Note: Descriptions are shown in the official language in which they were submitted.
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METHOD FOR TREATING CHRONIC PAIN
USING MEK INHIBITORS
BACKGROUND
The invention features a method for treating chronic pain using MEK
inhibitors. Chronic pain includes neuropathic pain, and chronic inflammatory
pain.
Abnormality anywhere in a nerve pathway disrupts nerve signals, which
in turn are abnormally interpreted in the brain, causing neuropathic pain.
Neuropathic pain may be, for example, a deep ache, a burning sensation, or
hypersensitivity to touch. Diseases or conditions associated with neuropathic
pain include, without limitation, diabetic neuropathy, causalgia, plexus
avulsion, neuroma, vasculitis, crush injury, viral infections (e.g., herpes
virus
infection or HIV), constriction injury, tissue injury, nerve injury from the
periphery to the central nervous system, limb amputation, hypothyroidism,
uremia, chronic alcoholism, post-operative pain, arthritis, back pain, and
vitamin deficiencies.
Infections such as herpes zoster (shingles) can cause nerve
inflammation and produce postherpetic neuralgia, a chronic burning localized
to the area of viral infection. Hyperalgesia is when an already noxious
stimulus becomes more painful, and allodynia, when a previously non-noxious
stimulus becomes painful (such as contact of clothing or a breeze). Reflex
sympathetic dystrophy is accompanied by swelling and sweating or changes
in local blood flow, tissue atrophy, or osteoporosis. Causalgia, including
severe burning pain and swelling, sweating, and changes in blood flow, may
follow an injury or disease of a major nerve such as the sciatic nerve. Some
types of chronic low back pain can have a neuropathic component (e.g.,
sciatica, postpoliomyelitis and CPRM). Neuropathic pain may also be induced
by cancer or chemotherapy.
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Neuropathic pain is currently treated with anticonvulsants such as
carbamazepine and antidepressants such as amitryptaline. NSAIDS and
opioids generally have little effect (Fields et al 1994 Textbook of Pain p 991-
996 (pub: Churchill Livingstone), James & Page 1994
J.Am.Pediatr.Med.Assoc, 8: 439-447, Galer, 1995 Neurology 45 S17-S25.
Neuropathic conditions that have been treated with gabapentin include:
postherpetic neuralgia, postpoliomyelitis, CPRM, HIV-related neuropathy,
trigeminal neuralgia, and reflex sympathetic dystrophy (RSD).
The generally weak efficacy of antiinflammatory agents suggests that the
mechanism for chronic pain is separate from hyperalgesia.
SUMMARY OF THE INVENTION
The invention features a method for treating chronic pain, which
method includes the step of administering a composition including a MEK
inhibitor to a patient in need of such treatment. Chronic pain includes
neuropathic pain, idiopathic pain, and pain associated with vitamin
deficiencies, uremia, hypothyroidism post-operative pain, arthritis, back
pain,
and chronic alcoholism. The invention also features compositions as
disclosed, formulated for the treatment of chronic pain. Such a composition
may include one or more MEK inhibitor compounds having a structure
disclosed in patent applications USSN 60/115,873, filed January 13, 1999,
PCT/US99/30483, international filing date December 21, 1999.
Examples of MEK inhibitors include a compound having the formula (I)
below:
O Rio
Q
R~ 1 I
2
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In formula (I), W is ORS, NR20R,, NRARB, NR2NRARB, O(CH2)2~NRARB, or
NR2(CH2)2.~ NRARB. R~ is H, C ~_$ alkyl, C 3_8 alkenyl, C 3_8 alkynyl, C 3_a
cycloalkyl, phenyl, (phenyl)C ~~ alkyl, (phenyl)C 3_4 alkenyl, (phenyl)C 3-a
alkynyl, (C 3_$ cycloalkyl)C » alkyl, (C 3_$ cycloalkyl)C 3~ alkenyl, (C 3_8
cycloalkyl)C 3_4 alkynyl, C 3_8 heterocyclic radical, (C 3_8 heterocyclic
radical)C
~_4 alkyl, (C 3_$ heterocyclic radical)C 3~ alkenyl, (C 3_$ heterocyclic
radical)C 3.~
alkynyl or (CH2)2_4 NR~Rp. R2 is H, C ~_4 alkyl, phenyl, C 3_s cycloalkyl, C
3_s
heterocyclic radical, or (C 3_s cycloalkyl) methyl. RA is H, C ~_s alkyl, C
3_$
alkenyl, C 3_$ alkynyl, C 3_$ cycloalkyl, phenyl, (C 3_8 cycloalkyl)C ,~
alkyl, (C 3_8
cycloalkyl)C 3~ alkenyl, (C 3_$ cycloalkyl)C 3~ alkynyl, C 3_8 heterocyclic
radical,
(C 3_$ heterocyclic radical)C ,_4 alkyl, (aminosulfonyl)phenyl,
[(aminosulfonyl)phenyl]C ~_4 alkyl, (aminosulfonyl)C ~_s alkyl,
(aminosulfonyl)C
3-s cycloalkyl, [(aminosulfonyl)C 3_s cycloalkyl]C ~_4 alkyl, or (CH2)2~
NRcRp. R8
is H, C ~_8 alkyl, C 3_8 alkenyl, C 3_8 alkynyl, C 3_8 cycloalkyl, or phenyl.
I S Q is one of the following formulae (i) - (iii):
R3 / \ R3 ~ \
\ / ),
Z R4 R4
Z~
(i) (ii) (iii)
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R3 is H or F; R4 is halo, NOz, S02NRo(CHz)z~NRERF, SOzNRERF or (CO)T. T
is C ~_$ alkyl, C 3_$ cycloalkyl, (NRERF)C ,_4 alkyl, ORF, -NRo(CHz)2~ NRERF,
or
NRERF; Z is one of the following formulae (iv) - (viii):
N N N~ X~ X2\/ N
~R~ ~R~
Rs R6
(iv) (v) (vi)
N. .N ,N. :N
X3 R~ N
(vii) (viii)
One of R5 and R6 is H or methyl and the other of R5 and R6 is H, C~_s alkyl,
Cz_6 alkenyl, Cz_s alkynyl, phenyl, benzyl, or -M-E-G. M is O, CO, SOz, NR~,
(CO)NRH, NRH (CO), NRH (SOz), (SOz)NRH, or CHz. E is (CHz),~ or (CHz)m
O(CHz)P where 1 <_ (each of m and p) <_ 3 and 2 <_ (m + p) 5 4; or E is
absent.
G is RK, OR, or NR~R,c, provided that if p = 1, then G is H. R7 is H, C ~_a
alkyl,
C z~ alkenyl, C z~ alkynyl, C 3_6 cycloalkyl, phenyl, 2-pyridyl, 3-pyridyl,
4-pyridyl, (CHz)~_zAr, where Ar is phenyl, 2-pyridyl, 3-pyridyl, or 4-pyridyl,
S02NRH(CHz)z~ NR~RK, (CO)(CHz)z~NR~RK or (CO)NRH(CHz)z~NR~RK. X, is
O, S, NRB, or CHR9; Xz is O, S, or CHR9; and X3 is O or S. In one
embodiment, if X~ or Xz is CHR9, the disclosed compound may also be a
tautomerized indole. R$ is H, C ~~ alkyl, phenyl, 2-pyridyl, 3-pyridyl, 4-
pyridyl,
(CHz)~_zAr, where Ar is phenyl, 2-pyridyl, 3-pyridyl, or 4-pyridyl, C z~
alkenyl,
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C 2~ alkynyl, C 3_6 cycloalkyl, or (C z~ alkyl)NR~RM provided R~ and R8
together have no more than 14 carbon atoms, exclusive of R~, RM, R~ and RK.
Rc is C ~_4 alkyl, phenyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, C 3~ alkenyl, C 3~
alkynyl, C 3_6 cycloalkyl, (CO)ORP , (C 2_4 alkyl)NR~RM, (CO)NRN(CH2)2_
4NR~RM, (CO)NR~RM, (CO)(CH2)2~-NR~RM, or (CH2)~_2Ar, where Ar is phenyl,
2-pyridyl, 3-pyridyl, or 4-pyridyl. R9 is C ~~ alkyl, phenyl, 2-pyridyl, 3-
pyridyl,
4-pyridyl, C 2~ alkenyl, C 2~ alkynyl, C 3_6 cycloalkyl, (CO)ORP ,
(C 2~ alkyl)NR~RM, (CO)NRN(CHZ)2~NR~RM, (CO)NR~RM, (CO)(CH2)2-a-
NR~Rnn, or (CH2)~_2Ar', where Ar' is phenyl, 2-pyridyl, 3-pyridyl, or 4-
pyridyl.
RP is H, C ~_6 alkyl, phenyl, C 3~ alkenyl, C 3~ alkynyl, C 3_6 cycloalkyl, or
(CH2)2~ NR~Rnn; Rio is H, methyl, halo, or N02; R,~ is H, methyl, halo, or
N02.
Each of Rc, Rp, RE, RF, R,, R~, RK, R~ and RM is independently selected from
H, C ,~ alkyl, C 3_4 alkenyl, C 3~ alkynyl, C 3_6 cycloalkyl, and phenyl; each
of
NRcRo,NRERF, NR~RK, and NR~RM can also independently be morpholinyl,
piperazinyl, pyrrolidinyl, or piperadinyl. Each of RH, RN, and Ro is
independently H, methyl, or ethyl. Finally, each hydrocarbon radical or
heterocyclic radical above is optionally substituted with between 1 and 3
substituents independently selected from halo, C ,~ alkyl, C 3_6 cycloalkyl, C
2_
4 alkenyl, C 2~ alkynyl, phenyl, hydroxyl, amino, (amino)sulfonyl, and N02,
wherein each substituent alkyl, cycloalkyl, alkenyl, alkynyl or phenyl is in
turn
optionally substituted with between 1 and 3 substituents independently
selected from halo, C 1_2 alkyl, hydroxyl, amino, and N02. In addition to the
above compounds, the invention also provides a pharmaceutically-acceptable
salt or C ~_~ ester thereof.
Preferred embodiments of the invention include methods using one or
more of the following compounds:
(a) said MEK inhibitor has a structure selected from: 7-fluoro-6-(4-iodo-2-
methyl-phenylamino)-1 H-benzoimidazole-5-carboxylic acid
cyclopropylmethoxy-amide; 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-6,7-
dihydro-1 H-benzoimidazole-5-carboxylic acid (hydrochloride); 7-fluoro-6-(4-
iodo-2-methyl-phenylamino)-1 H-benzoimidazole-5-carboxylic acid; 7-fluoro-6-
(4-iodo-2-methyl-phenylamino)-3H-benzoimidazole-5-carboxylic acid
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(2-hydroxy-ethoxy)-amide; 6-(2-chloro-4-iodo-phenylamino)-7-fluoro-1H-
benzoimidazole-5-carboxylic acid; and 7-fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylic acid pentafluorophenyl ester;
and (b) said MEK inhibitor has a structure selected from: 7-fluoro-6-(4-iodo-2-
methyl-phenylamino)-1 H-benzoimidazole-5-carboxylic acid
cyclopropylmethoxy-amide; and 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-3H-
benzoimidazole-5-carboxylic acid (2-hydroxy-ethoxy)-amide.
The invention also relates to a pharmaceutical composition including
(a) a benzoheterocycle (e.g., of formula I) and (b) a pharmaceutically-
acceptable carrier.
BRIEF DESCRIPTION OF THE FIGURES
FIG. 1 is a bar graph representing the paw withdrawal threshold (PWT)
in grams as a function of time in days. The empty, cross-hatched, and single
hatched bars are vehicle, PD 198306, and pregabalin, respectively. The
arrows indicate time of drug administration (30 mg/kg, p.o.).
