate as
described in
Example 1 to prepare compound 13. The final product was purified by flash
chromatography on
silica gel (ether/petroleum ether 13/7). Yield 84%.
1H NMR (CDCl3, 300 MHz) 8 7.16 (d, 2H, J = 8.5 Hz), 7.11 (d, 2H, J = 8.5 Hz),
3.5-3.6 (m,
6H), 3.34 (s, 3H), 2.81 (t, 2H, J= 6.9 Hz).
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CA 02568609 2006-11-28
EXAMPLE 14: Preparation of 1-(2-chloroethyl)-3-[4-(4-ethoxybutyl)phenyl)urea
[14]
O ~ ( O
N~N~CI
H H
To a suspension of powdered potassium hydroxide (2.42 g, 43.1 mmol) in DMSO
(20 mL) was
added a solution of 4-(4-nitrophenyl)butanol (2.09 mg, 10.7 mmol) and ethyl
iodide (3.4 mL, 42
mmol) in DMSO (10 mL) over a 1 h period while the temperature was maintained
below 25 °C.
The resulting mixture was stirred for an additional hour, poured into water
(150 mL), and
extracted with methylene chloride (3 times). The combined organic extracts
were washed with
10% sodium bisulfate (twice), water and brine, dried over MgS04, and
concentrated ih vacuo to
yield 1-(4-ethoxybutyl)-4-nitrobenzene (838 mg, 35%).
A mixture of 1-(4-ethoxybutyl)-4-nitrobenzene (838 mg, 3.75 mmol), iron powder
(1.58 g), and
concd HCl (0.05 rnL) in of a mixture of EtOH, CH3COOH and H2O (2:2:1, 25 mL)
was refluxed
for 4h. The solution was filtered, diluted with H20 (100 mL), and extracted
with CHZCIa (3 x 50
mL). The combined organic layers were washed with a saturated aqueous solution
of NaHC03,
water, and dried over KzC03. Evaporation of the solvent afforded 4-(4-
ethoxybutyl)aniline
which was used without further purification to prepare compound 14 by reacting
with 2-
Chloroethyl isocyanate as described for compound 1 to 13. The final product
was purified by
flash chromatography on silica gel (dichloromethane/ether 9/1). Yield 82%.
1H NMR (CDC13, 300 MHz) 8 7.14 (d, 2H, J = 8.4 Hz), 7.03 (d, 2H, J = 8.4 Hz),
3.3-3.5 (m,
8H), 2.53 (t, 2H, J= 6.8 Hz), 1.60 (m, 4H), 1.18 (t, 3H, J= 6.9 Hz).
EXAMPLE 15: Preparation of 1-(2-chloroethyl)-3-[4-(4-fluorobutyl)phenyl]urea
[15]
O
N~N~CI
H H
63
CA 02568609 2006-11-28
To a cold solution (-12 °C) of 4-(4-nitrophenyl) butanol (788 mg, 4.04
mmol) in dry CHZC12 (8
mL) was slowly added, under nitrogen, bis(2-methoxyethyl)aminosulfur
trifluoride (0.8 mL, 4.3
mmol). The reaction was kept at -12 °C for 15 min and warmed up to room
temperature. The
mixture was stirred for 24 h. The solution was poured into saturated aqueous
NaHC03 (25 mL),
and after COZ evolution ceased it was extracted three times with
dichloromethane. The organic
portions were reunited and washed with brine, dried over Na2S04, filtered, and
evaporated ih
vacuo. Flash chromatography on silica gel (ether/hexane 1/9) afforded 1-(4-
fluorobutyl)-4-
nitrobenzene (490 mg, 62%).
To a solution of 1-(4-fluorobutyl)-4-nitrobenzene (70 mg, 0.36 mmol) in
ethanol (2 mL) was
added SnC~z~2 HZO (409 mg, 1.81 mmol) and the mixture was refluxed for 2
hours. The solution
was cooled and poured into chilled water. The aqueous phase was alkalized by
adding a solution
of NaHC03 and extracted with ethyl acetate (3 times). The organic portions
were reunited,
washed with brine, dried over K2C03 and evaporated in vacuo to afford 4-(4-
fluorobutyl)aniline
which was purified by flash chromatography (CHC13) (53 mg, 88%).
4-(4-fluorobutyl)aniline was then reacted with 2-Chloroethyl isocyanate as
described in
examples 1-12 to obtain compound 15. Purified by flash chromatography on
silica gel
(dichloromethane/ether 85/15). Yield 74%.
1H NMR (CDC13, 300 MHz) 8 7.16 (d, 2H, J= 8.3 Hz), 7.07 (d, 2H, J= 8.3 Hz),
4.43 (dt, 2H, J
= 47.4, 5.5 Hz), 3.53 (m, 4H), 2.58 (t, 2H, J= 7.0 Hz), 1.65-1.75 (m, 4H).
EXAMPLE 16: Preparation of 1-(Z-chloroethyl)-3-(3-(5-hydroxypent-1-
ynyl)phenyl)urea
[16]
\ O
~ SCI
N- _N
HO ~ H H
64
CA 02568609 2006-11-28
3-Iodoaniline (5.05 g, 23.0 mmol), K2CO3 (7.97 g, 57.7 mmol), CuI (183 mg,
0.96 mmol), PPh3
(499 mg, 1.90 mmol), and 10% Pd/C (492 mg, 0.46 mmol Pd) were mixed in 1,2-
dimethoxyethane (30 mL) and water (30 mL) at 25 °C under a nitrogen
atmosphere. The
reaction mixture was stirred for 30 minutes and 4-pentyn-1-of (4.8 mL, 51.6
mmol) was added.
The mixture was heated at 80 °C for 16 hours, cooled to room
temperature and filtered through
Celite. The organic solvents were removed ira vacuo, and the aqueous residue
acidified with 1M
HCI. This solution was extracted with ethyl acetate, and the aqueous phase
basified with KOH.
The water layer was then extracted with ethyl acetate (3X). The organic
portions were reunited,
washed with brine, dried over Na2S04 and concentrated ifz vacuo. 3-(5-
hydroxypent-1-
ynyl)aniline was used for the next reaction without any purification. (5.62
g), which was then
reacted with 2-Chloroethyl isocyanate as described in examples 1-12 to obtain
compound 16.
Purified by flash chromatography on silica gel (chloroform/ethyl acetate 1/1).
Yield 72%.
1H NMR (CDC13, 300 MHz) ~ 7.26 (m, 2H), 7.10 (t, 1H, J= 7.9 Hz), 6.94 (d, 1H,
J= 7.5 Hz),
3.68 (t, 2H, J = 6.3 Hz), 3.55 (m, 2H), 3.47 (m, 2H), 3.31 (br s, 3H), 2.42
(t, 2H, J = 6.9 Hz),
1.75 (quint, 2H, J= 6.6 Hz).
EXAMPLE 17: Preparation of 1-(2-chloroethyl)-3-[3-(5-hydroxypentyl)phenyl)urea
[17)
O O
HO ~ / ~ ~CI
N- 'N
H H
The crude 3-(5-hydroxypent-1-ynyl) aniline (5.62 g, 32.0 mmol) (as prepared in
example 16)
was dissolved in ethanol (40 mL) and placed in a hydrogenation bottle with 10%
Pd/C (604 mg).
The bottle was filled with 40 psi of hydrogen and shaken for 4 h. The product
was filtered
through Celite, and the solvent was removed in vacuo. The aniline was purified
by vacuum
distillation to give 3-(5-hydroxypentyl)aniline as a clear oil (3.49 g, 84%
for two steps, from 3-
iodoaniline). 3-(5-hydroxypentyl)aniline was then reacted with 2-Chloroethyl
isocyanate as
described in examples 1-12 to obtain compound 17.
CA 02568609 2006-11-28
Purified by flash chromatography on silica gel (ethyl acetate/hexane 45/55.
Yield 36%.
1H NMR (CDC13, 300 MHz) ~ 7.0-7.15 (m, 3H), 6.77 (d, 1H, J= 7.2 Hz), 3.45-3.6
(m, 6H), 3.31
(br s, 3H), 2.50 (t, 2H, J= 7.6 Hz), 1.45-1.6 (m, 4H), 1.30 (m, 2H).
EXAMPLE 18: Preparation of Acetic acid 5-f4-[3-(2-
chloroethyl)ureido]phenyl}pentyl
ester [18]
O
'O \ O
SCI
N~N
H H
To a cold (0 °C) solution of 3-(5-hydroxypentyl)aniline (708 mg, 3.95
mmol) (as prepared in
example 17) in dry dichloromethane (20 mL) was added, under nitrogen, (Boc)20
(1.00 g, 4.60
nunol). The solution was allowed to warm to room temperature and stirred for 2
days (the
reaction was monitored by TLC). The solution was diluted with ethyl acetate.
The organic phase
was washed with a 5% solution of citric acid (twice), brine, dried over NaaS04
and concentrated
under vacuum. [3-(5-Hydroxypentyl)phenyl]carbamic acid tent-butyl ester was
used without any
further purification.
To a cold (0 °C) solution of [3-(5-Hydroxypentyl)phenyl]carbamic acid
test-butyl ester (112 mg,
0.40 mmol) in dry dichloromethane (3 mL) was added triethylamine (0.06 mL,
0.43 mmol) and
acetyl chloride (32 ~.L, 0.45 mmol) under nitrogen. The solution was allowed
to warm to room
temperature and stirred for 5 hours. The solution was diluted with ethyl
acetate. The organic
phase was washed successively with a 5% solution of citric acid (3 times), a
saturated solution of
NaHC03 (twice), brine, dried over Na2S04 and evaporated to give acetic acid 5-
(3-tert-
butoxycarbonylaminophenyl)pentyl ester which was purified by flash
chromatography on silica
gel (ethyl acetate/hexane 15/85) (88 mg, 68 %).
Acetic acid 5-(3-tent-butoxycarbonylamino-phenyl)-pentyl ester (88 mg, 0.27
mmol) was
dissolved in a mixture of trifluoroacetic acid (4.5 mL) and water (0.5 mL) and
the solution was
66
CA 02568609 2006-11-28
stirred for 10 min at room temperature. The solvents were evaporated and the
residue was taken
up in ethyl acetate. The organic phase was washed with a saturated solution of
NaHCO3 (twice),
brine, dried over Na2S04 and concentrated in vacuo to give acetic acid 5-(3-
aminophenyl)pentyl
ester (36 mg, 60%).
Acetic acid 5-(3-aminophenyl)pentyl ester was then reacted with 2-Chloroethyl
isocyanate as
described in examples 1-12 to obtain compound 18.
Purified by flash chromatography on silica gel (ethyl acetate/hexane 3/7).
Yield 86%
1H NMR (CDC13, 300 MHz) 8 7.19 (d, 2H, J= 8.3 Hz), 7.10 (d, 2H, J= 8.3 Hz),
4.04 (t, 2H, J=
6.7 Hz), 3.60 (m, 2H), 3.55 (m, 2H), 2.56 (t, 2H, J= 7.6 Hz), 2.04 (s, 3H),
1.63 (m, 4H), 1.38 (m,
2H).
EXAMPLE 19: Preparation of 6-{3-[3-(2-chloroethyl)ureido]phenoxy~hexanoic acid
ethyl
ester [19]
O
~ SCI
O O / N. 'N
H H
O
To a solution of 3-nitrophenol (1.72 g, 12.3 mmol) and KzC03 (3.40 g, 24.6
mmol) in acetone
(30 mL) was added dropwise 6-bromohexanoic acid ethyl ester (2.3 mL, 15 mmol).
The mixture
was refluxed under nitrogen for 48 hours. The solution was diluted with ethyl
acetate. The
organic phase was successively washed with water, a saturated solution of
NaHC03 (3 times),
brine, dried over MgS04 and evaporated under reduced pressure. The residue was
purified by
flash chromatography on silica gel (hexane/ethyl acetate 9/1) to yield 6-(3-
nitrophenoxy)hexanoic acid ethyl ester (3.34 g, 48%).
To a solution of 6-(3-nitrophenoxy)hexanoic acid ethyl ester (442 mg, 1.57
mmol) in ethanol (20
mL) was added SnC~2 ~ 2 H20 (1.78 g, 7.90 mmol) and the mixture was refluxed
for 4 hours.
The solution was cooled and poured into chilled water. The aqueous phase was
alkalized by
67
CA 02568609 2006-11-28
adding a solution of 1M NaOH and extracted with ethyl acetate (3 times). The
organic portions
were reunited, washed with brine (twice), dried over Na2S04 and evaporated
under reduced
pressure to yield 6-(3-aminophenoxy)hexanoic acid ethyl ester which was used
without any
further purification-(371 mg, 94%).
6-(3-aminophenoxy)hexanoic acid ethyl ester was then reacted with 2-
Chloroethyl isocyanate as
described in examples 1-12 to obtain compound 19. Purified by flash
chromatography
(hexane/ethyl acetate 3/2). Yield 85%.
1H NMR (CDC13, 300 MHz) 8 7.11 (t, 1H, J = 8.1 Hz), 6.96 (t, 1H, J = 2.1 Hz),
6.77 (dd, 1H, J =
8.1, 1.1 Hz), 6.54 (dd, 1H, J = 8.1, 2.1 Hz), 4.11 (q, 2H, J = 7.1 Hz), 3.86
(t, 2H, J = 6.4 Hz),
3.54 (m, 4H), 2.30 (t, 2H, J = 7.5 Hz), 1.6-1.8 (m, 4H), 1.4-1.5 (m, 2H), 1.24
(t, 3H, J = 7.1 Hz).
EXAMPLE 20: Preparation of 1-(2-chloroethyl)-2-(2-heptylphenyl)urea [20]
O
~ CI
N~N~
H H
2-Iodoaniline (1.31 g, 5.97 mmol), KZC03 (2.07 g, 15.0 mmol), CuI (47 mg, 0.25
mmol), PPh3
(134 mg, 0.51 mmol), and 10% Pd/C (125 mg, 0.12 mmol Pd) were mixed in 1,2-
dimethoxyethane (15 mL) and water (15 mL) at 25 °C under a nitrogen
atmosphere. This was
stirred for 30 minutes and hept-1-yne (2.0 mL, 15 mmol) was added. The mixture
was heated at
80 °C for 16 hours, cooled to room temperature and filtered through
Celite. The organic solvents
were removed in vacuo, and the aqueous residue acidified with 1M HCI. This
solution was
extracted with ethyl acetate, and the aqueous phase basified with KOH. The
water layer was
then extracted with ethyl acetate (3X). The organic portions were reunited,
washed with brine,
dried over NazS04 and concentrated in vacuo. The residue was purified by flash
chromatography
on silica gel (THF/hexane 5/95) to yield 2-(hept-1-ynyl)aniline (909 mg, 81%).
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2-(hept-1-ynyl)aniline (628 mg, 3.35 mmol) was dissolved in ethanol (20 mL)
and placed in a
hydrogenation bottle with 10% Pd/C (88 mg). The bottle was filled with 40 psi
of hydrogen and
shaken for 4 h. The product was filtered through Celite, and the solvent was
removed in vacuo
to obtain 2-heptanylaniline. The aniline was used without any further
purification (602 mg,
94%).
2-heptanylaniline was then reacted with 2-Chloroethyl isocyanate as described
in examples 1-12
to obtain compound 20. Purified by flash chromatography (ethyl acetate/hexane
3/7). Yield
83%.
1H NMR (CDC13, 300 MHz) 8 7.37 (m, 1H), 7.1-7.25 (m, 3H), 6.63 (br s, 1H),
5.44 (t, 1H, J =
5.5 Hz), 3.58 (m, 2H), 3.49 (m, 2H), 2.57 (t, 2H, J = 7.7 Hz), 1.52 (m, 2H),
1.25-1.55 (m, 8H),
0.86 (t, 3H, J = 6.7 Hz).