FIG 2. is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days.
Baseline (BL) measurements were taken before treatment. Animals received
a single p.o. administration of PD 198306 (3-30mg/kg), or pregabalin
(30mg/kg) and withdrawal thresholds were re-assessed 1 h after treatment.
Treatments were repeated twice a day for two days. Results are expressed
median ~ 1St and 3~d quartiles. *P<0.05, **P<0.01, ***P<0.001 significantly
different from vehicle treated animals (Mann-Whitney t test; n=7-8).
FIG. 3. is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days.
Baseline (BL) measurements were taken before treatment. Animals received
a single p.o. administration of PD 198306 (3-30mg/kg), or pregabalin
(30mg/kg) and withdrawal thresholds were re-assessed 1 h after treatment.
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Treatments were repeated twice a day for two days. Results are expressed
median ~ 1St and 3'd quartiles. **P<0.01 significantly different from vehicle
treated animals (Mann-Whitney t test; n=6).
FIG. 4. is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days.
Baseline (BL) measurements were taken before treatment. Animals received
a single i.t. administration of PD 198306 (1-30~,g/10~.1), or pregabalin
(100pg/10p1) and withdrawal thresholds were re-assessed at 30min, 1 h and
2h after treatment. Results are expressed median ~ 1 St and 3'd quartiles.
*P<0.05, ***P<0.001 significantly different from vehicle treated animals (Mann-
Whitney t test; n=7-9).
FIG. 5. is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days.
Baseline (BL) measurements were taken before treatment. Animals received
a single i.t. administration of PD 198306 (1-30pg/10p1), or pregabalin
(100~g/10p1) and withdrawal thresholds were re-assessed at 30min, 1 h and
2h after treatment. Results are expressed median ~ 1 St and 3'd quartiles.
*P<0.05, **P<0.01, ***P<0.001 significantly different from vehicle treated
animals (Mann-Whitney t test; n=6-8).
FIG. 6 is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days .
Animals received a single intraplantar (i.pl.) administration of PD 198306
(3mg/100~1), or an intrathecal injection of PD 198306 (30~g/10p1) and
withdrawal thresholds were re-assessed 1 h after treatment. Results are
expressed median ~ 1St and 3'd quartiles. **P<0.01 significantly different
from
vehicle treated animals (Mann-Whitney t test; n=6-9).
JO
FIG. 7. is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments as a function of time in days.
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Animals received a single intraplantar (i.pl.) administration of PD 198306
(3mg/100~1), or an intrathecal injection of PD 198306 (30~g/10~1) and
withdrawal thresholds were re-assessed 1 h after treatment. Results are
expressed median ~ 1St and 3'd quartiles. **P<0.01 significantly different
from
vehicle treated animals (Mann-Whitney t test; n=6).
FIG. 8 is a bar graph representing the force required in grams to elicit
paw withdrawal using von Frey hair filaments. Baseline (BL) measurements
were taken before treatment. Animals received a single i.t. administration of
PD219622, PD297447, PD 184352, or PD 254552 (30~g/10~1), or pregabalin
(100~g/10~1) and withdrawal thresholds were re-assessed at 30min, 1h and
2h after treatment. Results are expressed median ~ 1St and 3'd quartiles.
*P<0.05, **P<0.01, ***P<0.001 significantly different from vehicle treated
animals (Mann-Whitney t test; n=7-8).
DETAILED DESCRIPTION
The compounds disclosed herein are pharmaceutically active, for
example, they inhibit MEK. MEK enzymes are dual specificity kinases
involved in, for example, immunomodulation, inflammation, and proliferative
diseases such as cancer and restenosis.
Proliferative diseases are caused by a defect in the intracellular
signaling system, or the signal transduction mechanism of certain proteins.
Defects include a change either in the intrinsic activity or in the cellular
concentration of one or more signaling proteins in the signaling cascade . The
cell may produce a growth factor that binds to its own receptors, resulting in
an autocrine loop, which continually stimulates proliferation. Mutations or
overexpression of intracellular signaling proteins can lead to spurious
mitogenic signals within the cell. Some of the most common mutations occur
in genes encoding the protein known as Ras, a G-protein that is activated
when bound to GTP, and inactivated when bound to GDP. The above-
mentioned growth factor receptors, and many other mitogenic receptors, when
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activated, lead to Ras being converted from the GDP-bound state to the GTP-
bound state. This signal is an absolute prerequisite for proliferation in most
cell types. Defects in this signaling system, especially in the deactivation
of
the Ras-GTP complex, are common in cancers, and lead to the signaling
cascade below Ras being chronically activated.
Activated Ras leads in turn to the activation of a cascade of
serine/threonine kinases. One of the groups of kinases known to require an
active Ras-GTP for its own activation is the Raf family. These in turn
activate
MEK (e.g., MEK1 and MEK2) which then activates MAP kinase, ERK (ERK~
and ERK2). Activation of MAP kinase by mitogens appears to be essential for
proliferation; constitutive activation of this kinase is sufficient to induce
cellular
transformation. Blockade of downstream Ras signaling, for example by use of
a dominant negative Raf-1 protein, can completely inhibit mitogenesis,
whether induced from cell surface receptors or from oncogenic Ras mutants.
Although Ras is not itself a protein kinase, it participates in the activation
of
Raf and other kinases, most likely through a phosphorylation mechanism.
Once activated, Raf and other kinases phosphorylate MEK on two closely
adjacent serine residues, S21$ and S222 in the case of MEK-1, which are the
prerequisite for activation of MEK as a kinase. MEK in turn phosphorylates
MAP kinase on both a tyrosine, Y185, and a threonine residue, T183,
separated by a single amino acid.
This double phosphorylation activates MAP kinase at least 100-fold.
Activated MAP kinase can then catalyze the phosphorylation of a large
number of proteins, including several transcription factors and other kinases.
Many of these MAP kinase phosphorylations are mitogenically activating for
the target protein, such as a kinase, a transcription factor, or another
cellular
protein. In addition to Raf-1 and MEKK, other kinases activate MEK, and
MEK itself appears to be a signal integrating kinase. Current understanding
is that MEK is highly specific for the phosphorylation of MAP kinase. In fact,
no substrate for MEK other than the MAP kinase , ERK, has been
demonstrated to date and MEK does not phosphorylate peptides based on
the MAP kinase phosphorylation sequence, or even phosphorylate denatured
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MAP kinase. MEK also appears to associate strongly with MAP kinase prior
to phosphorylating it, suggesting that phosphorylation of MAP kinase by MEK
may require a prior strong interaction between the two proteins. Both this
requirement and the unusual specificity of MEK are suggestive that it
may have enough difference in its mechanism of action to other protein
kinases that selective inhibitors of MEK, possibly operating through
allosteric
mechanisms rather than through the usual blockade of the ATP binding site,
may be found.
The effect of the MEK inhibitor PD 198306 has been investigated in two
animal models of neuropathic pain by assessing static allodynia with von Frey
hairs.
Oral administration of PD 198306 (3-30mg/kg) had no effect in the model
of chronic constriction injury of the sciatic nerve (CCI). However, after
repeated
administration (3 doses over two days) it had a transient effect in the
diabetic
neuropathy model (streptozocin). This may be due to disorders of the blood-
brain barrier induced by the diabetic condition in these animals, thus
allowing
central action of the compound. Intrathecal administration of PD 198306 (1-
30pg) dose-dependently blocked static allodynia in both the streptozocin and
the
CCI models of neuropathic pain, with minimum effective doses (MED) of 3 and
10~g respectively. The highest dose used (30pg) totally blocked the
maintenance of static allodynia, for up to 1 h. Intraplantar administration of
PD
198306 (3mg/100~1) at a dose 100-fold higher than the dose shown to be
effective intrathecally (30~g/10~1) had no effect on static allodynia in
either of the
neuropathic pain models. This finding confirms the lack of effect seen after
systemic administration and suggests a central site of action for the
compound.
From this study we can suggest the use of MEK inhibitors as potential
new therapeutic tools for chronic pain. The study of potential side-effects,
especially related to memory, of future brain-penetrant MEK inhibitors will
indicate the therapeutic window for this novel class of compounds in the
treatment of pain.
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A. Terms
Certain terms are defined below and by their usage throughout this
disclosure.
Alkyl groups include aliphatic (i.e., hydrocarbyl or hydrocarbon radical
structures containing hydrogen and carbon atoms) with a free valence. Alkyl
groups are understood to include straight chain and branched structures.
Examples include methyl, ethyl, propyl, isopropyl, butyl, n-butyl, isobutyl, t-
butyl, pentyl, isopentyl, 2,3-dimethylpropyl, hexyl, 2,3-dimethylhexyl, 1,1-
dimethylpentyl, heptyl, and octyl. Cycloalkyl groups include cyclopropyl,
cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
Alkyl groups can be substituted with 1, 2, 3 or more substituents which
are independently selected from halo (fluoro, chloro, bromo, or iodo),
hydroxy,
amino, alkoxy, alkylamino, dialkylamino, cycloalkyl, aryl, aryloxy,
arylalkyloxy,
heterocyclic radical, and (heterocyclic radical)oxy. Specific examples include
fluoromethyl, hydroxyethyl, 2,3-dihydroxyethyl, (2- or 3-furanyl)methyl,
cyclopropylmethyl, benzyloxyethyl, (3-pyridinyl)methyl, (2- or 3-
furanyl)methyl,
(2-thienyl)ethyl, hydroxypropyl, aminocyclohexyl, 2-dimethylaminobutyl,
methoxymethyl, N-pyridinylethyl, diethylaminoethyl, and cyclobutylmethyl.
Alkenyl groups are analogous to alkyl groups, but have at least one
double bond (two adjacent sp2 carbon atoms). Depending on the placement
of a double bond and substituents, if any, the geometry of the double bond
may be entgegen (E), or zusammen (Z), cis, or trans. Similarly, alkynyl
groups have at least one triple bond (two adjacent sp carbon atoms).
Unsaturated alkenyl or alkynyl groups may have one or more double or triple
bonds, respectively, or a mixture thereof; like alkyl groups, unsaturated
groups
may be straight chain or branched, and they may be substituted as described
both above for alkyl groups and throughout the disclosure by example.
Examples of alkenyls, alkynyls, and substituted forms include cis-2-butenyl,
trans-2-butenyl, 3-butynyl, 3-phenyl-2-propynyl, 3-(2'-fluorophenyl)-2-
propynyl,
3-methyl(5-phenyl)-4-pentynyl, 2-hydroxy-2-propynyl, 2-methyl-2-propynyl, 2-
propenyl, 4-hydroxy-3-butynyl, 3-(3-fluorophenyl)-2-propynyl, and 2-methyl-2-
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propenyl. In formula (I), alkenyls and alkynyls can be C 2~ or C 2_8, for
example, and are preferably C 3~ or C 3_8.
More general forms of substituted hydrocarbon radicals include
hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxycycloalkyl, hydroxyaryl,
and corresponding forms for the prefixes amino-, halo- (e.g., fluoro-, chloro-
,
or bromo-), nitro-, alkyl-, phenyl-, cycloalkyl- and so on, or combinations of
substituents. According to formula (I), therefore, substituted alkyls include
hydroxyalkyl, aminoalkyl, nitroalkyl, haloalkyl, alkylalkyl (branched alkyls,
such
as methylpentyl), (cycloalkyl)alkyl, phenylalkyl, alkoxy, alkylaminoalkyl,
dialkylaminoalkyl, arylalkyl, aryloxyalkyl, arylalkyloxyalkyl, (heterocyclic
radical)alkyl, and (heterocyclic radical)oxyalkyl. R~ thus includes
hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxycycloalkyl, hydroxyaryl,
aminoalkyl, aminoalkenyl, aminoalkynyl, aminocycloalkyl, aminoaryl,
alkylalkenyl, (alkylaryl)alkyl, (haloaryl)alkyl, (hydroxyaryl)alkynyl, and so
forth.
I S Similarly, RA includes hydroxyalkyl and aminoaryl, and Rg includes
hydroxyalkyl, aminoalkyl, and hydroxyalkyl(heterocyclic radical)alkyl.
Heterocyclic radicals, which include but are not limited to heteroaryls,
include: furyl, oxazolyl, isoxazolyl, thiophenyl, thiazolyl, pyrrolyl,
imidazolyl,
1,3,4-triazolyl, tetrazolyl, pyridinyl, pyrimidinyl, pyridazinyl, indolyl, and
their
nonaromatic counterparts. Further examples of heterocyclic radicals include
piperidyl, quinolyl, isothiazolyl, piperidinyl, morpholinyl, piperazinyl,
tetrahydrofuryl, tetrahydropyrrolyl, pyrrolidinyl, octahydroindolyl,
octahydrobenzothiofuranyl, and octahydrobenzofuranyl.
Selective MEK 1 or MEK 2 inhibitors are those compounds which
inhibit the MEK 1 or MEK 2 enzymes, respectively, without substantially
inhibiting other enzymes such as MKK3, PKC, Cdk2A, phosphorylase kinase,
EGF, and PDGF receptor kinases, and C-src. In general, a selective MEK 1
or MEK 2 inhibitor has an ICSO for MEK 1 or MEK 2 that is at least one-
fiftieth
(1/50) that of its ICSO for one of the above-named other enzymes. Preferably,
a selective inhibitor has an ICSO that is at least 1/100, more preferably
1/500,
and even more preferably 1/1000, 1/5000, or less than that of its ICSO or one
or
more of the above-named enzymes.
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B. Compounds
One aspect of the invention features the use of compounds shown in
formula (I) in the Summary section. Embodiments of the invention includes
compounds of formula (I) wherein: (a) Q is formula (i); (b) R3 is H or fluoro;
(c)
R4 is fluoro, chloro, or bromo; (d) Rio is H, methyl, fluoro, or chloro; (e)
R~~ is
methyl, chloro, fluoro, nitro, or hydrogen; (f) R» is H; (g) R» is fluoro; (h)
each
of Rio and R» is fluoro; (i) R~ is H, methyl, ethyl, propyl, isopropyl,
isobutyl,
benzyl, phenethyl, allyl, C 3_5 alkenyl, C 3_6 cycloalkyl, (C 3_5 cycloalkyl)C
~_2
alkyl, (C 3_5 heterocyclic radical)C ~_2 alkyl, or (CHZ)2.~ NRcRp; (j) R~ is H
or (C
3~ cycloalkyl)C ~_2 alkyl; (k) R2 is H or methyl; (I) RA has at least one
hydroxyl
substituent; (m) RA is H, methyl, ethyl, isobutyl, hydroxyethyl, phenyl, 2-
piperidin-1-yl-ethyl, 2,3-dihydroxy-propyl, 3-[4-(2-hydroxyethyl)-piperazin-1-
yl]-
propyl, 2-pyrrolidin-1-yl-ethyl, or 2-diethylamino-ethyl; and RB is H; or
where
RB is methyl and RA is phenyl.; (n) W is NRARB or NR2NRARB; (o) W is
NR2(CH2)2.~ NRARB or O(CH2)2_3 NRARB; (p) W is NR20R~; (q) W is ORS; (r) Z
is formula (v); or (s) X~ is NRB, and R7 is H; or (t) combinations thereof. In
formula (I), the values for Z are shown left to right, or in a counter-
clockwise
orientation around the phenyl ring of Q.
According to one aspect of the invention, the compound of formula (I)
has a structure wherein: Q is formula (i) or (ii); R3 is H or fluoro; R4 is
fluoro,
chloro, or bromo; R,o is H, methyl, or chloro; R» is chloro, fluoro, or
hydrogen;
R, is H, methyl, ethyl, propyl, isopropyl, isobutyl, benzyl, phenethyl, allyl,
C 3-5
alkenyl, C 3_6 cycloalkyl, (C 3_5 cycloalkyl)C ,_2 alkyl, (C 3_5 heterocyclic
radical)C ~_2 alkyl, or (CH2)2~ NRcRp; R~ is H or (C 3~ cycloalkyl)C ~_2
alkyl; R2
is H or methyl; and Z is formula (v) or (vi). One embodiment of this aspect,
X~
is NRB. An example would be 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1[(2'-
morpholinyl)-ethyl]-2-(phenyl)-benzoimidazole-5-carboxylic acid
cyclopropylmethoxy-amide.
Embodiments of the invention also include compounds wherein Rio is
H; Rio is methyl or chloro; and where Rio is chloro. In some embodiments,
R7 and R8 together have no more than 14 carbon atoms, exclusive of R~, RM,
R~ and RK. Examples of this include compounds wherein R7 and R8 together
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have no more than 13 carbon atoms; no more than 7, 8, or 10 carbon atoms;
between 4 and 8 carbon atoms; between 1 and 10 carbon atoms; between 1
and 8 carbon atoms; and no more than 6 carbon atoms.
Preferably, where one of R~, R2, RA, RB, R~, Ro, RE, RF, R,, R~, RK, R~ ,
RM, R~, RH, RN, Ro, and RP is an alkenyl or alkynyl group, its double or
triple
bond, respectively, is not adjacent the point of attachment. For example,
where W is NR20R~, R2 is preferably prop-2-ynyl, or but-2 or 3-enyl, and less
preferably prop-1-ynyl or but-1-enyl.
Listed below are some of the preferred structures which can be
I 0 synthesized utilizing Schemes 1, 2, 10, and 11. Free acids, free
hydroxamic
acids, and cyclopropylmethyl hydroxamates are grouped together. For
example, compounds 1, 11, and 21 differ only by "W" (as defined in the
claims); compounds 2, 12, and 22 are similarly related. Preferred
compounds also include the 2-chloro (replacing 2-methyl) analogs of the listed
compounds.
Examples of compounds include: 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylic acid (APK ICSO = 47~17 nM);
7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzooxazole-5-carboxylic acid; 7-
Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzothiazole-5-carboxylic acid; 7-
Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzo(1,2,5]thiadiazole-5-carboxylic
acid; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzo[1,2,5]oxadiazole-5-
carboxylic acid; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-2-(2-hydroxyethyl)-
1 H-benzoimidazole-5-carboxylic acid; 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-2-(2-dimethylamino-ethyl)-1 H-benzoimidazole-5-carboxylic acid;
7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-1-acetyl-benzoimidazole-5-
carboxylic acid; 8-Fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-6-
carboxylic acid; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzotriazole-
5-carboxylic acid; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid hydroxyamide; 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-benzooxazole-5-carboxylic acid hydroxyamide; 7-Fluoro-6-(4-
iodo-2-methyl-phenylamino)-benzothiazole-5-carboxylic acid hydroxyamide; 7
Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzo[1,2,5]thiadiazole-5-carboxylic
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acid hydroxyamide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylic acid hydroxyamide; 7-Fluoro-6-(4-iodo-2-
methyl-phenylamino)-2-(2-hydroxyethyl)-1 H-benzoimidazole-5-carboxylic acid
hydroxyamide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-2-(2-dimethylamino-
ethyl)-1 H-benzoimidazole-5-carboxylic acid hydroxyamide; 7-Fluoro-6-(4-iodo-
2-methyl-phenylamino)-1-acetyl-benzoimidazole-5-carboxylic acid
hydroxyamide; 8-Fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-6-
carboxylic acid hydroxyamide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid hydroxyamide; 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylic acid cyclopropylmethoxy-
amide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-benzooxazole-5-carboxylic
acid cyclopropylmethoxy-amide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzothiazole-5-carboxylic acid cyclopropylmethoxy-amide; 7-Fluoro-6-(4-
iodo-2-methyl-phenylamino)-benzo[1,2,5]thiadiazole-5-carboxylic acid
cyclopropylmethoxy-amide; 7-Fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylic acid cyclopropylmethoxy-amide; 7-
Fluoro-6-(4-iodo-2-methyl-phenylamino)-2-(2-hydroxyethyl)-1 H-
benzoimidazole-5-carboxylic acid cyclopropylmethoxy-amide; 7-Fluoro-6-(4-
iodo-2-methyl-phenylamino)-2-(2-dimethylamino-ethyl)-1 H-benzoimidazole-5-
carboxylic acid cyclopropylmethoxy-amide; 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-1-acetyl-benzoimidazole-5-carboxylic acid cyclopropylmethoxy-
amide; 8-Fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-6-carboxylic
acid cyclopropylmethoxy-amide; and 7-Fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzotriazole-5-carboxylic acid cyclopropylmethoxy-amide.
The following is a list of examples representing schemes 3-9. As
above, free acids, free hydroxamic acids, and cyclopropylmethyl
hydroxamates are grouped together. For example, compounds 31, 45, and
59 differ only by "W' (as defined in the claims); compounds 32, 46, and 60 are
similarly related. Preferred compounds also include the 2-chloro (replacing 2-
methyl) analogs of the listed compounds.