EXAMPLE 21: Preparation of 1-(2-Chloroacetyl)-3-(4-iodophenyl)urea [21]
I ~ O O
/ ~ ~CI
N- 'N
H H
2-Chloroacetyl isocyanate (1.15 mL, 13.5 mmol) was slowly added dropwise to a
magnetically
stirred and cooled solution (ice bath) of freshly recrystallized 4-iodoaniline
(2.60 g, 11.8 mmol)
in dichloromethane (70 mL). The ice bath was then removed and the reaction
mixture was stirred
at room temperature for 20 h. After completion of the reaction, the solvent
was removed under
reduced pressure to give a yellow solid, which was purified by
recrystallization (THF/hexane) to
give Z1 (3.23 g, 81%).
1H NMR (DMSO, 300 MHz) ~ 10.93 (br s, 1H), 10.15 (br s, 1H), 7.66 (d, 2H, J=
8.6 Hz), 7.37
(d, 2H, J= 8.6 Hz), 4.38 (s, 2H).
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EXAMPLE 22: Preparation of 1-(2-chloroacetyl)-3-(3-(5-
hydroxypentyl)phenyl]urea [22]
O O
HO ~ / N~N~CI
H H
Prepared as described for compound 21 from 3-(5-hydroxypentyl)aniline (see
previous example
for its preparation).
Purified by flash chromatography (chloroform/ethyl acetate 1/1). Yield 51%.
1H NMR (CDC13, 300 MHz) 8 7.34 (s, 1H), 7.15 (m, 2H), 6.87 (m, 1H), 4.43 (s,
2H), 4.11 (s,
2H), 4.08 (m, 2H), 2.55 (t, 2H, J= 7.2 Hz), 1.5-1.65 (m, 4H), 1.36 (m, 2H).
EXAMPLE 23: Preparation of (R)-1-(2-chloro-1-methylethyl)-3-(4-iodophenyl)urea
[(R)-
23]
I / ~ O
~ SCI
N- -N
H H
To a stirred solution of phenyl chloroformate (4.04 g, 25.8 mmol) in dry THF
(60 mL) at 0 °C
and under nitrogen was added 4-iodoaniline (5.59 g, 25.5 mmol) over 5 minutes.
After the
addition was complete, triethylamine (4.0 mL, 28.7 mmol) was added. The ice
bath was
removed and the mixture was stirred at room temperature for 4 h. The mixture
was diluted with
ethyl acetate and the organic solution was washed successively with 1 M HCl
(2X), 1 M NaOH
(3X), brine (1X), dried over MgS04 and evaporated down to afford a white solid
(6.99 g, 81%)
which was used directly.
In a round bottom flask equipped with a drying tube filled with CaCl2, (R)-
alaninol (698 mg,
9.17 mmol) was dissolved in acetonitrile (35 mL). To the solution was added
the carbamate
CA 02568609 2006-11-28
(2.575 g, 7.59 mmol) and the mixture was stirred at room temperature for 3
days. The solution
was then diluted with ethyl acetate (heating might be necessary to dissolve
completely the urea).
The organic portion was washed successively with 1M HCl (2X), 1M NaOH (3X),
brine (1X),
dried over MgS04 and the solvent was evaporated under reduce pressure. The
pure product was
obtained after crystallization from CHC13/THF/hexanes (1.542 g, 63%).
To a suspension of the alcohol (1.79 g, 5.61 mmol) in dry THF (30 mL) and
under nitrogen was
added SOCl2 (0.6 mL, 8.2 mmol). The mixture was then heated for 30 min. The
cooled solution
was poured into brine and the product was extracted three times with ethyl
acetate. The organic
portions were reunited, washed successively with 1M HCl (2X), brine, dried
over MgS04 and
the solvent s were removed under reduced pressure. The residue was purified
twice by
crystallization from THF/hexane (1.42 g, 75%).
1H NMR (CDCl3 + MeOD, 300 MHz) 8 7.44 (d, 2H, J = 8.8 Hz), 7.07 (d, 2H, J
=.8.8 Hz), 4.09
(m, 1H), 3.59 (dd, 1H, J = 11.0, 4.6 Hz), 3.50 (dd, 1H, J =11.0; 3.6 Hz), 1.17
(d, 3H, J = 6.8 Hz).
EXAMPLE 24: Preparation of (.S~-1-(2-chloro-1-methylethyl)-3-(4-
iodophenyl)urea [(f~-23]
I
O
~CI
N N
H H
Prepared as described for (R)-23 using (~-alaninol instead of (R)-alaninol.
1H NMR (CDC13 + MeOD, 300 MHz) 8 7.44 (d, 2H, J = 8.8 Hz), 7.07 (d, 2H, J =
8.8 Hz), 4.09
(m, 1H), 3.59 (dd, 1H, J = 11.0, 4.6 Hz), 3.50 (dd, 1H, J = 11.0, 3.6 Hz),
1.17 (d, 3H, J = 6.8 Hz).
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EXAMPLE 25: Preparation of (R)-1-(4-tent-Butylphenyl)-3-(2-chloro-1-
methylethyl)urea
[(R)-24]
O
\ ~ ~ j~Cl
N N
H H
Prepared as described for (R)-23 starting with 4-tent-butylaniline.
EXAMPLE 26: Preparation of (S~-1-(4-tent-Butylphenyl)-3-(2-chloro-1-
methylethyl)urea
[(S~-24]
O
\ ~ ~ j~Cl
N N
H H
Prepared as described for (R)-23 starting with 4-tart-butylaniline and using
(~-alaninol instead
of (R)-alaninol.
EXAMPLE 27: Preparation of 1-(4-tent-Butylphenyl)-3-(2-chloro-1,1-
dimethylethyl)urea
[25]
O
\ ~ N- 'N CI
H H
Prepared as described for (R)-23 starting with 4-test-butylaniline and using 2-
amino-2-methyl-1-
propanol instead of (R)-alaninol.
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EXAMPLE 28: Preparation of 1-(2-Bromoethyl)-3-(3-iodophenyl)urea [26]
O
I / N~N~Br
H H
2-Bromoethyl isocyanate (0.21 mL, 2.32 mmol) was slowly added dropwise to a
magnetically
stirred and cooled solution (ice bath) of freshly recrystallized 3-iodoaniline
(465 mg, 2.12 mmol)
in dichloromethane (10 mL). The ice bath was then removed and the reaction
mixture was stirred
at room temperature for 20 h. After completion of the reaction, the solvent
was removed under
reduced pressure to give a solid, which was purified by recrystallization
(THF/hexane) to give 26
(650 mg, 83%)
1H NMR (CDC13, 300 MHz) 8 7.72 (d, 1H, J= 2.0 Hz), 7.25 (m, 2H), 6.89 (t, 1H,
J= 8.0 Hz),
3.53 (M, 2H), 3.41 (M, 2H).
EXAMPLE 29: Preparation of 3-[3-(2-bromoethyl)ureido]benzoic acid ethyl ester
[27]
O
O / N~N~Br
i H H
O
Prepared as described for compound 26 using 3-aminobenzoic acid ethyl ester.
Purified by flash chromatography on silica gel (ethyl acetate/hexane 2/3).
Yield 87%.
1H NMR (CDCl3, 300 MHz) 8 7.87 (s, 1H), 7.72 (d, 2H, J= 8.0 Hz), 7.36 (t, 1H,
J= 8.0 Hz),
4.36 (q, 2H, J= 7.1 Hz), 3.67 (m, 2H), 3.51 (m, 2H), 1.38 (t, 3H, J= 7.1 Hz).
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EXAMPLE 30: Preparation of 4-tent-Butylphenyl(4,5-dihydrooxazol-2-yl)amine
[27]
~N N
H
4-(test-Butylphenyl)-3-(2-chloroethyl)urea (102 mg, 0.40 mmol) and KF
supported on silica gel
40% (220 mg) were suspended in acetonitrile (5 mL). The mixture was stirred at
room
temperature for 3 days. The solvent was removed under reduced pressure and the
solid was
purified by flash chromatography on silica gel (methanol/dichloromethane 5/95)
to yield the
oxazoline (41 mg, 47%).
1H NMR (CDC13, 300 MHz) 8 7.28 (d, 2H, J = 8.7 Hz), 7.18 (d, 2H, J = 8.7 Hz),
4.37 (t, 2H, J =
8.4 Hz), 3.80 (t, 2H, J = 8.4 Hz), 1.27 (s, 9H).
EXAMPLE 31: Preparation of 1-(Z-Chloro-ethyl)-3-(3-iodo-phenyl)-urea [28]
0
~ ~cl
I \ N- 'N
H H
2-Chloroethyl isocyanate (1,2 Eq, 1.640 mmol, 0.173 g) was slowly added
dropwise to a
magnetically stirred and cooled solution (ice bath) of freshly distilled 3-
iodobenzenamine (1.0
Eq, 1.370 mmol, 0.300 g) in dichloromethane (15 mL of solvent/g of aniline).
The ice bath was
then removed and the reaction mixture was kept at room temperature for 20 h.
After completion
of the reaction, the solvent was removed by vacuum distillation to give a
white solid, which was
purified by flash chromatography on silica gel: dichloromethane/ MeOH (98 /
2). Yield = 100%
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NMR 1H (Acetone) 8: 8.18 (s, NHCONH, 1H), 8.10 (s, Ar, 1H), 7.37 (d, Ar, 1H J
= 8,01), 7.30
(d, Ar, 1H, J = 7,86), 7,02 (t, Ar, 1H, J = 8,01), 6.17 (s, NHCONH, 1H), 3.68
(m, 2H), 3.53 (q,
2H, J = 5,85) NMR 13C (Acetone) : 158.0, 143.0, 131.1, 127.4, 125.0, 118.0,
102.8, 44.7, 42.4.
EXAMPLE 32: Preparation of 1-(2-Chloro-ethyl)-3-[3-(7-hydroxy-heptyl)-phenyl]-
urea (29)
O
HO I / N~N~CI
H H
3-iodonitrobenzene (1.0 Eq, 3.655 mmol, 0.910 g), Pd/C 10% (0.02 Eq, 0.073
mmol, 0.078 g),
PPh3 (0.08 Eq, 0.292 mmol, 0.077 g), CuI (0.04 Eq, 0.146 rnmol, 0.028 g),
KZCO3 (2.52 Eq,
9.212 mmol, 1.273 g), 1,2-DME (5 mL), H20 (5 mL) were mixed at 25 °C
under a nitrogen
atmosphere. The reaction mixture was stirred for 30 minutes and 6-Heptyn-1-of
(1.0 Eq, 3.655
mmol, 0.410 g) was added. The mixture was heated at 80 °C for 16 hours,
cooled to room
temperature. The organic solvents were removed ifa vacuo, and the product was
purified by flash
crhomatographie on silica gel: Hexane / AcOEt (75/25).
7-(3-nitrophenyl)hept-6-yn-1-of (1.0 Eq, 0.276 mmol, 0.064 g) was dissolved in
ethanol (10 mL)
and placed in a hydrogenation bottle with 10% Pd/C (0.03 Eq, 0.009 mmol, 0.010
g). The bottle
was filled with 38 psi of hydrogen and shaken for 4 h. The product was
filtered through Celite,
and the solvent was removed in vacuo. The resulting aniline was purified by
flash
crhomatography on silica gel: Hexane / AcOEt (70 l 30).
In a round bottom flask equipped with a dry tube filled with CaClz, 7-(3-
aminophenyl)heptan-1-
ol (1.0 Eq, 0.060 mmol, 0.012 g) was dissolved in dichloromethane (10 mL). 2-
chloroethyl
isocyanate (1.1 Eq, 0.067 mmol, 0.007 g) was added and the mixture was stirred
at room
temperature overnight. The solvent was removed and the residue was purified by
flash
chromatography on silica gel: Dichloromethane / MeOH (98 l 2).
CA 02568609 2006-11-28
NMR 1H (Acetone) 8: 8.01 (s, NHCONH, 1H), 7.29 (m, Ar, 3H), 6.78 (m, Ar, 1H),
6.09 (s,
NHCONH, 1H), 3.67 (m, 2H), 3.52 (m, 4H), 2.87 (m, 4H), 2.54 (m, 2H), 1.35 (m,
6H).
EXAMPLE 33: Preparation of 1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pentyl)-phenyl]-
urea
(30)
0
HO ~ / ~ SCI
N N
H H
3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g), 4-Pentyn-1-of (2.67 Eq,
21.400 rmnol, 1.800
g), PdIC 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08 Eq, 0.630 mmol, 0.166
g), CuI (0.04
Eq, 0.320 mmol, 0.061 g), KZC03 (2.52 Eq, 20.200 mmol, 2.790 g), 1.2-DME (10
mL), H20 (10
mL) were mixed as described in Example 32. The procedure was similar. The
purification
technique was flash chromatography on silica gel : Hexane / AcOEt (70 / 30).
5-(3-nitrophenyl)pent-4-yn-1-of (1.0 Eq, 0.970 mmol, 0.200 g) in Pd / C 10%
(0.05 Eq, 0.047
mmol, 0.050 g), H2 (38 PSI), ETOH (10 mL) was hydrogenated as described in
Example 32 to
yield 5-(3-aminophenyl)pentan-1-ol. The product was purified by flash
chromatography on silica
gel : Hexane / AcOEt (70 / 30).
2-Chloroethyl isocyanate (1.2 Eq, 0.600 mmol, 0.063 g) and 5-(3-
aminophenyl)pentan-1-of (1.0
Eq, 0.500 mmol, 0.090 g) were mixed in dichloromethane (10 mL) as described in
Example 31.
The product was purified by flash chromatography on silica gel:
dichloromethane/ MeOH (98 /
2). Yield = 20%
1H NMR (CDC13 and MD30D) 8: 7.03 (m, Ar, 3H), 6.70 (t, Ar, 1H, J = 6.54) 3.89
(s, 3H), 4.03
(t, 2H, J = 6.54), 3.45 (m, 6H), 2.45 (t, 2H, J = 7.56), 1.45 (m, 4H), 1.23
(m, 2H). 13C NMR
(CDC13 and MD30D) 8: 156.3, 143.4, 138.9, 128.6, 122.8, 119.3, 116.6, 62.1,
44.2, 41.6, 35.7,
32.2, 30.8, 25.2.
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EXAMPLE 34: Preparation of 1-(2-Chloro-ethyl)-3-[3-(6-hydroxy-hexyl)-phenyl]-
urea
(31)
0
/ N~N~CI
H H
OH
3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g) was mixed with 5-Hexyn-1-of
(2.58 Eq,
20.670 rmnol, 2.030 g), PdIC 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08
Eq, 0.630 mmol,
0.166 g), CuI (0.04 Eq, 0.320 mmol, 0.061 g), K2C03 (2.52 Eq, 20.200 mmol,
2.790 g), 1,2-
DME (10 mL), HZO (10 mL) as described in Example 32.
6-(3-nitrophenyl)hex-5-yn-1-of (1.0 Eq, 0.958 mmol, 0.210 g) was hydrogenated
with Pd / C
10% (0.05 Eq, 0.047 mmol, 0.050 g), H2 (38 PSI), ETOH (10 mL) as described in
Example 32.
6-(3-aminophenyl)hexan-1-of (1.0 Eq, 0.671 mmol, 0.147 g) was mixed with 2-
chloroethyl
isocyanate (1.l Eq, 0.739 mmol, 0.078 g) in dichloromethane (10 mL) as
described in Example
31. The product was purified by flash chromatography on silica gel:
Dichloromethane/ MeOH
(97 / 3). Yield = 42%
1H NMR (Acetone) 8: 8.17 (s, NHCONH, 1H), 7.20 (m, Ar, 3H), 6.79 (d, Ar, 1H J
= 6.42), 6.25
(s, NHCONH, 1H), 3.60 (m, 6H), 2.51 (t, 2H, J = 6,87), 1.48 (m, 8H). 13C NMR
(Acetone) 8:
156.2, 144.1, 141.0, 129.2, 122.7, 119.2, 116.6, 62.4, 44.8, 42.5, 36.5, 33.6,
32.1, 29.7, 26.4.