Examples of compounds from schemes 3-9 include: 4-Fluoro-5-(4-
iodo-2-methyl-phenylamino)-benzothiazole-6-carboxylic acid; 4-Fluoro-5-(4-
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iodo-2-methyl-phenylamino)-benzooxazole-6-carboxylic acid; 5-(2-Chloro-4-
iodo-phenylamino)-6,7-difluoro-3H-benzoimidazole-4-carboxylic acid; 6,7-
Difluoro-2-(2-hydroxy-ethyl)-5-(4-iodo-2-methyl-phenylamino)-3H-
benzoimidazole-4-carboxylic acid; 6,7-Difluoro-5-(4-iodo-2-methyl-
phenylamino)-benzooxazole-4-carboxylic acid; 6,7-Difluoro-5-(4-iodo-2-
methyl-phenylamino)-benzothiazole-4-carboxylic acid; 7,8-Difluoro-6-(4-iodo-
2-methyl-phenylamino)-quinoxaline-5-carboxylic acid; 6-(4-lodo-2-methyl-
phenylamino)-8-nitro-quinoxaline-5-carboxylic acid; 5-(4-lodo-2-methyl-
phenylamino)-8-nitro-quinoxaline-6-carboxylic acid; 8-Chloro-5-(4-iodo-2-
methyl-phenylamino)-quinoxaline-6-carboxylic acid; 3-Cyclopropyl-7-(4-iodo-
2-methyl-phenylamino)-3H-benzoimidazole-4,6-dicarboxylic acid 4-
dimethylamide; 7-Bromo-4-(4-iodo-2-methyl-phenylamino)-benzooxazole-5-
carboxylic acid; 7-(2-Chloro-4-iodo-phenylamino)-4-fluoro-benzothiazole-6-
carboxylic acid; 7-(4-lodo-2-methyl-phenylamino)-4-nitro-benzooxazole-6-
carboxylic acid; 4-Fluoro-5-(4-iodo-2-methyl-phenylamino)-benzothiazole-6-
carboxylic acid hydroxyamide; 4-Fluoro-5-(4-iodo-2-methyl-phenylamino)-
benzooxazole-6-carboxylic acid hydroxyamide; 5-(2-Chloro-4-iodo-
phenylamino)-6,7-difluoro-3H-benzoimidazole-4-carboxylic acid
hydroxyamide; 6,7-Difluoro-2-(2-hydroxy-ethyl)-5-(4-iodo-2-methyl-
phenylamino)-3H-benzoimidazole-4-carboxylic acid hydroxyamide; 6,7-
Difluoro-5-(4-iodo-2-methyl-phenylamino)-benzooxazole-4-carboxylic acid
hydroxyamide; 6,7-Difluoro-5-(4-iodo-2-methyl-phenylamino)-benzothiazole-4-
carboxylic acid hydroxyamide; 7,8-Difluoro-6-(4-iodo-2-methyl-phenylamino)-
quinoxaline-5-carboxylic acid hydroxyamide; 6-(4-lodo-2-methyl-
phenylamino)-8-nitro-quinoxaline-5-carboxylic acid hydroxyamide; 5-(4-lodo-
2-methyl-phenylamino)-8-nitro-quinoxaline-6-carboxylic acid hydroxyamide; 8-
Chloro-5-(4-iodo-2-methyl-phenylamino)-quinoxaline-6-carboxylic acid
hydroxyamide; 3-Cyclopropyl-7-(4-iodo-2-methyl-phenylamino)-3H-
benzoimidazole-4,6-dicarboxylic acid 4-dimethylamide 6-hydroxyamide; 7-
Bromo-4-(4-iodo-2-methyl-phenylamino)-benzooxazole-5-carboxylic acid
hydroxyamide; 7-(2-Chloro-4-iodo-phenylamino)-4-fluoro-benzothiazole-6-
carboxylic acid hydroxyamide; 7-(4-lodo-2-methyl-phenylamino)-4-nitro-
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benzooxazole-6-carboxylic acid hydroxyamide; 4-Fluoro-5-(4-iodo-2-methyl-
phenylamino)-benzothiazole-6-carboxylic acid cyclopropylmethoxy-amide; 4-
Fluoro-5-(4-iodo-2-methyl-phenylamino)-benzooxazole-6-carboxylic acid
cyclopropylmethoxy-amide; 5-(2-Chloro-4-iodo-phenylamino)-6,7-difluoro-3H-
benzoimidazole-4-carboxylic acid cyclopropylmethoxy-amide; 6,7-Difluoro-2-
(2-hydroxy-ethyl)-5-(4-iodo-2-methyl-phenylamino)-3H-benzoimidazole-4-
carboxylic acid cyclopropylmethoxy-amide; 6,7-Difluoro-5-(4-iodo-2-methyl-
phenylamino)-benzooxazole-4-carboxylic acid cyclopropylmethoxy-amide;
6,7-Difluoro-5-(4-iodo-2-methyl-phenylamino)-benzothiazole-4-carboxylic acid
cyclopropylmethoxy-amide; 7,8-Difluoro-6-(4-iodo-2-methyl-phenylamino)-
quinoxaline-5-carboxylic acid cyclopropylmethoxy-amide; 6-(4-lodo-2-methyl-
phenylamino)-8-vitro-quinoxaline-5-carboxylic acid cyclopropylmethoxy-
amide; 5-(4-lodo-2-methyl-phenylamino)-8-vitro-quinoxaline-6-carboxylic acid
cyclopropylmethoxy-amide; 8-Chloro-5-(4-iodo-2-methyl-phenylamino)-
quinoxaline-6-carboxylic acid cyclopropylmethoxy-amide; 3-Cyclopropyl-7-
(4-iodo-2-methyl-phenylamino)-3H-benzoimidazole-4,6-dicarboxylic acid 4-
dimethylamide 6-cyclopropylmethoxy-amide; 7-Bromo-4-(4-iodo-2-methyl-
phenylamino)-benzooxazole-5-carboxylic acid cyclopropylmethoxy-amide;
7-(2-Chloro-4-iodo-phenylamino)-4-fluoro-benzothiazole-6-carboxylic acid
cyclopropylmethoxy-amide; and 7-(4-lodo-2-methyl-phenylamino)-4-nitro-
benzooxazole-6-carboxylic acid cyclopropylmethoxy-amide.
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C. Synthesis
The disclosed compounds can be synthesized according to the
following eleven Schemes, or variants thereof. These synthetic strategies are
further exemplified in Examples 1-22 below.
Scheme 1
HO O~ HO O
H-X1-R8 a 9 I ~ L~ H O R12
OZN ~ R3 OZN ~ R3 Catalyst '
Lz X i
H R8 a 9
,O O
Ri 2 O ~O Ri z H Rio
Rio Base w N w
HZN
+ I i I i
OZN R3 I i OzN R3 R~~
X~ RI1 Xt
H R8a 9 H R8a 9
,O O
R~ 2 O O H Rio O R~ 2 H Rio
Reducing Agent ~ N I ~ L ~ R7 I ~ N
i i
HzN X R3 R~ ~ N~X~ R3 Ra
1
H R8 a 9 R7 R8 a 9
R~ 2 O O H Rto
Iodinating Reagent I w N I w Deesterification
i i
N~X~ R3 Ri ~
R~ R8 a 9
R2
HO O Ri. .N O
H Rto O H Rlo
w N I w Rz-N-O-R~ ~ N
i i
N~X~ R3 R~~ I N~X~ R3 R»
R~r R8 a 9 R7 R8 a 9
18
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Scheme 2
,0 0
Rt2 O O Rto Rt2 H Rto
Base N
i Lt + H2N I i I i I i
O2N N \ R3 R t t ~N N H2R3 R t t
2
O ,O O
O R ~ R6 Rt2 H Rto
Reducing Agent Rt2 N Rto 5 O ~ N
w w I~ I
HN I ~ I N ~N R3 Rtt
2 R3 Rt t R5~
N H2
,O O
Rt2 H Rto
Iodinating Reagent w N w Deesterification
N ~N R3 Rtt I
R5~
R2
HO O Rt. .N O
H Rto H O H Rto
w N w R2-N-O-Ri ~ N
I ~ I ~ I ~ I
N
R3 Rt t I j1 N R3 Rt t
R ~ N RS
R6 R6
19
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Scheme 3
HO O HO O
Rt3 Lt
H-N-Rta I \ H-O-Rt2
OZN I ~ R3 Base OZN ~ R3 Catalyst
N.
~z Rt3 Rta
~O~O R ,O O
Rt2 to Rt2 H Rto
HZN ~ Base N
i
OzN I ~ R3 Rt t ~N I / R3 R
,N. ,N. tt
Rt3 Rta Rt3 Rta
,O O ~O O
Rt2 H Rto Rtz H Rto
Reducing Agent N I ) DiazotiTation ~ N
I I
I , I , 2) HXz HXZ i i
HZN ~ \R3 Rt t ~ N. R3 Rt t
~N_
Rt3 Rta Rt3 Rta
,O O
Rt2 H Rto O ,O O
Deprotection I ~ N I w Rt2 H Rto
i i ~~ R7 \ N
HXZ I / I /
NHZR3 Rtt X~N R3 Rtt
R/~
,O O
Rtz H Rto
Iodinating Reagent w N w Deesterification
I ~ I ~ t
X2 _N R3 Rtt
R~
R2
HO O Rt. .N O
H Rto O H Rto
N I ~ R2 N_O_Rt I ~ N
i i
I I
R~N R3 Rtt X~N R3 Rtt
R/~
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Scheme 4
HO O HO O HO O
Lt \ Lt NltCation Lt \ Lt H-X~-Rg or g Rg or 9 X~ \ Lt
R3 ~ O,N I ~ R; O,N I ~ R3
R4 R4 Ra
,O O
,O O R R
Rt'- Rto tz H to
H-O-Rt, R ~ X~ \ Lt HEN \ Base Rs or 9 X~ \ N \
8or9
Catalyst O.,N ~ R3 R I ~ O'-N / R R
it ~ 3 tt
R4
,O O OII
Reducing Agent Rt' H Rto ~~R~ Rsor Rt? O O H Rto
X N
R8 or 9 X~ I \ N I \ R~~ Z ~ \
i i N i i
H,N
Rs Rt t ~ R3 Rt t
HO O
~O O R
Rte H Rto R H to
Iodinating Reagent Rg °r 9 X, \ N \ Deesterification g or ~9 Xz \
N \
R~ N ~ ~ ~ r I R~~ ~ i
N I
Rs Rtt ~ R3 Rtt
Rz
Rt~ .N O
O Rto
H H
R,-N-O-R t Rg or 9' X, ~ N \
R7 \\
R3 Rt t
21
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Scheme 5
HO O HO O HO O
Li I ~ L~ Nitration L~ I w L' R,3_N_R~a Rya N ~ L~
i i O,N I ~ R3
R3 O,N ~ R3
Ra Ra Ra
,O O ,O O
R>> Rio Ria H Rio
H-O-R>> Ri3~N ~ Li H~_N Base Ri3~N ~ N
Ria I + I ~ Rya ~
Catalyst O,N ~ R i O,N ~
3 R~ i Rs R~ i
Ra Ra
O O O O
Rig H Rio Ri? H Rio
Reducing Agent R~ 3; N ~ N ~ 1 )Diazotization R~ 3~ N ~ N
R'H,N I ~ I ~ ~) W RHXi I ~ R I i
R3 Rig ~ 3 Rii
,O ,O ,O ~O
Ri2 H Rio O~~ Ri2 H Rto
Deprotection H,N I ~ N I ~ L~~ R7 yN I ~ N I w
R
i i X i i
i
HX ~ R3 Rig ~ ~ Rs Rig
R' O ~O R HO O Rio
i_ 11 io 1-1
Iodinating Reagent R7~N I ~ N I ~ Deesterification R7~N I ~ N I
r r
X i i Xi i i 1
~ R3 Ri ~ 1 ~ R3 Rt i
R,
Ri, .N O
H O H Rio
R,-N-O-Ri N w N w
R~-~rX~ I i I i I
R3 R~ i
22
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Scheme 6
HO O HO O HO O
Li Nitration Li ~ Li NH3 H,N ~ Li
R3 O,N I ~ R3 O,N I ~ R3
Ra Ra Ra
R'? O O Rio R~~ O O H Rio
H-O-Ri, H,N ~ Li H,N Base H,N ~ N
i ~ i
Catalyst O'N ~ R3 R, I i O,N ~ R3 R
ii
O
Reducing Agent R~' O O H R'o Rs~R6 RI' O O H RIo
H,N ~ N ~ JO( RS ~N ~ N
N
H,N ~ R3 R' ~ R~' ~- R3 Ri t
R'? O O H Rlo HO O H Rio
Iodinating Reagent Rs N \ N ~ Deesterification RS N ~ N
i I i
N R4 R3 Ri i 1 ~ N Ra \R3 Ra I
R,
R~~ .N~ O
1I O Rio
N N
R,-N-O-Ri Rs~ ~, w
\N I ~ I ~ 1
R6 R R3 R i i
a
23
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Scheme 7
HO O HO O HO O
Ra ~ Li Nitration Ra ~ Li NH3 Ra ~ Li
R3 I ~ R; I ~ NOz ~ R3 I / NOZ
NH,
,O O
R~Z O O Rio R~' H Rto
H-O-R ~, Ra ~ L i H,N ~ Base Ra I ~ N
Catalyst R3 I ~ NO, R I ~ R3 / NO, R /
ii tt
NH, NHZ
O ,O O
,O O R6 R~? H Rio
R~'- H Rio Rs~ Ra ~ N w
Reducing Agent R N O
a ~ w ~ w R3 ~ i N ~ i
i i ~ Rn
2 6
R3 NH NH~R~~ N\ R
Rs
O HO O R
Rig H Rio H io
Iodinating Reagent Ra ~ N ~ Deesterification Ra ~ N
R3 ~ i N ~ i I R3 ~ i N ~ i I
N~ R~i N~ Rii
Rb R6
Rs Rs
R,
R , .N O
H ~ O H Rio
Ra_N-O_Ri Ra w N w
R3 I ~ N I ~ I
N~ R~ i
R6
Rs
24
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Scheme 8
HO O HO O HO O
R.~ ~ Lt Nitration Ra ~ Lt H-X,~R8or9 R4 ~ Lt
R I i R3 I / NO~ R3 i N02
X2.
Lt Lt H R8or9
Rt' O p Rt? O O H Rto
H-O-Rt? R4 \ Lt to Base R4 w N w
I + HZN
I
Catalyst R3 ~ NO, I i R3 / NO, R /
)(, Rt t X, t t
H -. R8 or 9 H _ R8 or 9
,O O Q ,O O
Rt, H Rto ~ Rt? 1I Rto
Reducing Agent R4 ~ N ~ L, R~ Ra w N w
I~ I~ I~ I~
R3 XZ NH,Rtt R3 X~!~ Rtt
H Rg or g R8 or 9 R~
,O O HO O
Rt? H Rto H Rto
Iodinating Reagent R4 ~ N ~ Deesterification Ra w N w
I ~ I ~ I ~ I ~ I
R3 X,~ Rit I R3 X~!~ Rtt
i i
R8 or 9 R~ R8 or 9 R~
R,
Rt. .N O
H O Lt Rto
R,-N-O-Rt Ra ~ N W
I~
R3 XzJ Rtt I
\i
Rs or 9 R~
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Scheme 9
HO O HO O HO O
Ra \ Li Ra w Lt H Ra ( w Lt
I Nitration I R,3-N-R,a
i
R3 ~ R3 ~ NO, R3 NO,
N.
L~ Rts Rta
RI; O O R,' O O H Rto
H-O-Rte ~ I \ Li H'N Rto Base Ra I ~ N I
Catalyst R3 ~ NO~ I i R3
~N, Rn ~N_ NOZR~t
Rt3 Ria Rt3 Ria
Rt' O p H Rlo RIB O O H Rio
Reducing Agent Ra ~ N ~ 1)Diazotization Ra ~ N
I , I , 2) Hx,- I ~ I
R3 NH,Rtt R3 X~HRit
N. N.
Rt3 R,a Rts Rta
O ~ Rt? O O H Rto
Deprotection R~ \ N \ L" R~ Ra ~ N
i I i
I ~ I ~ R3 X~ R
R3 NH,X~HRt~ N~ tt
- R~
p HO~ O
Riz fi Rio H Rto
Iodinating Reagent Ra ~ N ~ Deesterification Ra ~ N
I
R3 N~ ~ Rt t 1 R3 N~ ~ Rt t 1
R~ R~
R,
R . .N O
H t O H Rto
RyN-O-R, Ra ~ N w
I
R3 N~ t Rtt
R~
26
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Scheme 10
,o o ,o 0
Rt 2 H Rtp Rt 2 H Rt0
N ~ Reagents , N
A I ~ I i Ni / I R
N ~R3 Rtt X3_N 3 Rtt
A=N02 or NH2
,O O
Rt 2 H Rt0
Iodinating Reagent i I N I w Deesterification
1
R3 R t t
X3 N
R2
HO O Rt. ,N O
Rtp O H R10
H
I N I w R2-N-O-Rt ~ I N
I N~ ~ I
X -N R3 Rtt X _N R3 Rit
3 3
27
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Scheme 11
,o o ,o 0
Rt2 H Rto Rtz H Rto
w N w RG_L3 i N w
I ~ I ~ RG. w I
HZN N \R3 Rtt H N. Rs Rtt
Rt3 'Rt4 Rt3 Rt4
,O O
Rtz H Rto
Deprotection ~ N ~ Diazotization
~'N ~ I I ~
E-I N H R3 R t t
2
,O O
Rt2 O o H Rto Rt2 H Rto
N ~ Iodinating Reagent ~ I N
I I ~ _ ~ i
R I
RG N=N R3 R t t G N=N R3 R t t
HO~O
Rto
Deesterification , I N I w R2-N-O-Rt
i
RG-N I
N=N Rs Rt t
R,
R , . N~ O
t o H Rto
N
~I I~
G' N
R N=N R3 Rt t I
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D. Uses
The disclosed compositions are useful as both prophylactic and
therapeutic treatments for diseases or conditions relating to chronic pain,
including neuropathic pain, as provided in the Summary section, as well as
diseases or conditions modulated by the MEK cascade. For example, in one
embodiment, the disclosed method relates to postoperative pain, phantom
limb pain, burn pain, gout, trigeminal neuralgia, acute herpetic and
postherpetic pain, causalgia, diabetic neuropathy, plexus avulsion, neuroma,
vasculitis, crush injury, constriction injury, tissue injury, post-surgical
pain,
arthritis pain, or limb amputation
For example, local injuries can be treated with local or topical
administration. Chronic pain affecting the entire body, such as diabetic
neuropathy can be treated with systemic administration (injection or orally)
of
a disclosed composition. Treatment for chronic pain (e.g., post-operative
pain) confined to the lower body can be administered centrally, e.g.,
epidurally. Formulations and methods of administration can include the use of
more than one MEK inhibitor, or a combination of a MEK inhibitor and another
pharmaceutical agent, such as an anti-inflammatory, analgesic, muscle
relaxing, or anti-infective agent. Preferred routes of administration are
oral,
intrathecal or epidural, subcutaneous, intravenous, intramuscular, and, for
non-human mammals, intraplantar, and are preferably epidural.
1. Dosages
Those skilled in the art will be able to determine, according to known
methods, the appropriate dosage for a patient, taking into account factors
such as age, weight, general health, the type of pain requiring treatment, and
the presence of other medications. In general, an effective amount will be
between 0.1 and 1000 mg/kg per day, preferably between 1 and 300 mg/kg
body weight, and daily dosages will be between 10 and 5000 mg for an adult
subject of normal weight. Commercially available capsules or other
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formulations (such as liquids and film-coated tablets) of 100 mg, 200 mg, 300
mg, or 400 mg can be administered according to the disclosed methods.
2. Formulations
Dosage unit forms include tablets, capsules, pills, powders, granules,
aqueous and nonaqueous oral solutions and suspensions, and parenteral
solutions packaged in containers adapted for subdivision into individual
doses.
Dosage unit forms can also be adapted for various methods of administration,
including controlled release formulations, such as subcutaneous implants.
Administration methods include oral, rectal, parenteral (intravenous,
intramuscular, subcutaneous), intracisternal, intravaginal, intraperitoneal,
intravesical, local (drops, powders, ointments, gels, or cream), and by
inhalation (a buccal or nasal spray).
Parenteral formulations include pharmaceutically acceptable aqueous
or nonaqueous solutions, dispersion, suspensions, emulsions, and sterile
powders for the preparation thereof. Examples of carriers include water,
ethanol, polyols (propylene glycol, polyethylene glycol), vegetable oils, and
injectable organic esters such as ethyl oleate. Fluidity can be maintained by
the use of a coating such as lecithin, a surfactant, or maintaining
appropriate
particle size. Carriers for solid dosage forms include (a) fillers or
extenders,
(b) binders, (c) humectants, (d) disintegrating agents, (e) solution
retarders, (f)
absorption acccelerators, (g) adsorbants, (h) lubricants, (i) buffering
agents,
and (j) propellants.
Compositions may also contain adjuvants such as preserving, wetting,
emulsifying, and dispensing agents; antimicrobial agents such as parabens,
chlorobutanol, phenol, and sorbic acid; isotonic agents such as a sugar or
sodium chloride; absorption-prolonging agents such as aluminum
monostearate and gelatin; and absorption-enhancing agents.
3. Related compounds
The invention provides the disclosed compounds and closely related,
pharmaceutically acceptable forms of the disclosed compounds, such as
salts, esters, amides, hydrates or solvated forms thereof; masked or protected
forms; and racemic mixtures, or enantiomerically or optically pure forms.
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Pharmaceutically acceptable salts, esters, and amides include
carboxylate salts (e.g., C ~_a alkyl, cycloalkyl, aryl, heteroaryl, or non-
aromatic
heterocyclic), amino acid addition salts, esters, and amides which are within
a
reasonable benefit/risk ratio, pharmacologically effective, and suitable for
contact with the tissues of patients without undue toxicity, irritation, or
allergic
response. Representative salts include hydrobromide, hydrochloride, sulfate,
bisulfate, nitrate, acetate, oxalate, valerate, oleate, palmitate, stearate,
laurate, borate, benzoate, lactate, phosphate, tosylate, citrate, maleate,
fumarate, succinate, tartrate, naphthylate, mesylate, glucoheptonate,
lactiobionate, and laurylsulfonate. These may include alkali metal and alkali
earth cations such as sodium, potassium, calcium, and magnesium, as well as
non-toxic ammonium, quaternary ammonium, and amine cations such as
tetramethyl ammonium, methylamine, trimethylamine, and ethylamine. See,
for example, S.M. Berge, et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977,
66:1-19 which is incorporated herein by reference. Representative
pharmaceutically acceptable amides of the invention include those derived
from ammonia, primary C ~_s alkyl amines and secondary di (C ,_6 alkyl)
amines. Secondary amines include 5- or 6-membered heterocyclic or
heteroaromatic ring moieties containing at least one nitrogen atom and
optionally between 1 and 2 additional heteroatoms. Preferred amides are
derived from ammonia, C ~_3 alkyl primary amines, and di (C ~_2 alkyl)amines.
Representative pharmaceutically acceptable esters of the invention include
C ~_~ alkyl, C 5_~ cycloalkyl, phenyl, and phenyl(C ,_6)alkyl esters.
Preferred
esters include methyl esters.
The invention also includes disclosed compounds having one or more
functional groups (e.g., hydroxyl, amino, or carboxyl) masked by a protecting
group. Some of these masked or protected compounds are pharmaceutically
acceptable; others will be useful as intermediates. Synthetic intermediates
and processes disclosed herein, and minor modifications thereof, are also
within the scope of the invention.
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HYDROXYL PROTECTING GROUPS
Hydroxyl protecting groups include: ethers, esters, and protection for 1,2-
and
1,3-diols. The ether protecting groups include: methyl, substituted methyl
ethers, substituted ethyl ethers, substituted benzyl ethers, silyl ethers and
conversion of silyl ethers to other functional groups.
Substituted Methyl Ethers
Substituted methyl ethers include: methoxymethyl, methylthiomethyl, t-
utylthiomethyl, (phenyldimethylsilyl) methoxymethyl, benzyloxymethyl, p-
ethoxybenzyloxymethyl, (4-methoxyphenoxy) methyl, guaiacolmethyl, t-
butoxymethyl, 4-pentenyloxymethyl, siloxymethyl, 2-methoxyethoxymethyl,
2,2,2-trichloroethoxymethyl, bis(2-chloro- ethoxy)methyl, 2-
(trimethylsilyl)ethoxymethyl, tetrahydropyranyl, 3-bromotetrahydro-pyranyl,
tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl, 4-
methoxytetrahydrothio-pyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxido,
1-[(2-chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl, 1,4-dioxan-2-yl,
tetrahydrofuranyl, tetrahydrothiofuranyl, and 2,3,3a,4,5,6,7,7a-octahydro-
7,8,8-trimethyl-4,7-ethanobenzofuran-2-yl.