EXAMPLE 35: Preparation of 1-(2-Chloro-ethyl)-3-[3-(4-hydroxy-butyl)-phenyl]-
urea (32)
~ o
HO / N~N/~CI
H H
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3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g) was mixed with 3-Butyn-1-of
(2.58 Eq,
20.670 mmol, 1.449 g), PdIC 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08 Eq,
0.630 mmol,
0.166 g), CuI (0.04 Eq, 0.320 mmol, 0.061 g), KZC03 (2.52 Eq, 20.200 mmol,
2.790 g), 1,2-
DME (10 mL), HZO (10 mL) as described in Example 32.
4-(3-nitrophenyl)but-3-yn-1-of (1.0 Eq, 1.046 mmol, 0.200 g) was hydrogenated
with PdIC 10%
(0.05 Eq, 0.047 mmol, 0.050 g), H2 (38 PSI), ETOH (10 mL) as described in
Example 32.
4-(3-aminophenyl)butan-1-of (1.0 Eq, 0.726 mmol, 0.120 g) was mixed with 2-
chloroethyl
isocyanate (1.2 Eq, 0.870 mmol, 0.091 g) in dichloromethane (10 mL) as
described in Example
31. The product was purified by flash chromatography on silica gel: Hexane /
AcOEt (65 / 35).
Yield = 18%
1H NMR (Acetone) ~: 8.17 (s, NHCONH, 1H), 7.20 (m, Ar, 3H), 6.79 (d, Ar, 1H J
= 6.42), 6.25
(s, NHCONH, 1H), 3.60 (m, 6H), 2.51 (t, 2H, J = 6,87), 1.48 (m, 8H). 13C NMR
(Acetone) b:
156.2, 144.1, 141.0, 129.2, 122.7, 119.2, 116.6, 62.4, 44.8, 42.5, 36.5, 33.6,
32.1, 29.7, 26.4.
EXAMPLE 36: Preparation of 1-(2-Chloro-ethyl)-3-[3-(3-hydroxy-propyl)-phenyl]-
urea
(33)
\ O
HO / N~N~CI
H H
To a mixture of 3-iodonitrobenzene (7 g, 28.1 mmoles), K2CO3 (11.63 g, 84.3
mmoles) in 80 mL
1,2-DME/water (1:1) were added successively CuI (229.50 mg, 1.21 mmoles), PPh3
(591.20 mg,
2.25 mmoles), Pd/C 10% (598.0 mg, 0.562 mmoles). The mixture was stirred at
room
temperature for 1 hour. 4-butyn-1-of (5.90 g, 84.30 mmoles) was added, then
the mixture was
heated to reflux overnight. After cooling, the mixture was filtered on Celite
and the organic layer
was evaporated under reduced pressure. Tha aqueous layer was acidified with
concentrated
Chlorhydric acid and extracted with AcOEt. The organic layer were washed with
brine, dried,
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CA 02568609 2006-11-28
filtered and evaporated. Purified by flash chrmatography on silica gel
AcOEt/Hexanes (35 :65).
Yield : 81
1H NMR (CDC13, 300 MHz) b: 8.29 (s, Ar, 1H), 8.17 (m, Ar, 1H), 7.74 (d, Ar,
1H, J = 8Hz),
7.51 (t, Ar, 1H, J = 8Hz), 4.53 (d, 2H, J = 6Hz).
A mixture of 3-(3-nitrophenyl)-prop-2-yn-1-of (100 mg, 0.564 mmoles), PdIC 10%
(10 mg,
0.094 mmoles) was hydrogenated under 38 psi overnight. The mixture was
filtered on Celite and
the filtrate was evaporated to dryness. Purified by flash chrmatography on
silica gel
AcOEt/Hexanes (25:75). Yield : 99%
1H NMR (CDC13, 300 MHz) 8: 7.08 (t, Ar, 1H, J = B.OHz), 6.61 (d, Ar, 1H, J =
7.5Hz), 6.53 (m,
Ar, 2H), 3.67 (t, 2H, J = 6.5Hz), 2.84 (s, 3H), 2.62 (t, 2H, J = 8.OHz), 1.87
(m, 2H).
3-(3-aminophenyl)propan-1-of was then reacted with 2-chloroethylisocyanate as
described in
examples 1-12 to obtain desired product. Purified by flash chromatography on
silica gel
EtOH/CH2C12 (2:98). Yield: 28%
1H NMR (Acetone) b: 8.00 (s, NHCONH, 1H), 7.36 (s, Ar; 1H), 7.29 (d, Ar, 1H J
= 8.01), 7.13
(t, Ar, 1H J = 7.83), 6.80 (d, Ar, 1H J = 7.32), 6.09 (s, NHCONH, 1H), 3.68
(m, 2H), 3.55 (m,
4H), 2.63 (t, 2H, J = 7.62), 1.79 (m, 2H). 13C NMR (Acetone) 8: 155.9, 143.8,
141.2, 129.2,
122.6, 119.1, 116.5, 61.7, 42.6, 42.4, 35.3, 32.8.
EXAMPLE 37: Preparation of 1-(3-Sromo-phenyl)-3-(2-chloro-ethyl)-urea (34)
O
SCI
Br N N
H H
3-bromobenzenamine (1.0 Eq, 1.740 mmol, 0.300 g) was mixed with 2-chlroethyl
isocyanate
(1,2 Eq, 2.090 mmol, 0.220 g) in dichloromethane (10 mL) as described in
Example 31. The
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solvent was removed and the product was purified by flash chromatography on
silica gel:
dichloromethane / MeOH (98 / 2). Yield = 100%
1H NMR (Acetone) 8: 8.26 (s, NHCONH, 1H), 7.93 (m, Ar, 1H), 7.31 (m, Ar, 1H),
7.15 (m, Ar,
2H), 6.20 (s, NHCONH, 1H), 3.67 (m, 2H), 3.52 (q, 2H, J = 2,11). 13C NMR
(Acetone) 8: 155.6,
142.9, 131.0, 125.0, 122.7, 121.3, 117.4, 44.7, 42.5.
EXAMPLE 38: Preparation of 1-(2-Chloro-ethyl)-3-(3-chloro-phenyl)-urea (35)
O
SCI
CI N N
H H
3-chlorobenzenamine (1.0 Eq, 2.350 mmol, 0.300 g) was mixed with 2-chlroethyl
isocyanate
(1,2 Eq, 2.820 mmol, 0.298 g) in dichloromethane (15 mL) as described in
Example 31. The
solvent was removed and the product was purified by flash chromatography on
silica gel:
dichloromethane / MeOH (98 / 2). Yield = 99%.
1H NMR (Acetone) b: 8.28 (s, NHCONH, 1H), 7.76 (m, Ar, 1H), 7.24 (m, Ar, 2H),
6.95 (m, Ar,
1H), 6.21 (s, NHCONH, 1H), 3.68 (m, 2H), 3.54 (q, 2H, J = 2,13). 13C NMR
(Acetone) 8: 155.6,
142.7, 134.6, 130.7, 122.0, 118.6, 117.0, 44.6, 42.4.
EXAMPLE 39: Preparation of (36)
O
SCI
HO N N
H H
3-aminophenol (1.0 Eq, 2.740 mmol, 0.300 g) was mixed with 2-chloroethyl
isocyanate (1.2 Eq,
3.300 mmol, 0.348 g) in THF (4 mL) and dichloromethane (10 mL) as described in
Example 31.
The solvent was removed and the product was purified by flash chromatography
on silica gel:
dichloromethane / MeOH (99 / 1). Yield = 96%.
CA 02568609 2006-11-28
1H NMR (Acetone)8: 8.57 (s, Ph, 1H), 8.23 (s, NHCONH, 1H), 7.25 (s, NHCONH,
1H), 7.06
(m, Ar, 1 H), 6. 8 0 (d, Ar, 1 H, J = 7. 71 ), 6. 5 0 (m, Ar, 1 H), 6. 3 5 (m,
Ar, 1 H), 3 .64 (m, 2H), 3 . 5 5
(q, 2H, J = 5.73). 13C NMR (Acetone) 8: 158.7, 156.8, 141.7, 130.4, 110.9,
110.3, 107.0, 44.8,
42.6.
EXAMPLE 40: Preparation of Acetic acid 3-[3-(2-chloro-ethyl)-ureido]-phenyl
ester (37)
0
~a
~O ~ N N
H H
1-(2-chloroethyl)-3-(3-hydroxyphenyl)urea (IMO-365) was obtained as described
in example 39.
1-(2-chloroethyl)-3-(3-hydroxyphenyl)urea (1.0 Eq, 0.326 mmol, 0.070 g) was
mixed with
triethylamine (3.0 Eq, 0.978 mmol, 0.099 g), acetic anhydride (3.0 Eq, 0.978
mmol, 0.100 g) and
4-pyrrolidinopyridine (0.02 Eq, 0.007 mmol, 0.001 g) at room temperature. The
solvent was
removed and the residue was purified by flash chromatography on silica gel,
Dichloromethane /
MeOH (98 / 2). Yield = 27%
1H NMR (Acetone) 8: 8.32 (s, NHCONH, 1H), 7.48 (s, Ar, 1H), 7.20 (m, Ar, 2H),
6.68 (m, Ar,
1H), 6.22 (s, NHCONH, 1H), 3.67 (m, 2H), 3.55 (q, 2H, J = 5.82). 13C NMR
(Acetone) ~: 169.5,
155.4, 152.2, 142.4, 129.8, 115.8, 115.5, 112.3, 44.7, 42.4, 20.8.
EXAMPLE 41: Preparation of 1-(2-Chloro-ethyl)-3-(3-hydroxymethyl-phenyl)-urea
(38)
O
HO I / ~ SCI
N N
H H
3-nitrobenzylalcohol (1.0 Eq, 1.959 mmol, 0.300 g) was reduced on SnC12.2H20
(6.0 Eq, 11.750
mmol, 2.650 g) and EtOH (20 mL). (3-aminophenyl)methanol (1.0 Eq, 1.620 mmol,
0.200 g)
was mixed with 2-chlroethyl isocyanate (1.2 Eq, 1.940 mmol, 0.206 g)in THF (20
mL) as
81
CA 02568609 2006-11-28
described in Example 31. The solvent was removed and the product was purified
by flash
chromatography on silica gel: dichloromethane / MeOH (98 / 2). Yield = 56%
1H NMR (Acetone) 8: 8.09 (s, NHCONH, 1H), 7.46 (s, Ar, 1H), 7.37 (m, Ar, 1H),
7.18 (t, Ar,
1H, J = 7.86), 6.95 (d, Ar, 1H, J = 7,35), 6.15 (s, NHCONH, 1H), 4.57 (s, 2H),
4.23 (s, OH, 1H),
3.67 (m, 2H), 3.53 (q, 2H, J = 6,00). 13C NMR (Acetone) 8: 156.0, 143.9,
141.0, 129.2, 120.7,
117.6, 117.3, 64.6, 44.8, 42.4.
EXAMPLE 42: Preparation of Acetic acid 5-{3-[3-(2-chloro-ethyl)-ureido]-
phenyl}-pentyl
ester (39)
O
O I ~ N~N~CI
IpI H H
3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g) was mixed with 4-pentyn-1-of
(2.67 Eq,
21.400 mmol, 1.800 g), Pd/C 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08 Eq,
0.630 mmol,
0.166 g), CuI (0.04 Eq, 0.320 mmol, 0.061 g), KzC03 (2.52 Eq, 20.200 mmol,
2.790 g), 1,2-
DME (10 mL), HZO (10 mL) as described in Example 32. The solvent was removed
and the
residue was purified by flash chromatography on silica gel: hexane / AcOEt (70
/ 30).
5-(3-nitrophenyl)pent-4-yn-1-of (1.0 Eq, 1.460 mmol, 0.300 g) was mixed with
triethylamine
(3.0 Eq, 4.380 mmol, 0.443 g), acetic anhydride (3.0 Eq, 4.380 mmol, 0.447 g)
and 4-
pyrrolidinopyridine (0.02 Eq, 0.029 mmol, 0.004 g) at room temperature. The
solvent was
removed and the residue was purified by flash chromatography on silica gel:
hexane / AcOEt (75
/ 25).
5-(3-nitrophenyl)pent-4-ynyl acetate (1.0 Eq, 0.978 mmol, 0.242 g) in Pd / C
10% (0.05 Eq,
0.047 mmol, 0.050 g), H2 (38 PSI) and EtOH (10 mL) was hydrogenated and
purified as
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CA 02568609 2006-11-28
described in Example 32. The solvent was removed and the product was purified
by flash
chromatography on silica gel: hexane / AcOEt (75 / 25).
5-(3-aminophenyl)pentyl acetate (1.0 Eq, 0.704 mmol, 0.156 g) was mixed with 2-
chlroethyl
isocyanate (1.2 Eq, 0.808 mmol, 0.085 g) in dichloromethane (15 mL) as
described in Example
31. The solvent was removed and the product was purified by flash
chromatography on silica
gel: dichloromethane / AcOEt (70 / 30). Yield = 54%
1H NMR (CDC13) ~: 7.81 (s, NHCONH, 1H), 7.09 (m, Ar, 3H), 6.80 (d, Ar, 1H, J =
7.32), 6.07
(s, NHCONH, 1H), 3.48 (m, 4H), 2.48 (t, 2H, J= 7.68), 2.02 (s, Ac, 3H), 1.56
(m, 4H), 1.31 (m,
2H). 13C NMR (CDC13) 8: 171.5, 156.4, 143.5, 138.8, 128.9, 123.3, 120.1,
117.5, 64.5, 44.3,
41.9, 35.7, 30.9, 28.4, 25.5, 21Ø
EXAMPLE 43: Preparation of Acetic acid 4-{3-[3-(2-chloro-ethyl)-ureido]-
phenyl}-butyl
ester (40)
o ~ w
~ci
~O ~ N N
H H
3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g) was mixed with 3-Butyn-1-of
(2.58 Eq,
20.670 mmol, 1.449 g), Pd/C 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08 Eq,
0.630 mmol,
0.166 g), CuI (0.04 Eq, 0.320 mmol, 0.061 g), KzC03 (2.52 Eq, 20.200 mmol,
2.790 g), 1,2-
DME (10 mL), H2O (10 mL) as described in Example 32. The solvent was removed
and the
residue was purified by flash chromatography on silica gel: Hexane / AcOEt (65
/ 35).
4-(3-nitrophenyl)but-3-yn-1-of (1.0 Eq, 1.046 mmol, 0.200 g) was mixed with
triethylamine (3.0
Eq, 3.1380 mmol, 0.318 g), acetic anhydride (3.0 Eq, 3.138 mmol, 0.320 g), and
4-
pyrrolidinopyridine (0.02 Eq, 0.021 mmol, 0.003 g) at room temperature. The
solvent was
removed.
83
CA 02568609 2006-11-28
4-(3-nitrophenyl)but-3-ynyl acetate (1.0 Eq, 0.988 mmol, 0.231 g) in Pd / C
10% (0.05 Eq, 0.047
mmol, 0.050 g), H2 (38 PS)7, ETOH (10 mL) was hydrogenated and purified as
described in
Example 32. The solvent was removed and the product was purified by flash
chromatography on
silica gel: hexane / AcOEt (75 / 25).