Substituted Ethyl Ethers
Substituted ethyl ethers include: 1-ethoxyethyl, 1-(2,chloroethoxy)ethyl,
1-methyl-1-methoxyethyl, 1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-
fluoroethyl, 2,2,2-trichloroethyl, 2-trimethylsilyethyl, 2-
(phenylselenyl)ethyl, t
butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl, and benzyl.
Substituted Benzyl Ethers
Substituted benzyl ethers include: p-methoxybenzyl, 3,4-dimethoxybenzyl,
o-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl,
p-phenylbenzyl, 2- and 4-picolyl, 3-methyl-2-picolyl N-oxido, diphenylmethyl,
p, p'-dinitrobenzhydryl, 5-dibenzosuberyl, triphenylmethyl, a-naphthyldiphenyl-
methyl, p-methoxyphenyldiphenylmethyl, di(p-methoxyphenyl)phenylmethyl,
tri-(p-methoxyphenyl)methyl, 4-(4'-bromophenacyloxy)phenyldiphenylmethyl,
4,4',4"-tris(4,5-dichlorophthalimidophenyl)methyl, 4,4',4"-
tris(levulinoyloxyphenyl) methyl, 4,4',4"tris(benzoyloxyphenyl)methyl, 3-
(imidazol-1-ylmethyl)bis(4',4"-dimethoxyphenyl)-methyl, 1,1-bis(4-
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methoxyphenyl)-1'-pyrenylmethyl, 9-anthryl, 9-(9-phenyl) xanthenyl, 9-(9-
phenyl-10-oxo) anthryl, 1,3-benzodithiolan-2-yl, and benzisothiazolyl S,S-
dioxido.
Silyl Ethers
Silyl ethers include: trimethylsilyl, triethylsilyl, triisopropylsilyl,
dimethylisopropylsilyl, diethylisopropylsilyl, dimethylthexylsilyl, t-
butyldimethylsilyl, t butyldiphenylsilyl, tribenzylsilyl, tri-p-xylylsilyl,
triphenylsilyl,
diphenylmethylsilyl, and t-butylmethoxyphenylsilyl.
ESTERS
Esters protecting groups include: esters, carbonates, assisted cleavage,
miscellaneous esters, and sulfonates.
Esters
Examples of protective esters include: formate, benzoylformate, acetate,
chloroacetate, dichloroacetate, trichloroacetate, trifluoroacetate,
methoxyacetate, triphenylmethoxyacetate, phenoxyacetate, p-
chlorophenoxyacetate, p-P-phenylacetate, 3-phenylpropionate, 4-
oxopentanoate (levulinate), 4,4-(ethylenedithio) pentanoate, pivaloate,
adamantoate,crotonate,4-methoxycrotonate, benzoate, p-phenylbenzoate,
and 2,4,6-trimethylbenzoate (mesitoate).
Carbonates
Carbonates include: methyl, 9-fluorenylmethyl, ethyl, 2,2,2-trichloroethyl,
2-(trimethylsilyl) ethyl, 2-(phenylsulfonyl) ethyl, 2-(triphenylphosphonio)
ethyl,
isobutyl, vinyl, allyl, p-nitrophenyl, benzyl, p-methoxybenzyl, 3,4-
dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, S-benzyl thiocarbonate, 4-
ethoxy-1-naphthyl, and methyl dithiocarbonate.
Assisted Cleavage
Examples of assisted cleavage protecting groups include: 2-iodobenzoate, 4-
azido-butyrate, 4-nitro-4-methylpentanoate, o-(dibromomethyl) benzoate, 2-
formylbenzene-sulfonate, 2-(methylthiomethoxy) ethyl carbonate, 4-
(methylthiomethoxymethyl) benzoate, and 2-(methylthiomethoxymethyl)
benzoate.
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Miscellaneous Esters
In addition to the above classes, miscellaneous esters include: 2,6-dichloro-4-
methylphenoxyacetate, 2,6-dichloro-4-(1,1,3,3-tetramethylbutyl)
phenoxyacetate, 2,4-bis(1,1-dimethylpropyl) phenoxyacetate,
chlorodiphenylacetate, isobutyrate, monosuccinoate, (E]-2-methyl-2-
butenoate (tigloate), o-(methoxycarbonyl) benzoate, p-P-benzoate,
a-naphthoate, nitrate, alkyl N,N,N' N'-tetramethylphosphorodiamidate,
N-phenylcarbamate, borate, dimethylphosphinothioyl, and
2,4-dinitrophenylsulfenate.
Sulfonates
Protective sulfates includes: sulfate, methanesulfonate(mesylate),
benzylsulfonate, and tosylate.
PROTECTION FOR 1.2- AND 1.3-DIOLS
The protection for 1,2 and 1,3-diols group includes: cyclic acetals and
ketals,
cyclic ortho esters, and silyl derivatives.
Cyclic Acetals and Ketals
Cyclic acetals and ketals include: methylene, ethylidene, 1-t-butylethylidene,
1-phenylethylidene, (4-methoxyphenyl) ethylidene, 2,2,2-trichloroethylidene,
acetonide (isopropylidene), cyclopentylidene, cyclohexylidene,
cycloheptylidene, benzylidene, p-methoxybenzylidene, 2,4-
dimethoxybenzylidene, 3,4-dimethoxybenzylidene, and 2-nitrobenzylidene.
Cyclic Ortho Esters
Cyclic ortho esters include: methoxymethylene, ethoxymethylene, dimethoxy-
methylene, 1-methoxyethylidene, 1-ethoxyethylidine, 1,2-
dimethoxyethylidene, a-methoxybenzylidene, 1-(N,N-
dimethylamino)ethylidene derivative, a-(N,N-dimethylamino) benzylidene
derivative, and 2-oxacyclopentylidene.
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PROTECTION FOR THE CARBOXYL GROUP
ESTERS
Ester protecting groups include: esters, substituted methyl esters, 2-
substituted ethyl esters, substituted benzyl esters, silyl esters, activated
esters, miscellaneous derivatives, and stannyl esters.
Substituted Methyl Esters
Substituted methyl esters include: 9-fluorenylmethyl, methoxymethyl,
methylthiomethyl, tetrahydropyranyl, tetrahydrofuranyl, methoxyethoxymethyl,
2-(trimethylsilyl)ethoxy-methyl, benzyloxymethyl, phenacyl, p-bromophenacyl,
a-methylphenacyl, p-methoxyphenacyl, carboxamidomethyl, and N-
phthalimidomethyl.
2-Substituted Ethyl Esters
2-Substituted ethyl esters include: 2,2,2-trichloroethyl, 2-haloethyl, a-
chloroalkyl, 2-(trimethylsily)ethyl, 2-methylthioethyl, 1,3-dithianyl-2-
methyl,
2(p-nitrophenylsulfenyl)-ethyl, 2-(p-toluenesulfonyl)ethyl, 2-(2'-
pyridyl)ethyl, 2-
(diphenylphosphino)ethyl, 1-methyl-1-phenylethyl, t-butyl, cyclopentyl,
cyclohexyl, allyl, 3-buten-1-yl, 4-(trimethylsily)-2-buten-1-yl, cinnamyl, a-
methylcinnamyl, phenyl, p-(methylmercapto)-phenyl, and benzyl.
Substituted Benzyl Esters
Substituted benzyl esters include: triphenylmethyl, diphenylmethyl,
bis(o-nitrophenyl)methyl, 9-anthrylmethyl, 2-(9,10-dioxo)anthrylmethyl, 5-
dibenzo-suberyl, 1-pyrenylmethyl,2-(trifluoromethyl)-6-chromylmethyl, 2,4,6-
trimethylbenzyl, p-bromobenzyl, o-nitrobenzyl, p-nitrobenzyl, p-
methoxybenzyl, 2,6-dimethoxybenzyl, 4-(methylsulfinyl)benzyl, 4-sulfobenzyl,
piperonyl, and 4-P-benzyl.
Silyl Esters
Silyl esters include: trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, i-
propyldimethylsilyl, phenyldimethylsilyl, and di- t-butylmethylsilyl.
Miscellaneous Derivatives
Miscellaneous derivatives includes: oxazoles, 2-alkyl-1,3-oxazolines, 4-alkyl-
5-oxo-1,3-oxazolidines, 5-alkyl-4-oxo-1,3-dioxolanes, ortho esters, phenyl
group, and pentaaminocobalt(III) complex.
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Stannyl Esters
Examples of stannyl esters include: triethylstannyl and tri-n-butylstannyl.
AMIDES AND HYDRAZIDES
Amides include: N,N-dimethyl, pyrrolidinyl, piperidinyl, 5,6-
dihydrophenanthridinyl, o-nitroanilides, N-7-nitroindolyl, N-8-nitro-1,2,3,4-
tetrahydroquinolyl, and p-P-benzenesulfonamides. Hydrazides include: N-
phenyl, N,N'-diisopropyl and other dialkyl hydrazides.
PROTECTION FOR THE AMINO GROUP
CARBAMATES
Carbamates include: carbamates, substituted ethyl, assisted cleavage,
photolytic cleavage, urea-type derivatives, and miscellaneous carbamates.
Carbamates
Carbamates include: methyl and ethyl, 9-fluorenylmethyl, 9-(2-
sulfo)fluorenylmethyl, 9-(2,7-dibromo)fluorenylmethyl, 2,7-di-t-butyl-[9-
(10,10-
dioxo-10,10,10,10-tetrahydro- thioxanthyl)]methyl, and 4-methoxyphenacyl.
Substituted Ethyl
Substituted ethyl protective groups include: 2,2,2-trichloroethyl, 2-
trimethylsilylethyl, 2-phenylethyl, 1-(1-adamantyl)-1-methylethyl, 1,1-
dimethyl-
2-haloethyl, 1,1dimethyl-2,2-dibromoethyl, 1,1-dimethyl-2,2,2-trichloroethyl,
1-
methyl-1-(4-biphenylyl)ethyl, 1-(3,5-di-t-butylphenyl)-1-methylethyl, 2-(2'-
and
4'-pyridyl)ethyl, 2-(N,N-icyclohexylcarboxamido)- ethyl, t-butyl, 1-adamantyl,
vinyl, allyl, 1-isopropylallyl, connamyl, 4-nitrocinnamyl, quinolyl, N-
hydroxypiperidinyl, alkyldithio, benzyl, p-methoxybenzyl, p-nitrobenzyl, p-
bromobenzyl, p-chlorobenzyl, 2,4dichlorobenzyl, 4-methylsulfinylbenzyl,
9-anthrylmethyl, and diphenylmethyl.
Assisted Cleavage
Protection via assisted cleavage includes: 2-methylthioethyl,
2-methylsulfonylethyl, 2-(p-toluenesulfonyl)ethyl, [2-(1,3-dithianyl)]methyl,
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4-methylthiophenyl, 2,4-dimethyl-thiophenyl, 2-phosphonioethyl,
2-triphenylphosphonioisopropyl, 1,1-dimethyl-2cyanoethyl, m-chloro-p-
acyloxybenzyl, p-(dihydroxyboryl)benzyl, 5-benzisoxazolyl-methyl, and
2-(trifluoromethyl)-6-chromonylmethyl.
Photolytic Cleavage
Photolytic cleavage methods use groups such as: m-nitrophenyl, 3,5-
dimethoxybenzyl, o-nitrobenzyl, 3,4-dimethoxy-6-nitrobenzyl, and phenyl(o-
nitrophenyl)methyl.
Urea-Type Derivatives
Examples of of urea-type derivatives include: phenothiazinyl-(10)-carbonyl
derivative, N'-p-toluenesulfonylaminocarbonyl, and N'-
phenylaminothiocarbonyl.
Miscellaneous Carbamates
In addition to the above, miscellaneous carbamates include: t amyl, S-benzyl
thiocarbamate, p-cyanobenzyl, cyclobutyl, cyclohexyl, cyclopentyl,
cyclopropylmethyl, p-decyloxy-benzyl, diisopropylmethyl, 2,2-
dimethoxycarbonylvinyl, o-(N,N-dimethyl-carboxamido)-benzyl, 1,1-dimethyl-
3(N,N-dimethylcarboxamido)propyl, 1,1-dimethyl-propynyl, di(2-pyridyl)methyl,
2-furanylmethyl, 2-iodoethyl, isobornyl, isobutyl, isonicotinyl, p(p'-
methoxyphenyl- azo)benzyl, 1-methylcyclobutyl, 1-methylcyclohexyl, 1-
methyl-1-cyclopropyl- methyl, 1-methyl-(3,5-dimethoxyphenyl)ethyl, 1-methyl-
1(p-henylazophenyl)- ethyl, 1-methyl-1-phenylethyl, 1-methyl-1-(4-
pyridyl)ethyl, phenyl, p-(phenylazo)benzyl, 2,4,6-tri-t butylphenyl, 4-
(trimethylammonium) benzyl, and 2,4,6-trimethylbenzyl.
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AMIDES
Amides
Amides includes: N-formyl, N-acetyl, N-chloroacetyl, N-trichloroacetyl,
N-trifluoroacetyl, N-phenylacetyl, N-3-phenylpropionyl, N-picolinoyl, N-3-
pyridyl-carboxamide, N-benzoylphenylalanyl derivative, N-benzoyl, and N-p-
phenylbenzoyl.
Assisted Cleavage
Assisted cleavage groups include: N-o-nitrophenylacetyl, N-o-
nitrophenoxyacetyl, N-acetoacetyl, (N'-dithiobenzyloxycarbonylamino)acetyl,
N-3-(p-hydroxphenyl) propionyl, N-3-(o-nitrophenyl)propionyl, N-2-methyl-2-
(o-nitrophenoxy)propionyl, N-2-methyl-2-(o-phenylazophenoxy)propionyl, N-4-
chlorobutyryl, N-3-methyl-3-nitrobutyryl, N-o-nitrocinnamoyl, N-
acetylmethionine derivative, N-o-nitrobenzoyl, N-o-
(benzoyloxymethyl)benzoyl, and 4,5-diphenyl-3-oxazolin-2-one.
C~iclic Imide Derivatives
Cyclic imide derivatives include: N-phthalimide, N-dithiasuccinoyl,
N-2,3-Biphenyl-maleoyl, N-2,5-dimethylpyrrolyl,
N-1,1,4,4-tetramethyldisilylazacyclopentane adduct, 5-substituted
1,3-dimethyl-1,3,5-triazacyclohexan-2-one, 5-substituted 1,3-dibenzyl-
1,3,5-triazacyclohexan-2-one, and 1-substituted 3,5-dinitro-4-pyridonyl.
SPECIAL -NH PROTECTIVE GROUPS
Protective groups for - NH include: N-alkyl and N-aryl amines, imine
derivatives, enamine derivatives, and N-hetero atom derivatives (such as N-
metal, N-N, N-P, N-Si, and N-S), N-sulfenyl, and N-sulfonyl.
N-Alkyl and N-Aryl Amines
N-alkyl and N-aryl amines include: N-methyl, N-allyl,
N-[2-(trimethylsilyl)ethoxyl]-methyl, N-3-acetoxypropyl,
N-(1-isopropyl-4-nitro-2-oxo-3-pyrrolin-3-yl), quaternary ammonium salts,
N-benzyl, N-di(4-methoxyphenyl)methyl, N-5-dibenzosuberyl,
N-triphenylmethyl, N-(4-methoxyphenyl)diphenylmethyl, N-9-phenylfluorenyl,
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N-2,7-dichloro-9-fluorenylmethylene, N-ferrocenylmethyl, and
N-2-picolylamine N'-oxide.
(mine Derivatives
Imine derivatives include: N-1,1-dimethylthiomethylene, N-benzylidene,
N-p-methoxybenzylidene, N-diphenylmethylene,
N-[(2-pyridyl)mesityl]methylene,
N-(N',N'-dimethylaminomethylene), N,N'-isopropylidene,
N-p-nitrobenzylidene,
N-salicylidene, N-5-chlorosalicylidene, N-(5-chloro-2-hydroxyphenyl)phenyl-
methylene, and N-cyclohexylidene.
Enamine Derivative
An example of an enamine derivative is N-
(5,5-dimethyl-3-oxo-1-cyclohexenyl).
N-Hetero Atom Derivatives
N-metal derivatives include: N-borane derivatives, N-diphenylborinic acid
derivative, N-[phenyl(pentacarbonylchromium- or -tungsten)]carbenyl, and
N-copper or N-zinc chelate. Examples of N-N derivatives include: N-vitro,
N-nitroso, and N-oxide. Examples of N-P derivatives include:
N-diphenylphosphinyl, N-dimethylthiophosphinyl, N-diphenylthiophosphinyl,
N-dialkyl phosphoryl, N-dibenzyl phosphoryl, and N-Biphenyl phosphoryl.
Examples of N-sulfenyl derivatives include: N-benzenesulfenyl,
N-o-nitrobenzenesulfenyl, N-2,4-dinitrobenzenesulfenyl,
N-pentachlorobenzenesulfenyl, N-2-vitro-4-methoxy-benzenesulfenyl,
N-triphenylmethylsulfenyl, and N-3-nitropyridinesulfenyl. N-sulfonyl
derivatives include: N-p-toluenesulfonyl, N-benzenesulfonyl,
N-2,3,6-trimethyl- 4-methoxybenzenesulfonyl,
N-2,4,6-trimethoxybenzenesulfonyl, N-2,6-dimethyl-4-methoxy-
benzenesulfonyl, N-pentamethylbenzenesulfonyl,
N-2,3,5,6-tetramethyl-4-methoxybenzene- sulfonyl,
N-4-methoxybenzenesulfonyl, N-2,4,6-trimethylbenzenesulfonyl, N-
2,6-dimethoxy- 4-methylbenzenesulfonyl,
N-2,2,5,7,8-pentamethylchroman-6-sulfonyl, N-methanesulfonyl,
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N-~i-trimethylsilylethanesulfonyl, N-9-anthracenesulfonyl,
N-4-(4',8'-dimethoxynaphthylmethyl)-benzenesulfonyl, N-benzylsulfonyl,
N-trifluoromethylsulfonyl, and N-phenacylsulfonyl.
Disclosed compounds which are masked or protected may be prodrugs,
compounds metabolized or otherwise transformed in vivo to yield a disclosed
compound, e.g., transiently during metabolism. This transformation may be a
hydrolysis or oxidation which results from contact with a bodily fluid such as
blood, or
the action of acids, or liver, gastrointestinal, or other enzymes.
Features of the invention are further described in the examples below.
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E. Examples
BIOLOGICAL EXAMPLES
Example 1
Effect of PD 198306 on streptozocin-induced static allodynia
Animals
Male Sprague Dawley rats (250-300g), obtained from Bantin and
Kingman, (Hull, U.K.) were housed in groups of 3. All animals were kept under
a 12h light/dark cycle (lights on at 07h OOmin) with food and water ad
libitum.
All experiments were carried out by an observer blind to drug treatments.
Development of diabetes in the rat
Diabetes was induced in rats by a single i.p. injection of streptozocin
(50 mg/kg) as described previously (Courteix et al., 1993).
Evaluation of static allodynia
Mechanical hypersensitivity was measured using Semmes-Weinstein
von Frey hairs (Stoelting, Illinois, U.S.A.). Animals were placed into wire
mesh
bottom cages allowing access to the underside of their paws. Animals were
habituated to this environment prior to the start of the experiment.
Mechanical
hypersensitivity was tested by touching the plantar surface of the animals
right
hind paw with von Frey hairs in ascending order of force ( 0.7, 1.2, 1.5, 2,
3.6,
5.5, 8.5, 11.8, 15.1 and 29g) for up to 6 sec. Once a withdrawal response was
established, the paw was re-tested, starting with the next descending von
Frey hair until no response occurred. The highest force of 29 g lifted the paw
as well as eliciting a response, thus represented the cut off point. The
lowest
amount of force required to elicit a response was recorded as the paw
withdrawal threshold (PWT) in grams.
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Dru s
PD 198306 [N-Cyclopropylmethoxy-3,4,5-trifluoro-2-(4-iodo-2-methyl-
phenylamino)-benzamide] and CI-1008 (pregabalin) were synthesized at
Parke-Davis (Ann Arbor, MI, USA). PD 198306 was suspended in
cremophor:ethanol:water (1:1:8) vehicle. Pregabalin was dissolved in water.
Both compounds were administered orally. Streptozocin (Aldrich, UK) was
dissolved in 0.9% w/v NaCI and administered intraperitoneally. Drug
administrations were made in a volume of 1 ml/kg.
Statistics
The static allodynia data were analysed using a Kruskall-Wallis ANOVA
for non-parametric results, followed when significant by Mann-Whitney's t
test.
Experimental protocol
Static allodynia was assessed with von Frey hairs, before (baseline,
BL) and 1 h after oral administration of PD 198306 (30mg/kg, p.o.), vehicle
(cremophor:ethanol:water, 1:1:8) or pregabalin (30mg/kg, p.o.) (test).
Animals were administered again the same compounds on the following day,
both in the morning and the afternoon. Static allodynia was assessed only
before and 1 h after the afternoon administration, in order to minimise the
habituation of the animals to the testing conditions. Animals treated with
pregabalin received water in the morning administration, in order to avoid the
potential development of tolerance to the compound with repeated
administration.
Day 1: Day 2:2:
a.m.: PD 198306
Water
Vehicle
p.m.: BL p.m.: BL
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PD 198306 PD 198306
Pregabalin Pregabalin
Vehicle Vehicle
Test Test
RESULTS
A single administration of pregabalin (30mg/kg, p.o.) significantly
blocked streptozocin-induced static allodynia 1 h after administration. In
contrast, a single administration of PD 198306 (30mg/kg, p.o) had no effect on
streptozocin-induced static allodynia 1 h after administration (see below).
However, after the compound had been administered twice more on the
following day, it significantly blocked streptozocin-induced static allodynia
1 h
after the third administration. The efFects had disappeared by the following
day (see FIG. 1 ).
Example 2
MATERIALS AND METHODS
Animals
Male Sprague Dawley rats (250-300g), obtained from Charles River,
Margate, U.K.) were housed in groups of 3-6. All animals were kept under a
12h light/dark cycle (lights on at 07h OOmin) with food and water ad libitum.
All
experiments were carried out by an observer blind to drug treatments.
Diabetes was induced in rats by a single i.p. injection of streptozocin
(50mg/kg) as described previously (Courteix et al., 1993).
Development of Chronic Constriction Iniury in the rat
Animals were anaesthetised with 2% isoflurane 1:4 02/N20 mixture
maintained during surgery via a nose cone. The sciatic nerve was ligated as
previously described by Bennett and Xie, 1988. Animals were placed on a
homeothermic blanket for the duration of the procedure. After surgical
preparation the common sciatic nerve was exposed at the middle of the thigh
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by blunt dissection through biceps femoris. Proximal to the sciatic
trifurcation,
about 7mm of nerve was freed of adhering tissue and 4 ligatures (4-0 silk)
were tied loosely around it with about 1 mm spacing. The incision was closed
in layers and the wound treated with topical antibiotics.