4-(3-aminophenyl)butyl acetate (1.0 Eq, 0.471 mmol, 0.098 g) was mixed with 2-
chlroethyl
isocyanate (1.2 Eq, 0.539 mmol, 0.057 g) in dichloromethane (15 mL) as
described in Example
31. The solvent was removed and the product was purified by flash
chromatography on silica
gel: hexane / AcOEt (65 / 35). Yield = 48%
1H NMR (CDC13) ~: 7.69 (s, NHCONH, 1H), 7.11 (m, Ar, 3H), 6.83 (d, Ar, 1H, J =
7.32), 6.07
(s, NHCONH, 1H), 4.03 (t, 2H, J = 6.57), 3.53 (m, 4H), 2.52 (t, 2H, J= 7.89),
2.03 (s, Ac, 3H),
1.60 (m, 4H). 13C NMR (CDC13) 8: 171.5, 156.3, 143.2, 138.7, 129.0, 123.5,
120.3, 117.8, 64.4,
44.4, 41.9, 35.4, 28.1, 27.5, 21Ø
EXAMPLE 44: Preparation of Acetic acid 3-[3-(2-chloro-ethyl)-ureido]-benzyl
ester (41)
O
O I ~ ~ SCI
N N
O H H
3-nitrobenzylalcohol (1.0 Eq, 1.959 mmol, 0.300 g) was mixed with
triethylamine (3.0 Eq, 5.880
mmol, 0.595 g), acetic anhydride (3.0 Eq, 5.880 mmol, 0.600 g) and 4-
pyrrolidinopyridine (0.02
Eq, 0.039 mmol, 0.006 g) at room temperature. The solvent was removed and the
product was
purified by flash chromatography on silica gel: hexane l AcOEt (75 / 25).
3-nitrobenzyl acetate (1.0 Eq, 0.649 mmol, 0.127 g) was reduced on SnC12.2Hz0
(6.0 Eq, 3.890
mmol, 0.879 g) and EtOH (20 mL). The solvent was removed and the product was
purified by
flash chromatography on silica gel: dichloromethane / MeOH (95 / 5).
84
CA 02568609 2006-11-28
2-(3-aminophenyl)ethyl acetate (1.0 Eq, 0.352 mmol, 0.058 g) was mixed with 2-
chlroethyl
isocyanate (1,2 Eq, 0.422 mmol, 0.045 g) in dichloromethane (10 mL) as
described in Example
31. The solvent was removed and the product was purified by flash
chromatography on silica
gel: dichloromethane / MeOH (95 / 5). Yield = 31
1H NMR (CDC13) 8: 7.79 (s, NHCONH, 1H), 7.23 (m, Ar, 3H), 6.98 (m, Ar, 1H),
6.08 (s,
NHCONH, 1H), 4.98 (s, 2H), 3.53 (m, 4H), 2.05 (s, Ac, 3H). 13C NMR (CDC13)
171.2, 156.2,
139.0, 136.9, 129.3, 122.9, 119.9, 119.8, 66.1, 44.4, 41.9, 21Ø
EXAMPLE 45: Preparation of Acetic acid 3-{3-[3-(2-chloro-ethyl)-ureido]-phenyl-
propyl
ester (42)
OI~
\/O I / N~N~CI
(O H H
To a mixture of 3-iodonitrobenzene (1 g, 4.56 mmoles), K~C03 (1.57 g, 11.4
mmoles) in 30 mL
1.2-DME/water (1:1) were added successively CuI (34.78 mg, 0.18 mmoles), PPh3
(95.80 mg,
0.36 mmoles), Pd/C 10% (97.05 mg, 0.09 mmoles). The mixture was stirred at
room temperature
for 1 hour. Propargyl alcohol (807 mg, 14.40 mmoles) was added, then the
mixture was heated to
reflux overnight. After cooling, the mixture was filtered on Celite and the
organic layer was
evaporated under reduced pressure. Tha aqueous layer was acidified with
concentrated
Chlorhydric acid and extracted with AcOEt. The combined organic layers were
washed with
brine, dried, filtered and evaporated. Purified by flash chromatography on
silica gel
CHZC12/EtOH (95: 5). Yield : 81
1H NMR (CDC13, 300 MHz) b: 8.29 (s, Ar, 1H), 8.17 (m, Ar, 1H), 7.74 (d, Ar,
1H, J = 8Hz),
7.51 (t, Ar, 1H, J = 8Hz), 4.53 (d, 2H, J = 6Hz).
To an ice-cold 3-(3-nitrophenyl)-prop-2-yn-1-of (150 mg, 0.85 mmoles) in
diethylether (10 mL)
were added acetic anhydride (254.23 mg, 2.54 mmoles), triethylamine (256.54
mg, 2.54
CA 02568609 2006-11-28
mmoles), 4-pyrrolidinopyridine (2.52 mg, 0.017 mmoles) and the mixture was
stirred at room
temperature for 12 hours. The reaction was quenched by saturated solution of
Na2C03 and the
mixture was extracted with AcOEt. The extracts were washed with brine, dried
and evaporated.
Purified by flash chromatography on silica gel AcOEt/Hexanes (8 :2). Yield :
99%
IH NMR (CDC13, 300 MHz) b: 8.24 (s, Ar, 1H), 8.14 (d, Ar, 1H, J = B.SHz), 7.71
(d, Ar, 1H, J =
7.SHz), 7.48 (t, Ar, 1H, J= 8Hz), 4.88 (s, 2H), 2.12 (s, 3H).
A mixture of acetic acid 3-(3-nitrophenyl)-prop-2-ynyl ester (100 mg, 0.48
mmoles), Pd/C 10%
(10 mg, 0.094 mmoles) in 30 mL of dry ethanol was hydrogenated under 38 psi
overnight. The
mixture was filtered on Celite and the filtrate was evaporated to dryness.
Purified by flash
chromatography on silica gel AcOEt/Hexanes (25:75). Yield : 81%
1H NMR (CDC13, 300 MHz) 8: 7.08 (m, Ar, 1H), 6.59 (d, Ar, 1H, J = 7.SHz), 6.53
(m, Ar, 2H),
4.09 (t, 2H, J = 6.SHz), 3.66 (s, 2H), 2.60 (t, 2H, J = 8Hz), 2.06 (s, 3H),
1.93 (m, 2H).
Acetic acid 3-(3-aminophenyl)-prop-2-ynyl ester was then reacted with 2-
chloroethylisocyanate
as described in examples 1-12 to obtain desired product. Purified by flash
chromatography on
silica gel AcOEt/CH2C12 (2:8). Yield : 93%
1H NMR (CDCl3) b: 7.51 (s, NHCONH, 1H), 7.11 (m, Ar, 3H), 6.98 (m, Ar, 1H),
6.84 (d, Ar,
1H, J = 7.38) 5.94 (s, NHCONH, 1H), 4.03 (t, 2H, J = 6.54), 3.55 (m, 4H), 2.59
(t, 2H, J = 7.92),
2.04 (s, Ac, 3H), 1.88 (m, 2H) NMR. 13C (CDC13) 8: 171.5, 156.2, 142.4, 138.7,
129.2, 123.6,
120.5, 118.1, 63.9, 44.5, 41.9, 32.1, 30.0, 21Ø
EXAMPLE 46: Preparation of 1-(2-Chloro-ethyl)-3-[3-(2-hydroxy-ethyl)-phenyl]-
urea (43)
O
HO \ N~N~CI
H H
86
CA 02568609 2006-11-28
2-(3-nitrophenyl)ethan-1-of (1.0 Eq, 1.200 mmol, 0.200 g) was reduced on
SnC12.2H20 (6.0 Eq,
7.200 mmol, 1.625 g) and EtOH (20 mL). The solvent was removed and the product
was
purified by flash chromatography on silica gel: dichloromethane / MeOH (95 /
~5).
2-(3-aminophenyl)ethan-1-of (1.0 Eq, 0.437mmo1, 0.060 g) was mixed with 2-
chloroethyl
isocyanate (1.2 Eq, 0.524 mmol, 0.055 g), in dichloromethane (15 mL) as
described in Example
31. The solvent was removed and the product was purified by flash
chromatography on silica
gel: Ether / Hexane (90 / 10). Yield = 20%
1H NMR (Acetone-d6) 8: 8.08 (brs, NH, 1H), 7.33 (m, Ar, 2H), 7.13 (m, Ar, 1H),
6.82 (d, Ar,
1H, J = 7.44), 6.15 (brs, NH, 1H), 3.69 (m, 4H), 3.53 (m, 2H), 2.93 (s, OH,
1H), 2.74 (t, 2H, J =
7.02). 13C NMR (Acetone-d6) b: 177.3, 155.9, 140.9, 129.2, 123.1, 119.5,
116.7, 63.7, 44.8,
42.4, 40.3.
EXAMPLE 47: Preparation of Acetic acid 2-{3-[3-(2-chloro-ethyl)-ureido]-
phenyl}-ethyl
ester (44)
o I~ o
C / N~N~CI
H H
2-(3-nitrophenyl) ethan-1-of (1.0 Eq, 1.495 mmol, 0.250 g) was mixed with
triethylamine (3.0
Eq, 4.485 mmol, 0.454 g), acetic anhydride (3.0 Eq, 4.485 mmol, 0.457 g) and 4-
pyrrolidinopyridine (0.02 Eq, 0.030 rmnol, 0.004 g) ) at room temperature. The
solvent was
removed and the product was purified by flash chromatography on silica gel:
Hexane / AcOEt
(75 / 25).
3-nitrophenethyl acetate (1.0 Eq, 0.871 mmol, 0.182 g) in Pd / C 10% (0.05 Eq,
0.047 mmol,
0.050 g), HZ (38 PSl~, and ETOH (10 mL) was hydrogenated and purified as
described in
Example 32. The solvent was removed and the product was purified by flash
chromatography on
silica gel: hexane / AcOEt (60 / 40).
87
CA 02568609 2006-11-28
2-(3-aminophenyl)ethyl acetate (1.0 Eq, 0.726 mmol, 0.130 g) was mixed with 2-
chloroethyl
isocyanate (1,2 Eq, 0.871 mmol, 0.092 g), in dichloromethane (15 mL) as
described in Example
31. The solvent was removed and the product was purified by flash
chromatography on silica
gel: dichloromethane / MeOH (98 / 2). Yield = 87%
1H NMR (Acetone) 8: 7.90 (s, NHCONH, 1H), 7.11 (m, Ar, 3H), 6.83 (d, Ar, 1H, J
= 6.63), 6.21
(s, NHCONH, 1H), 4.18 (t, 2H, J = 6.96), 3.50 (m, 4H), 2.79 (t, 2H, J = 6.99),
1.99 (s, Ac, 3H)
NMR 13C (Acetone) 8: 171.4, 156.5, 138.9, 138.8, 129.1, 123.8, 120.5, 118.4,
64.8, 44.3, 41.9,
34.9, 21Ø
EXAMPLE 48: Preparation of Acetic acid 6-[3-[3-(2-chloro-ethyl)-ureido]-
phenyl}-hexyl
ester (45)
O ~ O
O I / N~N~CI
H H
3-iodonitrobenzene (1.0 Eq, 8.010 mmol, 1.995 g) was mixed with 5-Hexyn-1-of
(2.58 Eq,
20.670 rninol, 2.030 g), Pd/C 10% (0.02 Eq, 0.160 mmol, 0.170 g), PPh3 (0.08
Eq, 0.630 mmol,
0.166 g), CuI (0.04 Eq, 0.320 mmol, 0.061 g), I~2C03 (2.52 Eq, 20.200 mmol,
2.790 g), 1,2-
DME (10 mL), H20 (10 mL) under a nitrogen atmosphere. The reaction was carried
out as
described in Example 32. The organic portion was purified by flash
chromatography on silica gel
Hexane / AcOEt (75 / 25).
6-(3-nitrophenyl)hex-5-yn-1-of (1.0 Eq, 1.140 mmol, 0.255 g) was mixed with
triethylamine (3.0
Eq, 3.420 mmol, 0.346 g), acetic anhydride (3.0 Eq, 3.420 mmol, 0.349 g) and 4-
pyrrolidinopyridine (0.02 Eq, 0.023 mmol, 0.003 g) ) at room temperature. The
solvent was
removed and the product was purified by flash chromatography on silica gel:
Hexane / AcOEt
(75 / 25).
88
CA 02568609 2006-11-28
6-(3-aminophenyl)hex-5-ynyl acetate (1.0 Eq, 0.643 mmol, 0.168 g) in Pd / C
10% (0.07 Eq,
0.047 mmol, 0.050 g), H2 (38 PSI), and ETOH (10 mL) was hydrogenated and
purified as
described in Example 32. The solvent was removed and the product was purified
by flash
chromatography on silica gel: dichloromethane / MeOH (99 / 1).
6-(3-aminophenyl)hexyl acetate (1.0 Eq, 0.348 mmol, 0.082 g) was mixed with 2-
chloroethyl
isocyanate (1.2 Eq, 0.418 mmol, 0.0446 g), in dichloromethane (15 mL) as
described in
Example 31. The solvent was removed and the product was purified by flash
chromatography on
silica gel: dichloromethane / MeOH (98 / 2). Yield = 64%
1H NMR (Acetone) 8: 7.80 (s, NHCONH, 1H), 7.12 (m, Ar, 3H), 6.84 (m, Ar, 1H),
6.15 (s,
NHCONH, 1H), 4.05 (m, 2H), 3.51 (m, 4H), 2.42 (m, 2H), 2.08 (s, Ac, 3H) 1.56
(m, 4H), 1.32
(m, 4H). 13C NMR (Acetone) S: 171.7, 156.4, 143.8, 138.8, 128.9, 123.4, 120.2,
117.6, 64.7,
44.4, 41.9, 35.8, 31.2, 28.9, 28.8, 25.7, 21Ø
EXAMPLE 49: Preparation of 1-(2-Chloro-ethyl)-3-[3-(3-methoxy-propyl)-phenyl]-
urea
(46)
O
~O I / N~N~CI
H H
NaH (60%) (97.40 mg, 4.06 mmoles) was suspended in dry THF (8 mL) and 3-(3-
nitrophenyl)-
pent-4-yn-1-of (see synthesis of IMO-371) (200 mg, 0.98 mmoles) in dry THF (3
Ml) was added
dropwide at 0°C. The mixture was stirred for 15 minutes at 0°C.
The MeI (332.5 mg, 2.34
mmoles) was added dropwise and the mixture was stirred at room temperature for
3 hours.
Saturated solution of NaHC03 (10 mL) and MeOH (10 mL) were added. The mixture
was
extracted with AcOEt, dried, filtered and evaporated to dryness. Purified by
flash
chromatography on silica gel AcOEt. Yield : 58%
89
CA 02568609 2006-11-28
1H NMR (CDC13, 300 MHz) 8: 8.17 (s, 1H), 8.07 (d, 1H, J = 8Hz), 7.64 (d, 1H, J
= 7.SHz), 7.42
(m, 1H), 3.49 (t, 2H, J = 6Hz), 3.34 (s, 3H), 2.49 (t, 2H, J = 6Hz), 1.86 (t,
2H, J = 6Hz), 1.22 (m,
2H).
A mixture of 1-(5-methoxypent-1-ynyl)-3-nitrobenzene (100 mg, 0.425 mmoles),
Pd/C 10% (10
mg, 0.094 mmoles), dry ehtanol was hydrogenated under 38 psi overnight. The
mixture was
filtered on Celite and the filtrate was evaporated to dryness. Purified by
flash chrmatography on
silica gel AcOEt. Yield : 85%
1H NMR (DMSO-d6, 300MHz) 8: 7.17 (m, 1H), 6.55 (m, 3H), 3.37 (m, SH), 2.51 (m,
2H), 1.61
(m, 2H).
3-(5-methoxypentyl) phenylamine was then reacted with 2-chloroethylisocyanate
as described in
examples 1-12 to obtain desired product. Purified by flash chromatography on
silica gel AcOEt.
Yield : 87%
'H NMR (CDC13, 300 MHz) 8: 8,18 (brs, NH, 1H), 7,14 (m, 3H), 6,86 (d, Ar, 1H,
J = 7,2Hz),
3,57 (m, 4H), 3,33 (m, 4H), 2,55 (t, 2H, J = 7Hz), 1,59 (m, 4H).
EXAMPLE 50: Preparation of (47) 1-(3-pentyl-phenyl)-3-(2-chloro-ethyl)-urea
(47)
i I O
N~N~~~
H H
To a mixture of 3-iodonitrobenzene (1 g, 4.01 mmoles), I~ZC03 (1.38 g, 10.0
mmoles) in 20 mL
1,2-DME/water (1 :1) were added successively CuI (30.54 mg, 0.16 mmoles), PPh3
(84.14 mg,
0.32 mmoles), Pd/C 10% (85.35 mg, 0.080 mmoles). The mixture was stirred at
room
temperature for 1 hour. 1-pentyne (725.40 mg, 84.30 mmoles) was added, then
the mixture was
heated to reflux overnight. After cooling, the mixture was filtered on Celite
and the organic layer
was evaporated under reduced pressure. Tha aqueous layer was acidified with
concentrated
CA 02568609 2006-11-28
Chlorhydric acid and extracted with AcOEt. The organic layer were washed with
brine, dried,
filtered and evaporated. Purified by flash chrmatography on silica gel
HexaneslAcOEt (60 :40).
Yield : 58%
1H NMR (CDC13, 300 MHz) 8: 8.17 (s, 1H), 8.06 (d, 1H, J = 8Hz), 7.63 (d, 1H, J
= 7.5Hz), 7.41
(m, 1H), 2.37 (m, 2H), 1.02 (m, 2H).
A mixture of 1-vitro-3-pentynylbenzene (100 mg, 0.523 mmoles), Pd/C 10% (10
mg, 0.094
mmoles) in 30 mL of dry ethanol was hydrogenated under 38 psi overnight. The
mixture was
filtered on Celite and the filtrate was evaporated to dryness. Purified by
flash chrmatography on
silica gel CHZC12/EtOH (95 :5). Yield : 97%
1H NMR (DMSO-d6, 300MHz) ~: 7.09 (m, 1H), 6.63 (d, 1H, J = 7.5Hz), 6.53 (m,
2H), 3.55 (brs,
2H), 2.54 (m, 2H), 1.63 (m, 2H), 1.37 (m, 4H), 0.93 (m, 2H).
3-pentylphenylamine was then reacted with 2-chloroethylisocyanate as described
in examples 1-
12 to obtain desired product. Purified by flash chrmatography on silica gel
CH2Clz/EtOH
(95 :5). Yield : 91
1H NMR (CDC13, 300 MHz) b: 7,94 (brs, NH, 1H), 7,01 (m, Ar, 1H), 6,77 (brs,
Nh, 1H), 3,47
(m, CH2, 8H), 2,45 (t, CH2, 2H, J = 7), 1,21 (m, CHZ, 4H).
EXAMPLE 51: Preparation of 5-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-pentanoic
acid
(48)
i O
HO ~ I N~N~CI
O H H
91
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To a mixture of 3-iodonitrobenzene (5 g, 20.3 mmoles), K2C03 (7.03 g, 50.9
mmoles) in 80 mL
1,2-DME/water (1 :1) were added successively CuI (166.0 mg, 0.87 mmoles), PPh3
(427.11 mg,
1.82 mmoles), Pd/C 10% (432.06 mg, 0.406 mmoles). The mixture was stirred at
room
temperature for 1 hour. 5-pentanoic acid (5.90 g, 84.30 mmoles) was added,
then the mixture
was heated to reflux overnight. After cooling, the mixture was filtered on
Celite and the organic
layer was evaporated under reduced pressure. Tha aqueous layer was acidified
with concentrated
Chlorhydric acid and extracted with AcOEt. The organic layer were washed with
brine, dried,
filtered and evaporated.
Purified by flash chromatography on silica gel CHzCl2/EtOH (95 :5). Yield :
45%
1H NMR (DMSO-d6, 300 MHz) 8: 12.39 (brs, 1H), 8.19 (d, 1H, J = 8Hz), 8.13 (s,
1H), 7.82 (d,
1H, J = 7.SHz), 7.66 (t, 1H, J = 8Hz), 2.63 (m, 4H).
A mixture of 5-(3-nitrophenyl)-pent-4-ynoic acid (100 mg, 0.456 mmoles), Pd/C
10% (10 mg,
0.094 mmoles) was hydrogenated under 38 psi overnight. The mixture was
filtered on Celite and
the filtrate was evaporated to dryness.
Purified by flash chromatography on silica gel EtOH/CH2Cl2 (2 :98). Yield :
57%
1H NMR (DMSO-a6, 300 MHz) b: 7.08 (t, 1H, J = 8Hz), 6.61 (d, 1H, J = 7Hz),
6.54 (m, 2H),
6.17 (m, 2H), 2.56 (m, 2H), 2.36 (m, 2H), 1.67 (m, 4H).
5-(3-aminophenyl) pent-4-ynoic acid was then reacted with 2-
chloroethylisocyanate as described
in examples 1-12 to obtain desired product.
Purified by flash chromatography on silica gel EtOH/CHZCl2 (5 :95). Yield :
37%
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IH NMR (Acetone-d6, 300MHz) 8: 7.57 (brs, NH, 1H), 7.36 (m, Ar, 2H), 7.07 (m,
Ar, 2H), 6.87
(d, Ar, 1H, J = ,0), 6.74 (brs, NH, 1H), 3.66 (m, CH2, 4H), 3.46 (m, CHZ, 4H),
2.59 (m, CH2,
2H), 1.63 (m, CHZ, 2H).
EXAMPLE 52: Preparation of 5- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-
pentanoic acid
ethyl ester (49)
i O
O \ I N~N~CI
H H
To a mixture of 3-iodonitrobenzene (5 g, 20.3 mmoles), K2CO3 (7.03 g, 50.9
mmoles) in 80 mL
1,2-DME/water (1 :1) were added successively CuI (166.0 mg, 0.87 mmoles), PPh3
(427.11 mg,
1.82 mmoles), Pd/C 10% (432.06 mg, 0.406 mmoles). The mixture was stirred at
room
temperature for 1 hour. 5-pentanoic acid (5.90 g, 84.30 mmoles) was added,
then the mixture
was heated to reflux overnight. After cooling, the mixture was filtered on
Celite and the organic
layer was evaporated under reduced pressure. Tha aqueous layer was acidified
with concentrated
Chlorhydric acid and extracted with AcOEt. The organic layer were washed with
brine, dried,
filtered and evaporated.
Purified by flash chromatography on silica gel CH2Cl2/EtOH (95 :5). Yield :
45%
IH NMR (DMSO-a6, 300 MHz) 8: 12.39 (brs, 1H), 8.19 (d, 1H, J = 8Hz), 8.13 (s,
1H), 7.82 (d,
1H, J = 7.SHz), 7.66 (t, 1H, J = 8Hz), 2.63 (m, 4H).
A mixture of 5-(3-nitrophenyl)-pent-4-ynoic acid (250 mg, 1.14 mmoles), 4 mL
of dry EtOH, 8.5
mL of dry CHZCIz and 8 mg of APTS was stirred at reflux for 12 hours. After
cooling, the
mixture was evaporated under reduce pressure. The residue was dissolved in
saturated solution
Na2C03 and extracted with CH2C12. The organic layer was dried, filtered and
evaporated to
dryness. Purified by flash chromatography on silica gel CHZC12/EtOH (98 :2).
Yield : 87%
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1H NMR (CDC13, 300 MHz) 8: 8.14 (s, 1H), 8.05 (d, 1H, J = 8Hz), 7.61 (d, 1H, J
= 8Hz), 7.41
(m, 1H), 4.15 (q, 2H, J = 7Hz), 2.71 (m, 2H), 2.59 (m, 2H), 1.23 (q, 3H, J =
7Hz).
A mixture of 5-ethyl-(3-nitrophenyl)-pent-4-ynoic acid ester (100 mg, 0.404
mmoles), Pd/C 10%
(10 mg, 0.094 mmoles) was hydrogenated under 38 psi overnight. The mixture was
filtered on
Celite and the filtrate was evaporated to dryness. Purified by flash
chromatography on silica gel
EtOH/CH2Clz (2 :98). Yield : 57%
1H NMR (CDCl3, 300 MHz) 8: 7.06 (m, 1H), 6.58 (d, 1H, J = 7.SHz), 6.50 (m,
2H), 4.11 (q, 2H,
J = 7Hz), 2.54 (m, 2H), 2.32 (m, 2H), 1.64 (m, 4H), 1.26 (t, 3H, J = 7Hz).
5-ethyl-(3-aminophenyl)pent-4-ynoic acid ester was then reacted with 2-
chloroethylisocyanate as
described in examples 1-12 to obtain desired product. Purified by flash
chromatography on
silica gel EtOH/CHZC12 (5 :95). Yield : 77%
1H NMR (CDCl3, 300 MHz) b: 7.23 (m, Ar, 3H), 6.89 (m, Ar, 2H), 5.63 (brs, NH,
1H), 4.31 (q,
CH2, 2H, J = 7,0), 2.56 (m, CH2, 2H), 2.39 (m, CH2, 2H), 1.81 (m, CH2, 4H),
1.23 (t, CH3, 3H, J
= 7.0).
EXAMPLE 53: Preparation of 1-(2-Chloro-ethyl)-3-(3-cyanomethyl-phenyl)-urea
(50)
i I O
N~N~CI
H H
A mixture of nitrobenzene acetic acid (1 g, 5.52 mmoles), 1.66 mL of SOC12, 10
mL dry CHC13
was refluxed for 14 hours. CHC13 and excess thionyl chlorid were removed in
vacuo, and the
residue was evaporated twice with 25 mL of toluene to remove traces of thionyl
chlorid. The
residue was taken into 10 mL of toluene and 30 mL of cold concentrated
ammonium hydroxyde
were added. The white solid formed was collected and dried with etanol in
vacuo. Yield : 79 %.
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1H NMR (DMSO-d6, 300MHz) 8: 8.14 (m, 2H), 7.71 (m, 2H), 7.06 (brs, 2H), 3.58
(s, 2H).
2-(3-nitrophenyl) acetamide was added with 10 mL of POC13 and the mixture was
heated at
reflux for 2 hours. After cooling, the mixture was poured into ice, basified
with NaaC03 and
extracted with CHZCIz. The organic layer was dried over NaZS04, filtered and
evaporated.
Purified by flash chromatography on silica gel CHZCIz. Yield : 41
1H NMR (CDC13, 300 MHz) ~: 8.14 (m, 2H), 7.69 (d, 1H, J = 8Hz), 7.56 (m, 1H),
3.88 (s, 2H).
A solution of 15 mL HBr 48% was cooled to 0°C. (430 mg, 2.65 mmoles) of
3-nitrobenzonitrile,
(574 mg, 4.85 mmoles) of Sn were added successively. The mixture was stirred
at room
temperature for 3 hours, then poured into ice. The solution was basified with
Na2C03, extracted
with CHZC12, dried, filtered and evaporated. Purified by flash chromatography
on silica gel
EtOH/CH2C12 (2 :98). Yield : 32%
1H NMR (DMSO-d6, 300MHz) 8: 7.37 (m, 2H), 6.81 (m, 2H), 3.86 (s, 2H).
3-aminobenzonitrile was then reacted with 2-chloroethylisocyanate as described
in examples 1-
12 to obtain desired product. Purified by flash chrmatography on silica gel
EtOH/CH2C12
(5 :95). Yield : 78%
1H NMR (CDC13, 300 MHz) 8: 8.12 (brs, NH, 1H), 7.26 (m, Ar, 4H), 6.97 (brs,
NH, 1H), 3.63
(m, CH2, 6H).
EXAMPLE 54: Preparation of 2-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl{-acetamide
(51)
o ~ ~ o
H2N ~ N~N~CI
H H
CA 02568609 2006-11-28
A mixture of 2-(3-nitrophenyl) acetamide (510 mg, 2.83 mmoles), Pd/C 10% (29
mg, 0.272
mmoles), dry ehtanol was hydrogenated under 38 psi overnight. The mixture was
filtered on
Celite and the filtrate was evaporated to dryness. The crude product was then
reacted with 2-
chloroethylisocyanate as described in examples 1-12 to obtain desired product.
Purified by flash
chromatography on silica gel EtOHICHZC12 (10 :90). bYield : 37%.
1H NMR (DMSO-d6, 300MHz) 8: 8.17 (brs, NH, 1H), 8.12 (d, Ar, 1H, J = 8.2Hz),
7.65 (m, Ar,
3H), 7.03 (brs, NH, 1H), 3.68 (s, CH2, 2H), 3.45 (m, CH2, 4H).
The following exemplary compounds were also prepared.
Example 55: 1-(2-Chloro-ethyl)-3-m-tolyl-urea (52):
O
\ N~N~CI
H H
1H NMR (CDC13) 8: 7.20-7,12 (m, Ar, 3H), 6.90 (d, Ar, 1H, J = 7), 3.57 (m,
Ch2, 4H), 2.29 (s,
CH3, 3H). 13C l~IMR 8: 156.1, 139.4, 138.0, 129.2, 125.1, 122.1, 118.4, 44.5,
42.1, 21,4.
Example 56: 1-(2-Chloro-ethyl)-3-(3-ethyl-phenyl)-urea (53);
0
\ ( N~N~CI
H H
1H NMR (Acetone-d6) 8: 8.09 (brs, NH, 1H), 7.29 (m, Ar, 2H), 7.13 (t, Ar, 1H,
J = 7.8), 6.80 (d,
Ar, J = 7.8), 6.20 ( brs, NH, 1H), 2.54 (q, CH2, 2H, J = 7.1), 3.57 (m, CH2,
4H), 1.19 (t, CH3,
3H, J = 7). 13C NMR 8: 156.2, 145.5, 141.0, 129.3, 122.1, 118.7, 116.7, 44.8,
42.5, 29.0, 14.9.
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Example 57: 1-(2-Chloro-ethyl)-3-(3-methoxy-phenyl)-urea (54);
0
\ ~ N~N~CI
H H
1H NMR (CDC13) 8: 8.12 (brs, NH, 1H), 7.21 (m, Ar, 3H), 6.87 (d, Ar, 1H, J =
7), 3.97 (s, CH3,
3H), 3.57 (m, CH2, 4H). 13C NMR ~: 156.1, 139.4, 138.0, 129.2, 125.1, 122.1,
118.4, 57.8, 44.5,
42.1.
Example 58: 1-(2-Chloro-ethyl)-3-[4-(4-hydroxy-butyl)-phenyl]-urea (55);
HO
O
/ N~ SCI
N
H H
1H NMR (Acetone-d6) ~: 8.00 (brs, NH, 1H), 7.39 (d, CH2, 2H, J = 8.2), 7.07
(d, CH2, 2H, J =
8.2), 6.50 ((brs, NH, 1H), 3.51 (m, 8H), 2.57 (t, 2H, J = 7.5), 1.58 (m, 2H).
13C NMR 8: 155.9,
139.0, 136.3, 130.0, 119.2, 64.8, 42.8, 42.5, 42.4, 35.2.
Example 59: 1-(2-Chloro-ethyl)-3-[4-(3-hydroxy-propyl)-phenyl]-urea (56);
HO v ~ \ O
/ N~N~CI
H H
1H NMR: 8.23 (brs, NH, 1H), 7.30 (d, Ar, 1H, J = 7.9), 7.06 (d, Ar, 1H, J =
8.2), 6.36 (brs, Nh,
1H), 3.58 (m, 4H), 3.42 (m, 6H). 13C NMR 8: 155.2, 138.2, 134.0, 128.5, 118.0,
63.3, 44.5, 43.1,
35.7, 30.4.
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Example 60: 1-(2-Chloro-ethyl)-3-[4-(5-hydroxy-pentyl)-phenyl]-urea (57);
\ o
OH / N~N~CI
H H
1H NMR: 8.63 (brs, NH, 1H), 7.36 (d, 2H, J = 7.9), 7.13 (d, 2H, J = 7.9), 6.43
(brs, NH, 1H),
3.96 (t, 2H, J = 6.5), 3.57 (m, 6H), 2.57 (t, 2H, J = 7.5), 1.54 (m, 2H), 1.42
(m, 2H). 13C NMR 8:
155.2, 129.5, 128.4, 123.2, 117.9, 60.6, 43.4, 40.9, 34.4, 32.7, 25Ø
Example 61: 1-(2-Chloro-ethyl)-3-[3-(5-hydroxy-pent-1-ynyl)-phenyl]-urea (58);
\ o
/ N~N~CI
HO ~ H H
1H NMR (acetone-d6) 8: 8.12 (brs, Nh, 1H), 7.03 (m, 1H), 6.79 (d, Ar, 1H, J =
7), 6.69 (brs, Ar,
1H), 6.57 (d, Ar, 1H, j= 8.0), 3.74 (t, 2H, j = 7), 3.49 (m, 4H), 2.47 (t, 2H,
J = 7), 1.81 (t, 2H, J =
7), 1.23 (t, 2H, j=7). '3C NMR 8: 156.1, 146.3, 129.2, 125.3, 124.4, 122.0,
118.0, 114.9, 88.9,
81.3, 61.6, 45.1, 42.8, 31.4, 16Ø
Example 62: 3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid ethyl ester (59);
0
O / N~N~CI
I H H
O
1H NMR (CDC13) S: 8.22 (brs, Nh, 1H), 7.93 (s, Ar, 1H), 7.63 (d, Ar, 1H, J =
7,8), 7.57 (d, Ar,
1H, J = 7), 7.30 (m, 1H), 6.35 (brs, Nh, 1H), 4.31 (q, CH2, 2H, J = 7), 3.57
(m, 4H), 1.34 (t,
98
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CH3, 3H, J = 7). 13C NMR ~: 166.7, 156.0, 139.1, 131.0, 129.1, 124.3, 124.1,
120.5, 61.2, 44.5,
42.0, 14.2.
Example 63: 1-(2-Chloro-ethyl)-3-[3-(5-methoxy-pentyl)-phenyl]-urea (60);
\ o
/O / N~N~CI
H H
1H NMR (CDC13) ~: 8.18 (brs, Nh, 1H), 7.14 (m, 3H), 6.86 (d, Ar, 1H, J = 7,2),
3.57 (m, 4H),
3.33 (m, 4H), 2.55 (t, 2h, J = 7), 1.59 (m, 4H). 13C NMR 8: 156.0, 144.0,
138.4, 129.1, 123.9,
120.8, 118.2, 58.5, 44.6, 42.0, 35.9, 31.2, 29.5, 25.9.
Example 64: f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl-acetic acid (61);
o I \ o
HO / N~N~CI
H H
1H NMR (DMSO-d6) ~: 8.95 (brs, OH), 7.89 (s, 1H, Ar), 7.61 (d, 1H, J = 8.0),
7.41 (d, 1H, J =
7.9), 7.29m, 1H, Ar), 6.61 (brs, NH, 1H), 3.68 (m, 4H), 3.34 (s, 2H). 13C NMR
8: 168.8, 155.7,
148.6, 135.2, 128.5, 116.5, 114.7, 113.2, 47.3, 44.8, 38.2.
Example 65: 3-[3-(2-Chloro-ethyl)-ureido]-phenyl]-carboxamide (62);
\ o
HZN ~ / ~ ~CI
~N N
H H
O
99
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1H NMR (DMSO-d6) b: 8.92 (brs, NH2, 2H), 7.89 (brs, 1H, NH), 7.83 (s, 1H, Ar),
7.40 (d, 1H, J
= 7.9), 7.32 (m, 2H), 6.57 (brs, NH, 1H), 3.69 (m, 4H). 13C NMRB: 168.1,
155.1, 140.4, 135.1,
128.5, 120.5, 120.1, 117.3, 44.4, 41.3.
Example 66: 1-(2-Chloro-ethyl)-3-(3-heptyl-phenyl)-urea (63);
0
~cl
N~N
H H
1H NMR: 7.83 (brs, NH, 1H), 7.08 (m, 3H), 6.79 (d, Ar, 1H, J = 7), 6.23 (brs,
NH, 1H), 3.50 (m,
6H), 2.48 (t, Ar, 2H, J = 7), 1.21 (m, 8H). 13C NMR 8: 156.0, 144.0, 138.9,
128.8, 123.3, 120.0,
117.2, 61.1, 44.7, 43.9, 42.7, 41.9, 36.0, 31.7, 31.4, 29.3.
Example 67: 5-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-pentanoic acid amide
(64);
0
H2N ~ / ~ ~CI
v v -N N
H H
O
IH NMR (DMSO-d6) 8: 8.03(brs, NH, 1H, 7.11 (m, Ar, 3H), 6.69 (d, Ar, 1H, J =
7), 6.21 (brs,
NH, 1H), 3.47 (m, 4H), 3.34 (m, 2H), 2.67 (t, CH2, 2H, J = 7), 2.53 (m, 2H),
2.39 (t, CH2, 2H, J
= 7). '3C NMR 8: 172.3, 155.8, 147.9, 137.5, 130.3, 125.6, 124.7, 122.9, 58.3,
44.9, 41.2, 33.9,
29.3, 15Ø
Example 68: Pentanedioic acid mono-{3-[3-(2-chloro-ethyl)-ureido]-phenyl}
ester (65);
0 0 ( ~ o
~ ~cl
HO O N N
H H
100
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iH NMR (Acetone-d6) 8: 8.23 (brs, NH, 1H), 7.23 (m, Ar, 3H), 6.69 (m, Ar, 1H),
6.20 (brs, NH,
1H), 3.67 (m, 2H), 3.52 (m, 2H), 2.66 (m, 2H), 2.44 (m, 2H), 2.04 (m, 2H). 13C
NMR (Acetone-
d6) 8: 174.1, 171.9, 155.7, 152.2, 142.3, 129.9, 115.8, 115.6, 112.4, 44.7,
42.4, 33.6, 33.0, 20.8.
Example 69: 1-(2-Chloro-ethyl)-3-(3-cyano-phenyl)-urea (66);
O
\ N~N~CI
H H
1H NMR (DMSO-d6) b: 8.92 (brs, NH, 1H), 7.76 ( s, Ar, 1H), 7.40 (d, Ar, 1H, j
= 6.7), 7.18 ( d,
Ar, 1H, J = 7.2), 6.43 (brs, Mi, 1H), 3.51 (t, CH2, 2H, J = 7.0), 3.47 (t,
CH2, 2H, J = 7.0). 13C
NMR b: 154.8, 141.2, 130.0, 123.9, 122.6, 120.8, 118.9, 111.4, 44.8, 41.4.
Example 70: 3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid (67);
O
HO \ N~N~CI
O H H
1H NMR (DMSO-d6) 8: 12.87 (brs, OH, 1H), 8.91 (brs, NH, 1H), 8.08 ( s, Ar,
1H), 7.68 (d, Ar,
1H, J = 7.3), 7.61 (d, Ar, 1H, J = 7.5), 7.36 (m, Ar, 1H), 6.48 (brs, Nh, 1H),
3.68 (m, CH2, 2H),
3.45 m, CH2, 2H. 13C NMR 8: 167.4, 155.0, 140.6, 131.3, 128.9, 122.1, 121.9,
118.5, 44.3, 41.3.
Example 71: 5-{3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-pentanoic acid methyl
ester (68);
O
,O I / N~N~CI
H H
101
CA 02568609 2006-11-28
1H NMR (CDC13) 8: 8.21 (brs, NH, 1H), 7.07 (m, Ar, 1H), 6.57 (m, Ar, 3H), 6.17
(brs, NH, 1H),
3.66, (s, CH3, 3H), 3.51 (m, CH2, 4H), 2.58 (m, CH2, 2H), 2.33 (t, CH2, 2H, J
= 7.0), 1.65 (m,
CH2, 2H), 1.25 (m, CH2, 2H). 13C NMR 8: 174.1, 155.3, 146.3, 1434, 129.2,
118.8, 115.3,
112.8, 51.3, 44.7, 41.8, 35.5, 34.2, 30.4, 24.6.
Example 72: 1-(2-Chloro-ethyl)-3-(3-hexyl-phenyl)-urea (69);
I ~ O
N~N~CI
H H
1H NMR (CDC13) b: 7.58 (brs, NH, 1H), 7.11 (m, Ar, 3H), 6.79 (d, Ar, 1H, J =
69), 3.53 (m,
CH2, 4H), 3.46 (m, CH2, 4H), 2.48 (m, CH2, 2H), 1.20 (m, CH2, 8H). 13C NMR 8:
156.6,
144.0, 138.8, 128.8, 123.3, 120.0, 117.2, 61.1, 44.7, 43.9, 42.7, 41.9, 36.0,
31.8, 31.4, 29.3, 29.1,
22.6.
Example 73: 6- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-hexanoic acid amide
(70);
o I~ o
H2N / N~N~CI
H H
1H NMR (DMSO-d6) 8: 8.21 (brs, NH, 1H), 7.38 (m, Ar, 2H), 6.55 (m, Ar, 2H),
6.38 (brs, NH,
1H), 3.45 (m, CH2, 6H), 2.56 (m, CH2, 2H), 2.23 (m, CH2, 4H). 13C NMR 8:
172.3, 154.8,
137.4, 129.2, 123.7, 121.1, 117.2, 60.1, 44.7, 42.1, 35.9, 32.4, 30.9, 23.8.
Example 74: 6- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-hexanoic acid ethyl
ester (71);
o ~ O
I / N~N~CI
H H
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1H NMR (CDC13) 8: 7.14 (m, Ar, NH, 4H), 6.86 (d, Ar, 1H, J = 7,2), 5.66 (brs,
NH, 1H), 4.10
(q, CH2, 2H, J = 7.0), 3.59 (m, CH2, 4H), 2.54 (m, CH2, 2H), 2.27 (m, CH2,
2H), 1.62 (m, CH2,
4H), 135 (m, CH2, CH3, SH). 13C NMR 8: 174.9, 155.9, 143.9, 138.4, 129.1,
124.0, 120.9,
118.3, 60.3, 44.8, 42.0, 35.6, 34.3, 30.9, 29.7, 28.7, 24.8, 14.2.
Example 75: 6- f 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-hexanoic acid (72);
o iI o
HO \ N~N~CI
H H
1H NMR (Acetone-d6) 8: 8.17 (brs, NH, 1H), 7.31 (m, Ar, 1H), 7.12 (m, Ar, 2H),
6.79 (d, Ar,
1H, J = 7.4), 6.23 (brs, Nh, 1H), 3.64 (m, CH2, 2H), 3.54 (m, CH2, 2H), 2.55
(m, CH2, 2H),
2.20 (m, CH2, 2H), 1.71 (m, CH2, 4H), 1.22 (m, CH2, 2H). 13C NMR b: 174.1,
156.1, 143.9,
141.1, 129.2, 122.6, 119.2, 116.8, 44.9, 44.8, 42.5, 36.3, 34.1, 31.8, 30.6,
30.2, 25.4.
Example 76: 6-}3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-hexanoic acid methyl
ester (73);
o i1 o
\ N~N~CI
H H
1H NMR (CDC13) 8: 8.11 (brs, Nh, 1H), 7.48 (m, Ar, 2H), 6.51 (m, Ar, 2H),
6.38, (brs, Nh, 1H),
3.57 (m, CH2, 6H), 3.81 (s, CH3, 3H), 2.51 (m, CH2, 2H), 2.17 (m, CH2, 4H).
13C NMR 8:
174.1, 155.3, 138.3, 131.7, 124.2, 121.1, 116.8, 51.4, 44.6, 41.9, 35.9, 32.6,
30.8, 23.6.
Example 77: 3-[3-(2-Chloro-ethyl)-ureido]-benzoic acid methyl ester (74);
O
,O I / N~N~CI
O H H
103
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1H NMR (CDC13) 8: 8.21 (brs, NH, 1H), 7.87 (s, Ar, 1H), 7.61, Ar, 1H, J =
7.0), 7.58 (d, Ar, 1H,
J = 7.), 7.21 brs, Nh, 1H), 3.76 (s, CH3, 3H), 3.57 (m, CH2, 4H). 13C NMR 8:
167.1, 156.2,
139.3, 131.1, 129.0, 125.0, 123.9, 120.4, 52.2, 46.4, 42Ø
Example 78: 1-(2-Chloro-ethyl)-3-[3-(4-hydroxy-but-1-ynyl)-phenyl]-urea (75);
O
/ ~ N~N~CI
HO / H H
1H NMR (Acetone-d6) ~: 8.21 (brs, NH, 1H), 7.03 (m, Ar, 1H), 6.79 (d, Ar, 1H,
J = 7.0), 6.71 (s,
Ar, 1H), 6.57 (d, Ar, 1H, J = 7.9), 6.23 (brs, NH, 1H), 3.71 (m, CH2, 6H),
3.45 (m, CH2, 2H)
2.601 (t, CH2, 2H, J = 7.0). '3C NMR b: 155.8, 146.3, 129.3, 124.1, 122.1,
118.2, 115.2, 86.2,
82.5, 61.1, 44.7, 42.3, 23.7.
Example 79: 1-(2-Chloro-ethyl)-3-[3-(3-hydroxy-prop-1-ynyl)-phenyl]-urea (76);
HO ~ \ N~N~CI
H H
1H NMR (Acetone-d6) 8: 8.06 (brs, NH, 1H), 7.08 (m, Ar, 1H), 6.83 (d, Ar, 1H,
J = 7.5), 6.75 (s,
Ar, 1H), 6.63 (d, Ar, 1H J = 7.9), 3.57 (m, CH2, 4H), 2.04 (s, CH2, 2H). 13C
NMR 8: 155.2,
129.8, 123.3, 123.1, 118.0, 115.6, 86.7, 85.9, 51.6, 44.8, 42.3.
Example 80: 3-[3-(2-Chloro-ethyl)-ureido]-phenyl}-acetic acid ethyl ester
(77);
o ~I o
O \ N~N~CI
H H
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CA 02568609 2006-11-28
1H NMR (CDC13) 8: 7.89 (brs, NH, 1H), 7.17 (s, Ar, 1H), 7.08 (m, Ar, 2H), 6.81
(d, Ar, 1H, J =
7.9), 4.09 (q, CH2, 2H, J = 7,0), 3.46 (m, CH2, 6H), 1.21 (t, CH3, 3H, J =
7.0). 13C NMR 8:
172.1, 156.2, 139.2, 134.9, 129.7, 123.8, 120.8, 118.6, 61.1, 44.4, 42.0,
41.8, 14.1.
Example 81: Acetic acid 3- f 3-[3-(2-chloro-ethyl)-ureido]-phenyl}-propyl
ester (78).
o ~I o
~O ~ N~N~CI
H H
1H NMR (CDC13) 8: 7.57 (brs, NH, 1H), 7.21 (m, Ar, 3H), 6.79 (m, Ar, 1H), 5.91
(m, Ar, 1H),
3.57 (m, CH2, CH3, 9H). 13C NMR b: 172.5, 156.0, 139.1, 134.7, 129.2, 124.0,
120.9, 118.8,
52.1, 44.4, 40.9, 40.3.
EXAMPLE 82: Inhibition of Proliferation of Fibroblasts by Compound 1
Human foreskin fibroblasts, obtained by biopsy, were cultured in RPMI-1640
medium
supplemented with 10% fetal calf serum. Cytotoxicity was assessed using a
modified Alamar
Blue assay as described by Lancaster et al. (U.S. Patent No. 5,501,959).
Briefly, 2 x 103 cells
in 100 ~.l were seeded in 96-well plates and preincubated for 24 hours. After
addition of 100 ~1
fresh medium containing increasing concentrations of compound 1, cells were
incubated at 37°C
for 72 hours. The culture medium was removed, cells were washed with a
phosphate-buffered
saline (PBS) solution and replaced by 50 ~1 of a resazurin solution (resazurin
125 ~g/ml in
PBSaerum free RPMI 1:4). Cell survival was calculated from fluorescence
(exitation 485 nm;
emission 590 nm) measured with a FL 600 Reader (BioTek Instruments).
Cytotoxicity was
expressed as the dose required to inhibit cell growth by 50% (GISO). Values
are the means of at
least three independent determinations. GISO of compound 1 on human foreskin
fibroblasts was
12.5 ~M. Results of this assay are shown in Figure 1.
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CA 02568609 2006-11-28
EXAMPLE 83: Inhibition of Proliferation of HUVECs by Compound 1
Human umbilical vein endothelial cells, isolated from umbilical cord, were
cultured in M199
medium supplemented with Endothelial Cell Growth Supplement (ECGS, 20 ~.g/ml),
heparin
(0.09 g/1), , L-glutamine (0.292 g/1), 1 % antibiotic-antimycotic, and 10%
fetal calf serum.
Cytotoxicity was assessed using a modified Alamar Blue assay as described by
Lancaster et al.
(U.S. Patent No. 5,501,959). Briefly, 2 x 103 cells in 100 ~,1 were seeded in
gelatine-coated 96-
well plates and preincubated for 24 hours. After addition of 100 ~,1 fresh
medium containing
increasing concentrations of compound 1, cells were incubated at 37°C
for 72 hours. The culture
medium was removed, cells were washed with a phosphate-buffered saline (PBS)
solution and
replaced by 50 ~l of a resazurin solution (resazurin 125 ~,g/ml in PBSaerurn
free RPMI 1:4).
Cell survival was calculated from fluorescence (exitation 4~5 nm; emission 590
nm) measured
with a FL 600 Reader (BioTek Instruments). Cytotoxicity was expressed as the
dose required to
inhibit cell growth by 50% (GIso). Values are the means of at least three
independent
determinations. GIso of compound 1 on HUVECs was 3.1 ~,M. Results of this
assay are shown
in Figure 2.
EXAMPLE 84: Inhibition of Proliferation of HUVECs by Various Compounds of
Formula
I
Inhibition of HUVEC Proliferation by Various Compounds of Formula I was
determined as
outlined in Example ~3.
Table 1: Inhibition of HWEC Proliferation by Various Compounds of Formula I
Compound Glso
#
1 6.3
2 ~.4
3 7.8
4 72.6
17.1
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CA 02568609 2006-11-28
Conzpound Glso
#
6 5.4
8 15.3
13 63.0
14 18.8
21 1.6
(R)-24 27.2
1 GISO is the dose required to inhibit cell growth by 50%.
Table 2: Inhibition of HUVEC Proliferation by Various compounds of Formula I
R2
R3 ~ ~ R~ O Y
~ ~ /Z
R4 / N- _N' I v
H H
R5
Compound Rl R2 R3 R4 RS X Y Z Glsol
(R)-79 H methoxy methoxy H H H methyl Cl 30.7
(R-
isomer)
(S)-80 H methoxy methoxy H H H methyl Cl >100
(S-
isomer)
(R)-81 H H I H H H methyl Br 1.8
(R-
isomer)
(S)-82 H H n-heptylH H H n-propyl Cl 8.5
(S-isomer)
(S)-83 H H n-heptylH H H i-propyl Cl 10.4
(S- ( 14.02)
isomer)
84 H H n-heptylH H H Methyl' Cl 18.4
(R)-85 H H n-butyl H H H methyl Cl 9.5
(R-
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CA 02568609 2006-11-28
Conzpouud R~ R2 R3 R4 RS X Y Z Glso
isomer)
1 GISO is the dose required to inhibit cell growth by 50%.
Z Different values obtained in separate experiments.
3 Racemic mixture.
Example 85: Inhibition of HUVECs by CEUs in a dose- and time- dependent
manner.
Human umbilical vein endothelial cells (HUVECs) were isolated as previously
described.
HUVECs were maintained in a gelatin-coated 75-cm2 flask in M199 (Invitrogen,
Burlington,
ON) supplemented with 20 % fetal bovine serum (FBS), 100 units/ml penicillin,
100 ~,g/ml
streptomycin, 3 ng/ml basic fibroblast growth factor (bFGF) (Invitrogen,
Burlington, ON) and 5
units/ml heparin (Invitrogen, Burlington, ON).
HUVECs were inoculated into 96 well tissue culture plates in 100 ~,L
containing 2 X 103 cells
and were incubated at 37 °C. After 24 h, freshly solubilized drugs in
DMSO were diluted in
fresh medium. Aliquots of 100 ~,1 containing escalating concentration of drugs
(0.3 ~M to 100
~,M) were added to the appropriate microtiter wells already containing 100 ~1
of culture medium.
The cells were incubated for different period of time ranging from 3 h to 48
h. The supernatant
was removed, the cells were washed and incubated with fresh medium to complete
the total
incubation time to 48 h for each condition. Assays were stopped by addition of
cold TCA to the
wells (final concentration, 10 %), followed by their incubation for 60 min at
4 °C. The
supernatant was discarded and the plates were washed five times with tap water
and air-dried.
Sulforhodamine B solution (50 ~1) at 0.1 % (w/v) in 1 % acetic acid was added
to each well, and
plates were incubated for 15 min at room temperature. After staining, unbound
dye was removed
by washing five times with 1 % acetic acid and the plates were air-dried.
Bound stain was
solubilized with 10 mM trizma base, and the absorbance was read using a
~,Quant Universal
Microplate Spectrophotometer (Biotek, Winooski, VT) at 585 nm. The results
were compared
with those of a control reference plate fixed on the treatment day and the
growth inhibition
percentage was calculated for each drug contact period. The growth inhibition
percentage is
expressed as the mean of triplicates for each drug contact period, compared
with those of a
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CA 02568609 2006-11-28
control reference plate fixed on the day of the treatment.
The cytotoxicity of soft alkylant compound 1 its bioisosteric derivative,
compound 5 and its non-
alkylating counterpart, tBEU, was compared with that of a classical and strong
allcylating agent,
namely cisplatin (cDDP). The antimicrotubule agents colchicine, vinblastin and
paclitaxel were
also tested in this assay but was found not cytotoxic until they reach 48 h of
exposure (data not
shown). When they were in contact for less than 6 h with either cell lines,
virtually none of the
tested compounds (CEUs) showed inhibition of tumor cell growth and
proliferation. However, as
the time of contact between the tested compounds (CEUs) and tumor cells was
increased from 6
to 48 h, the comparable GISO of compound 1 and compound 5 markedly shifted to
the left hand-
side of the graph Figure 3A and B and this was shown in the low micromolar
range for all tumor
cell lines tested. These was strikingly different from the cDDP effect, which
displayed
cytotoxicity after only 3 h of treatment, with a GISO that was essentially in
the same range for all
time of contact tested (Figure 3D). Interestingly, at all concentrations
tested, the non-alkylating
tBEU showed no apparent cytotoxicity (Figure 3C), suggesting that the chemical
alkylating
property of candidate compounds was essential for their cytotoxicity. Overall,
soft alkylating
CEUs were as much cytotoxic as cDDP. However they require a longer time of
contact to
display their proliferative inhibitory activity, which is compatible to the
incubation time of other
anti-antimicrotubule agents tested, such as colchicine and paclitaxel (data
not shown).
EXAMPLE 86: Inhibition of IL-2 Production in Jurkat Cells by Compounds of
Formula I
#1
The human leukemic T-cell line Jurlcat was maintained in suspension culture in
RPMI 1640
medium supplemented with 10% (v/v) heat-inactivated fetal calf serum. The
cultures were kept
at 37°C in a humidified 95% air, 5% C02 atmosphere.
Quantification of IL-2 production from Jurkat T lymphocytes was performed
using a commercial
enzyme immunoassay (EIA) system (Cayman Chemical Company, USA). Cells were
grown to
log phase, harvested by centrifugation, counted with a hemacytometer, and
resuspended in fresh
media at 106 cells/ml. Then 96-well culture plates were prepared with SO~M of
inhibitor
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CA 02568609 2006-11-28
(compounds 1, 2 or 3 in dimethyl sulfoxide) alone or either T-cell receptor
(TCR)-independent
stimulants (80 nM phorbol myristate acetate (PMA) and 10 ~g/ml
phytohemagglutinin (PHA))
or TCR-independent stimulants (~0 nM PMA and 1 ~M ionomycin). Cells suspension
(150 ~1)
was added to each well and incubated for 24 h at 37°C. Culture
supernatants were harvested and
diluted 1:22 and 1:220 to bring the IL-2 concentrations within the linear
range of the EIA. EIA
detection was performed as suggested by manufacturer (Cayman Chemical) and
colormetric
reading determined on automatic plate reader. The results are presented in
Figure 4 and Table 3.
Autonomous cell growth has been showrn to be regulated in part by IL-2. The
ability of
compounds of Formula I to affect the inhibition of IL-2 production, therefore,
can help to control
cell proliferation.
Table 3: Effect of Compound of Formula I on IL-2 production on stimulated
Jurkat cells
PMA + PMA +
Compound
#
IorZOrraycin PHA
DMSO (control)100% 100%
1 67% 47%
2 40% 33%
3 6% 24%
EXAMPLE 87: Inhibition of IL-2 Production in Jurkat Cells by Compounds of
Formula I
#2
The ability of compounds 1, 2, 3 and 86 to inhibit IL-2 production in Jurkat
cells was assayed as
described above (Example ~6). The results are presented in Figures SA and SB.
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CA 02568609 2006-11-28
N N~ ~
H H ~ N~N~
86 87
EXAMPLE 88: Effect of haloethyl urea derivatives on IL-2 production on human
lympho-
mono cells.
Lympho-mono cells were isolated from human blood, and kept in suspension
culture in RPMI
1640 medium supplemented with 10% (v/v) heat-inactivated fetal calf serum. The
cultures were
kept at 37°C in a humidified 95% air, 5% C02 atmosphere for 24 hours.
Quantification of IL-2 production from human lympho-mono cells was performed
using a
commercial EIA system (eBiosciences, USA). Cells were counted with a
hemacytometer, and
resuspended in fresh media at 106 cells/mL. Cell suspension (150 ~L) was added
to each well of
a 96-well culture plates. IMO compounds were dissolved in dimethyl sulfoxide.
50~,L of
inhibitor (compouns 1, 2, 3, 86, 5, 30, 87 was added to each well to yield a
final concentration of
3, 10 and 30~M. The control consisted of the blank solution used to dissolve
the test compounds,
added to the cell suspension. The mixture of cell suspension and inhibitor was
incubated for 24 h
at 37°C. Ori the second day, stimulant (80 nM PMA and 10~,g/mL PHA) was
added to each well
and incubated for 24 h at 37°C. Culture supernatants were harvested and
diluted 1:22 and 1:220
to bring the IL-2 concentrations within the linear range of the EIA. EIA
detection was performed
as suggested by manufacturer (eBiosciences) and colorimetric reading
determined on automatic
plate reader. The cells, from which the supernatant was collected, were
exposed to resazurin
(Alamard blue) to measure the number of cells alive. This measurement is
performed to
discriminate between cell death (cell toxicity) and IL-2 decrease after
exposure to the different
compounds of the invention. Normally, at 30~,M, all the compounds of the
invention tested here
are non toxic. (The compounds toxicity arises solely with dividing cells).
Nevertheless, the
addition of PMA and PHA to the mixture of compounds of the invention and cell
suspension
made some compounds of the invention toxic for the non-dividing lympho-mono
cells.
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CA 02568609 2006-11-28
The results are shown in Figure 6. Figure 6a illustrates the total amount of
IL-2 measured in the
culture supernatant. Figure 6b illustrates number of cells alive as determined
by exposure to
resazurin. While Figure 6c illustrates Normalized amount of IL-2 measured in
the supernatant.
The normalization procedure consisted in the ratio of total IL-2 value (Figure
6a) by the number
of live cells (Figure 6b).
EXAMPLE 89: Inhibition of Proliferation of Keratinocytes by Compound 1
Cells from the human immortalized keratinocyte cell line HaCat were cultured
in RPMI-1640
medium supplemented with 10% fetal calf serum.
Cytotoxicity was assessed using a modified sulforhodamine B assay as described
by Rubinstein
et al (JNatl Cancer Inst 82:113-118, 1990). Briefly, 2 x 103 cells in 100 ~,L
were seeded in 96-
well plates and preincubated for 24h. After that, 100 ~,L of fresh medium
containing increasing
concentration of compound 1 were added. Cells were incubated at 37°C
for 72 hours. Cells were
then fixed ifz situ by the gentle addition of 50 ~,1 of cold 50 % (w/v) TCA
(final concentration, 10
TCA) and incubated for 60 minutes at 4°C. The supernatant was
discarded, and the plates
were washed five times with tap water and air dried. Sulforhodamine B (SRB)
solution (50 ~1 at
0.2 % (w/v) in 1 % acetic acid) was added to each well, and plates were
incubated for 30 minutes
at room temperature. After staining, unbound dye was removed by washing five
times with 1
acetic acid and the plates were air dried. Bound stain was subsequently
solubilized with 10 mM
Trizma base, and the absorbance was read on an automated plate reader at a
wavelength of 580
nrn. Cytotoxicity was expressed as the dose of drug required to inhibit cell
growth by 50%
(GISO). Values are the means of at least three independent determinations.
GISO of compound 1
on HaCat cells was 11.1 ~M. The results of this assay are presented in Figure
7.
EXAMPLE 90: Inhibition of Proliferation of Keratinocytes by Compounds of the
Invention
The ability of other compounds to inhibit the proliferation of keratinocytes
was assayed as
described above (Example 89). The results are presented in Table 4.
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CA 02568609 2006-11-28
O ~ p
\ N~N~Br \ I N~N~Br
H H H H
88 89
Table 4: Cytotoxicity of compounds of the invention on proliferative HaCat
cells
Co~rzpoufad Glso
#
1 11.1
2 25.6
3 15.0
6.9
R-23 3.0
S-23 78.0
27 6.2
88 6.7
89 13.2
30 0.9
14 22.5
1 GISO is the dose required to inhibit cell growth by 50%.
Compound 30 in the above Table is 4-ter-butyl-C(K-E)U.
EXAMPLE 91: The Effect of Compounds of Formula I on Proliferating and Non-
proliferating Keratinocytes by Compounds of Formula I
The effect of compounds 1, 2, 3 and 17 on the growth of proliferating and non-
proliferating
HaCat cells was studied and compared to control compounds (5-FU, colchicine,
taxol and
vinblastine). The results are presented in Figures 8A-H.
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CA 02568609 2006-11-28
EXAMPLE 92: Effect of 1-(4-tert-butyl-phenyl)-3-3(2-chloro-ethyl)-urea (tB-
CEU) on
growth of the human immortalized keratinocyte cell line HaCat
Cells from the human immortalized lceratinocyte cell line HaCat were cultured
in RPMI-1640
medium supplemented with 10% fetal calf serum.
Growth inhibition was assessed using a modified sulforhodamine B assay as
described by
Rubinstein et all. Briefly, 2 x 103 cells in~ 100~L were seeded in 96-well
plates and preincubated
for 24h. After that, 100~L of fresh medium containing increasing concentration
of the test drugs
were added. Cells were incubated at 37°C for 72 hours. Cells are then
fixed in situ by the gentle
addition of 50 ~1 of cold 50 % (w/v) TCA (final concentration, 10 % TCA) and
incubated for 60
minutes at 4°C. The supernatant is discarded, and the plates axe washed
five times with tap water
and air dried. Sulforhodamine B (SRB) solution (50 ~,1 at 0.2 % (w/v) in 1 %
acetic acid) is
added to each well, and plates are incubated for 30 minutes at room
temperature. After staining,
unbound dye is removed by washing five times with 1 % acetic acid and the
plates are air dried.
Bound stain is subsequently solubilized with 10 mM trizma base, and the
absorbance is read on
an automated plate reader at a wavelength of 580 nm. Growth Inhibition was
expressed as the
dose of drug required to inhibit cell growth by 50% (GIso). Number of
untreated cells were
arbitrary assign to 100% growth . Values are the means of at least three
independent
determinations.
Results are presented in Table 5 and Figure 9
Table 5 Growth Inhibition of haloethyl urea derivatives on proliferative HaCat
cells
Compound GIso
1 11.1
2 25.6
3 15.0
6.9
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CA 02568609 2006-11-28
(R)23 3.0
(S)23 78.0
90 6.7
91 13.2
92 0.9
93 22.5
O O ~ O
~Br ~ I ~ ~Br
N N N N
H H H H
90 91
O O ~O ~ O
N~N~CI ~N~N~CI
H H H H
92 93
EXAMPLE 93: Matrigel Plug Assay
The Matrigel plug assay was conducted to determine the ability of compound 5
to inhibit
angiogenesis using known procedures and suriname as a control. The results are
shown in Figure
10.
EXAMPLE 94: Figure 6. CEUs inhibit the bFGF-induced angiogenesis in the mice
Matrigel~ plug assay.
The experimental protocol was conducted as previously described Auerbach et
al. Pharmacol
Ther. 51:1-11 (1991); Ribatti et al. Int. J. Biol. Markers 14:207-213 (1999);
Stefansson et al. J.
Biol. Chem. 276:8135-8141)). Briefly, the liquid Matrigel~ (Becton Dickinson,
Bedford, Mass.)
was kept on ice until the inj ection to avoid gelling of the colloid. The
Matrigel~ solution was
115
CA 02568609 2006-11-28
either loaded with bFGF (250 ng/ml) to induce angiogenesis, or left unloaded
to be used as
negative controls. Five hundreds microliters of cold Matrigel~ were injected
subcutaneously on
the ventral line of mice. After the injection, the Matrigel~ preparation was
allowed to polymerize
for 5 min. The drugs (50 and 100 mg/kg; in DMA: transcutol: chremophor;
Tween20~ [1:3:3:3])
was then administered intraperitoneally, according to the method described by
Stefansson et al.
(J. Biol. Chem. 276:8135-8141)). After 5 to 7 days, the mice were euthanized
by COZ
asphyxiation, the Matrigel~ plugs were excised and only those that did not
show any sign of
hematoma were solubilized in PBS-Triton X-100 (0.1 %). The hemoglobin content
of Matrigel~
plugs was compared to a hemoglobin standard-curve (405 nm) to evaluate the
level of
vascularization Stefansson et al. J. Biol. Chem. 276:8135-8141)).The results
are normalized
according to the Matrigel° total dry weight. The results are
representative of three independent
experiments. The hemoglobin content of bFGF-containing Matrigel~ was used as
the 100
reference. ANOVA test revealed a significant difference between the groups (p
< 0.05) and the
Dunnett test was performed (t p < 0.05, when compared to Matrigel~ alone, *p<
0.05 when
compared to bFGF-containing plugs). For the sake of clarity, the results
corresponding to the
Matrigel~ without bFGF are not shown but revealed that a significant
vascularization was
induced by bFGF, after its addition to the Matrigel~ plugs.
Matrigel~ plugs containing bFGF induced a significant two-fold increase in
hemoglobin content
over Matrigel~ alone (Figure 11). When compound 1 and compound 5 were
administrated at 100
mg/kg, they were potent inhibitors of bFGF-induced blood vessel formation
(Figure 11), causing
a significant reduction of the hemoglobin content of 62 % and 58 %,
respectively. At the same
dose, tBEU clearly failed to reduce bFGF-induced vascularization, rather
showing an
hemoglobin content statistically similar to that from mice inj ected with the
vehicle or not
(untreated). Accordingly, the holoethyl urea compounds of the instant
invention have
antiangiogenic action in vivo. The mice injected with the test compounds of
the instant invention
showed no sign of toxicity
EXAMPLE 95: Cell Migration Assay
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CA 02568609 2006-11-28
The ability of compound 1 and compound 5 to inhibit the migration of
endothelial cells
(HUVECs) through an artificial membrane when submitted to the influence of a
chemoattractant
molecule was evaluated using Boyden's chambers and following published
protocols (see the
website for National Cancer Institute, Developmental Therapeutic Program,
Angiogenesis
Resource Center). Generally, a 48-well plate was fitted with collagen coated
membrane inserts (8
p.M pore size). The lower chamber of each well contained culture medium (29
~l) with or
without the chemoattractant bFGF (250 ng/ml). HUVECs in culture medium plus
BSA (45 p1)
were added to the upper chamber. Once the cells had attached to the membrane,
the test
compound was added. After a five-hour incubation at 37°C, the migrating
cells were fixed and
labelled. The results are presented in Figures 12A and B.
Example 96: Inhibition of endothelial cell migration by compounds 1 and 5
The chemotaxis motility of HUVECS was assessed using Boyden chambers. Briefly,
the
underside of TranswellTM migration chamber membranes (8.0- mm pore size) were
coated with
collagen IV as described previously (Filardo et al. J. Cell Biol. 130: 441-450
(1995); Klemke et
al. J. Cell Biol. 127:859-866 (1994)) and as modified by Petitclerc et al. (J.
Biol. Chem.
275:8051-8061 (2000). The HUVECs (A, B, C) were pre-treated ( ~ ) or not ( O )
for 16 h with
escalating concentrations of different drugs, then they were added to the top
of the TranswellTM
in a DMEM medium containing 0.1 % BSA, in the presence of drugs. Soluble
fibronectin (25
~,g/ml) was added to the lower chamber to induce chemotaxis. The cells were
allowed to migrate
for 4-6 h at 37 °C, fixed and stained for quatification. The number of
migrating cells per well
were counted. The results expressed the mean ~ s.e. of triplicates.
CEUs abrogate the motogenic potential of endothelial cells. Figure 13
demonstrates that the
CEUs inhibited HLJVEC cell migration with an overall efficiency of 30-40 %.
Because the
antiproliferative effect of CEUs depends on the time of contact, the migration
assays were
conducted with either 0 or 16 hours pretreatments.
EXAMPLE 97: Chick Embryo Chorioallantoic Membrane (CAM) Assay
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CA 02568609 2006-11-28
The ability of compound 1 to inhibit formation of a solid tumour was assessed
in the chick CAM
assay using published protocols (Brooks et al., in Methods ifa Molecular
Biology, Vol. 129, pp.
257-269 (2000), ed. A.R. Howlett, Humana Press Inc., Totowa, NJ). 10-day old
chick embryos
were inoculated with cells from the hamster melanoma cell line. After 11 days,
the test
compound was injected intravenously and the eggs were incubated for a further
6 days. Tumours
were then harvested and weighed. The results are presented in Figure 14.
Inhibition of tumour
growth by compound 1 appears to be mediated by inhibition of formation of a
vascular network.
EXAMPLE 98: Compounds of the instant invention impede the growth of two
unrelated
tumor cell lines in the chick chorioallantoic membrane (CAM) assay.
Human HT1080 fibrosarcoma and hamster CSl melanoma cell lines were used to
assess the
antitumoral activity of CEUs in the chick chorioallantoic assay (Petitclerc et
al. J. Biol. Chem.
275:8051-9061 (2000); Kim et al. Cell 94:353-362 (1998); Lyu et al. Int. J.
Cancer 77:257-263
(1998)). In brief, day-0 fertilized chicken eggs were purchased from Couvoirs
Victoriaville
(Victoriaville, QC, Canada). The eggs were incubated for 10 days in a Pro-FI
egg incubator fitted
with an automatic egg turner, before being transferred to a Roll-X static
incubator for the rest of
the incubation time (incubators were purchased from Lyon Electric, Chula
Vista, San Diego).
The eggs were kept at 37 °C in a 60 % humidity atmosphere for the whole
incubation period.
Using a hobby drill (Dremel; Racine, WI), a hole was drilled on the side of
the egg, and a
negative pressure was applied to create a new air sac. A window was opened on
this new air sac
and was covered with transparent adhesive tape to prevent contamination. A
freshly prepared cell
suspension (40 ~,l) of either HT1080 (3.5 x 105 cells/egg) or CS1 (5 x 106
cells/egg) cells was
applied directly on the freshly exposed CAM tissue through the window. On day
11, the tested
drugs were injected intravenously in 10-12 eggs in a small volume (100 ~,1).
The eggs were
incubated until day 17, at which time the embryos were euthanized at 4
°C, followed by
decapitation. Tumors were collected, pictures were taken to illustrate the
different groups and the
tumor-wet weights were recorded. In all experiments, the number of dead
embryos from the
different groups was monitored for any sign of toxicity.
Panel A and B of Figure 15 show that incubation of CS1-derived tumors on the
CAM with
its
CA 02568609 2006-11-28
compounds 1 and 5 resulted in a significant dose-dependent reduction of the
tumors size, as
observed also with cDDP (Figure 15D). Moreover, both CEUs also inhibited the
formation of
HT1080 tumor mass in the same concentration range (data not shown). It is
noteworthy that the
non-alkylating homologue tBEU failed to influence the growth of CS1 tumors
(Figure 15C),
supporting the assumption that the antitumoral effect of the compounds 1 and 5
was dependent
on their alkylating activity. In the same experimental settings, only 10
~g/egg of cDDP was
sufficient to inhibit tumor cell growth. However, a high level of chick embryo
toxicity was
observed at higher concentration, since 95 to 100 % of embryos died at a cDDP
concentration
reaching 50 ~,g/egg (Figure 15). By contrast, the antitumoral effect of tBCEU
and COMPOUND
was shown at doses that were well tolerated by the chick embryo, as we
monitored by chick
necropsy (up to 150 ~,g/egg, data not shown).
Cumulative results are shown from three independent experiments. ANOVA test
showed a
significant difference between doses (p< 0.01). Dunnett test was performed (*
p < 0.05, ** p<
0.01). # indicates 95-100 % chick embryos mortality in the group when using 25
~,g/ml cDDP.
In panel A), the black circle corresponds to the injection of the solvent used
to solubilize the
drugs.
EXAMPLE 99: Compounds 1, 5, 2 and 22 impede the growth of CS1 tumor cell line
in the
chick chorioallantoic membrane (CAM) assay.
The ability of compounds 1, 5, 2 and 30 to impede the growth of CS 1 tumor
cell line was
determined using the chick chorioallantoic membrane (CAM) assay as described
in Example 98.
The results are present in Figure 16.
Example 99: Microtubule depolymerization and cytoskeleton disruption induced
by CEUs.
Morphological analysis of microtubules, actin cytoskeleton and nucleus by
immunocytochemistry.
Human umbilical vein endothelial cells (HUVECs) were as previously described.
HUVECs
were maintained in a gelatin-coated 75-cm~ flask in M199 (Invitrogen,
Burlington, ON)
supplemented with 20 % fetal bovine serum (FBS), 100 units/ml penicillin, 100
~g/ml
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CA 02568609 2006-11-28
streptomycin, 3 ng/ml basic fibroblast growth factor (bFGF) (Invitrogen,
Burlington, ON) and 5
units/ml heparin (Invitrogen, Burlington, ON).
HUVECs were seeded at 1x105 cells in in 35-mm Petri dishes and incubated for
16. h at 37 °C.
After treatments with either the test compounds of the instant invention or
classical
antimicrotubule agents, the cells were washed twice with phosphate-buffered
saline (PBS, pH
7.4) and then fixed with 3.7 % formaldehyde in PBS for 20 min. After two
washes, the cells
were permeabilized and blocked with 0.1 % saponin and 3 % (w/v) BSA in PBS,
during 1 h at 37
°C. The cells were then further incubated during 1 h at 37 °C
with anti-tubulin (clone TUB2.1,
that is specific to (3-tubulin and does not cross-react with other-tubulin
isoforms; Sigma-Aldrich;
St-Louis, MO) (l: 200) in 0.1 % saponin and 3 % BSA in PBS. The cells were
washed three
times with PBS containing 0.05 % of Tween 20 and incubated 1 h at 37 °C
in blocking buffer
containing anti-mouse IgG Alexa-488 (1: 1000), DAPI (2.5 ~,g/ml in PBS) to
stain nuclei and
Rhodamine-labeled phalloidin (l: 600) to stain the actin cytoskeleton. The
observations were
made using a Nikon Eclipse E800 microscope (Tokyo, Japan) equipped with a 40X
objective.
Images were captured as 16 bit TIFF files with a Hamamatsu ORCA ER cooled (-
20°C) digital
camera (Photonics Management Management Corp., Bridgewater, N.J.) driven by
SimplePCI
AIC software (Compix Inc. C Imaging systems, Pennsylvania).
The effect of compounds 1, 5 and tBEU on the microtubule network was compared
by an
immunofluorescence staining of (3-tubulin (Figure 17) In comparison with
untreated cells, 100
~,M of tBCEU during 24 h considerably affected the (3-tubulin fibers (Figure
17), showing a
punctated (3-tubulin staining that lead to a microtubule depolymerization
phenotype. This was
established by a comparison with a 24 h cells exposure to paclitaxel (50 ~,M),
colchicine (25
~,M), or to vinblastine (5 ~.M), classical antimicrotubule agents having
opposite effect on
microtubule network by their non-covalent binding to (3-tubulin. Paclitaxel
stabilizes the
microtubules, thus inhibiting their depolymerization, whereas the others
rather blocking their
polymerization, inducing therefore a depolymerization phenotype (Figure 17).
In fact, the
tBCEU effect on (3-tubulin was indeed drastically different from what was
observed after
120
CA 02568609 2006-11-28
paclitaxel treatment, not as severe as vinblastine, but rather similar to that
observed after
colchicine cell exposure. As expected, the bioisosteric derivative compound 5
showed a similar
microtubule dissolution activity as tBCEU. On the contrary, tBEU did not
exhibit any effect on
the microtubule network nor did affected the filamentous structure of actin.
However, tBCEU,
compound 5, colchicine and vinblastine considerably decreased the amounts of
filamentous actin
in both cell lines, presumably as an indirect consequence of (3-tubulin
depolymerization, showing
a more punctate actin distribution. This collapse of the actin structure was
not observed in
response to paclitaxel. Interestingly, the toxic effect of cDDP on (3-tubulin
and on actin filaments
seemed rather associated to its pro-apoptotic mechanism, actin and
microtubules being dissolved
only in cells showing a typical apoptotic nuclear fragmentation phenotype
Deschesnes et al.,
Mol. Biol. Cell. 12:1569-1582 (1996); Desbiens et al., Biochem. J. 372:631-641
(2003). Stress
fibers were rather observed in cDDP-treated cells that are still non-
apoptotic. This was still
contrasting with CEUs that were rather inducing a non-classical nuclear
condensation phenotype
consequently or in parallel to their microtubules disruption effect.
The invention being thus described, it will be obvious that the same may be
varied in many
ways. Such variations are not to be regarded as a departure from the spirit
and scope of the
invention, and all such modifications as would be obvious to one skilled in
the art are intended to
be included within the scope of the following claims.
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