Intrathecal infections
PD 198306 and pregabalin were administered intrathecally in a volume
of 10 ~.I using a 100 ~.I Hamilton syringe by exposing the spine of the rats
under
brief isoflurane anaesthesia. Injections were made into the intrathecal space
between lumbar region 5-6 with a 10 mm long 27 gauge needle. Penetrations
were judged successful if there was a tail flick response. The wound was
sealed
with an autoclip and rats appeared fully awake within 2-3 min following
injection.
Evaluation of static allodynia
Mechanical hypersensitivity was measured using Semmes-Weinstein
von Frey hairs (Stoelting, Illinois, U.S.A.). Animals were placed into wire
mesh
bottom cages allowing access to the underside of their paws. Animals were
habituated to this environment prior to the start of the experiment.
Mechanical
hypersensitivity was tested by touching the plantar surface of the animals
right
hind paw with von Frey hairs in ascending order of force ( 0.7, 1.2, 1.5, 2,
3.6,
5.5, 8.5, 11.8, 15.1 and 29g) for up to 6sec. Once a withdrawal response was
established, the paw was re-tested, starting with the next descending von
Frey hair until no response occurred. The highest force of 29g lifted the paw
as well as eliciting a response, thus represented the cut off point. The
lowest
amount of force required to elicit a response was recorded as the paw
withdrawal threshold (PWT) in grams.
Experimental protocol
Static allodynia was assessed with von Frey hairs, before (baseline,
BL) and 0.5h, 1 h and 2h after intrathecal or intraplantar administration of
PD
198306 (1-30~g, i.t.), vehicle (cremophor:ethanol:water, 1:1:8) or pregabalin
(10~.g, i.t). For oral administration experiments, static allodynia was
assessed
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with von Frey hairs, before (baseline, BL) and 1 h after oral administration
of
PD 198306 (3-30mg/kg, p.o.), vehicle (cremophor:ethanol:water, 1:1:8) or
pregabalin (30mg/kg, p.o.). Animals were administered again the same
compounds on the following day, both in the morning and the afternoon. Static
allodynia was assessed before and 1 h after the morning administration. In the
afternoon static allodynia was assessed before, 1 h, 2h and 3h after
administration for streptozocin treated animals. CCI animals were assessed
before, 1 h and 2h after administration
Drugs used
PD 198306 and pregabalin were synthesised at Parke-Davis (Ann
Arbor, MI, USA). PD 198306 was suspended in cremophor:ethanol:water
(1:1:8) vehicle. Pregabalin was dissolved in water. Both compounds were
administered orally, intrathecally or intraplantar in volumes of 1 ml/kg, 101
and
I S 1001 respectively. Streptozocin (Aldrich, UK) was dissolved in 0.9% w/v
NaCI
and administered intraperitoneally in a volume of 1 ml/kg.
Statistics
Data were analysed using a Kruskall-Wallis ANOVA for non-parametric
results, followed when significant by Mann-Whitney's t test vs vehicle group.
RESULTS
1. Effects of PD 198306 on static allodynia, following systemic
administration
1 1 Effect of PD198306 on streptozocin-induced static allodynia
A single administration of pregabalin (30mg/kg, p.o.) significantly blocked
streptozocin-induced static allodynia 1 h after administration. In contrast, a
single administration of PD 198306 (3-30mg/kg, p.o) had no effect on
streptozocin-induced static allodynia 1 h after administration (FIG. 2).
However, after the compound had been administered twice more on the
following day, PD 198306 (30mg/kg) significantly blocked streptozocin-
induced static allodynia for 2h after the third administration (FIG. 2).
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1.2. Effect of PD198306 on CCI-induced static allodynia
A single administration of pregabalin (30mg/kg, p.o.) significantly blocked
CCI-
induced static allodynia 1 h after administration. In contrast, neither a
single or
multiple administration of PD 198306 (3-30mg/kg, p.o) had any effect on CCI-
induced static allodynia (FIG. 3).
2. Effects of PD 198306 on static allodynia, following intrathecal
administration
Intrathecally administered PD198306 (1-30~g) dose-dependently blocked the
maintenance of static allodynia in both streptozocin (FIG. 4) and CCI animals
(FIG. 5) with respective MEDs of 3 and 10 fig. This antiallodynic effect
lasted
for 1 h.
3. Effects of PD 198306 on static allodynia, following intraplantar
administration
An intrathecal administration of PD 198306 (30~g) significantly blocked static
allodynia in both neuropathic pain models (FIGS. 6,7). In contrast, a single
administration of PD 198306 at a dose 100-fold higher (3mg/100~.1) directly
into the paw had no effect on streptozocin (FIG. 6) or CCI-induced static
allodynia (FIG. 7).
REFERENCES
Bennett GJ, Xie Y-K. A peripheral mononeuropathy in rat that produces
disorders of pain sensation like those seen in man. Pain 1988;33:87-107.
Courteix C, Eschalier A and Lavarenne J. Streptozocin -induced rats:
behavioural evidence for a model of chronic pain. Pain 1993;53:81-8
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Example 3
Effect of other MEK inhibitors in a neuropathic pain model in the rat
SUMMARY
S The effect of several MEK inhibitors, with different binding affinities, has
been investigated in the CCI model of neuropathic pain in the rat, by
assessing
static allodynia with von Frey hairs. Intrathecal administration of PD219622
or
PD297447 (30~g) had no significant effect on allodynia. This lack of effect
may reflect the low affinity or solubility of the compounds. However,
intrathecal administration of PD 254552 or PD 184352 (30~g), which posses
higher binding affinities, blocked the maintenance of static allodynia in CCI
animals. The antiallodynic effect was only evident for 30min post-injection
and
thus, shorter than the one observed for pregabalin (100~.g). The magnitude of
the effect was similar for 30~g of PD 184352 and 100~.g of pregabalin. From
this study it is concluded that MEK inhibitors exert an antiallodynic effect
in
CCI-induced neuropathic rats when administered intrathecally, and that the
antiallodynic effect correlates with the affinity of the compounds.
The animals and methods for developing chronic constriction injury in
the rat, injecting test compounds, and evaluation of static allodynia were
according to Example 2 above. PD219622, PD297447, PD 184352, PD
254552 and pregabalin were administered intrathecally at doses of 30~g for all
PD compounds and 100~g for pregabalin. Static allodynia was assessed with
von Frey hairs, before (baseline, BL) and 0.5h, 1 h and 2h after intrathecal
administration of the compounds
Drugs used
PD297447, PD219622, PD 254552, PD 184352 (CI-1040), and pregabalin
were synthesised at Parke-Davis (Ann Arbor, MI, USA). PD297447,
PD219622, PD 254552 and PD 184352 were suspended in
cremophor:ethanol:water (1:1:8) vehicle. Pregabalin was dissolved in water.
All
compounds were administered intrathecally in a 101 volume.
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Statistics
Data were analysed using a Kruskall-Wallis ANOVA for non-parametric
results, followed when significant by Mann-Whitney's t test vs vehicle group.
RESULTS
Intrathecally administered PD297447 or PD219622 (30~g) had no
significant effect on allodynia. This lack of effect may reflect the tow
affinity of
the compounds (965nM and 100nM respectively). However, intrathecal
administration of PD 184352 or PD 254552 (30~g) blocked the maintenance
of static allodynia in CCI animals (see FIG. 8). These compounds possess
higher affinity (2 and 5 nM respectively). The antiallodynic effect was only
evident for 30min post-injection and thus, shorter than the one observed for
pregabalin (100~.g). The magnitude of the effect was similar for 30~g of PD
184352 and 100~g of pregabalin.
The results indicate that MEK inhibitors exert an antiallodynic effect in
CCI-induced neuropathic rats when administered intrathecally, and that the
antiallodynic effect correlates with the affinity of the compounds.
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CHEMICAL EXAMPLES
EXAMPLE 1
S Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzoimidazole-
5-carboxylic acid (PD 205293) (APK ICSO = 14 nM; colon 26 cells, ICSO = > 10
micromolar)
Step a: Preparation of 5-nitro-2,3,4-trifluorobenzoic acid
To gently stirring concentrated sulfuric acid (50 ml) was added fuming
nitric acid (3.4 ml, 0.076 mol). Solid 2,3,4-trifluorobenzoic acid (10.00 g,
0.05565 mol) was added directly in increments. After stirring 45 minutes, the
reaction mixture had become an orange homogeneous solution which was
then poured over chilled water (400 ml). The resulting aqueous suspension
was extracted with diethyl ether (3 x 200 ml). The combined extracts were
dried with anhydrous magnesium sulfate and concentrated in vacuo to yield
12.30 g of a dull, light-yellow solid. Recrystallization from chloroform (50
ml)
afforded 9.54 g of the pale yellow microcrystalline product; 78 % yield; m.p.
;
' H-NMR (400 MHz; DMSO) 8 14.29 (broad s, 1 H), 8.43-8.38 (m, 1 H); '3C-
NMR (100 MHz; DMSO) 8 162.41, 154.24 (dd, J~_F=270.1, 10.7 Hz), 148.35
(dd, J~_F=267.0, 9.2 Hz), 141.23 (dt, J~_F=253.4 Hz), 133.95, 123.30 (d, J~_
F=2.2 Hz), 116.92 (dd, J~_F=18.2, 3.8 Hz); '9F-NMR (376 MHz; DMSO) s -
120.50 to -120.63 (m), -131.133 to -131.27 (m), -153.63 to -153.74 (m).
Step b: Preparation of 4-amino-2,3-difluoro-5-nitrobenzoic acid
Solid 5-vitro-2,3,4-trifluorobenzoic acid (0.75 g, 0.00339 mol) was
dissolved in concentrated ammonium hydroxide (25 ml) to give instantly a
yellow solution. A precipitate began to form within five minutes, after which
time the mixture was acidified to pH 0 with concentrated aqueous hydrochloric
acid. A yellow precipitate rapidly formed. The mixture was heated to boiling
and was filtered hot. The yellow solids were washed with 10 % aqueous
hydrochloric acid and were suction dried to afford 0.47 g of a yellow powder;
64 % yield; ' H-NMR (400 MHz; DMSO) 8 13.32 (s, 1 H), 8.36 (d, 1 H, J=7.6
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Hz), 7.98 (s, 2H); '9F-NMR (376 MHz; DMSO) 8 -128.69 to -128.76 (m),
153.60 (d).
Step c: Preparation of methyl 4-amino-2,3-difluoro-5-nitrobenzoate
Hydrogen chloride gas was dissolved in anhydrous methanol (30 ml)
until the solution was warm. The solid 4-amino-2,3-difluoro-5-nitrobenzoic
acid (0.47 g; 0.00215 mol) was dissolved in this solution and the reaction
mixture was brought to reflux with vigorous stirring for 23 hours under a
nitrogen atmosphere. The reaction mixture was allowed to cool slowly on the
bench. A yellow precipitate formed and was collected by vacuum filtration and
dried with suction to afford 0.35 g of yellow microfilaments; 70 % yield; m.p.
183.5-184 °C; ' H-NMR (400 MHz; DMSO) 8 8.36 (dd, 1 H, J=7.3, 1.7 Hz),
8.06 (s, 2H), 3.78 (s, 3H); '9F-NMR (376 MHz; DMSO) b -128.85 to -128.92
(m), -153.29 (d); MS (APCI-) 231 (M-1, 100); IR (KBr) 3433, 3322, 1700,
1650, 1549, 1343, 1285 cm~'; Anal. calcd/found for: C8H6F2N204 C,
41.39/41.40; H, 2.61 /2.50; N, 12.07/11.98; F, 16.37/16.58.
Step d: Preparation of methyl 4-amino-3-fluoro-2-(2-methyl-phenylamino)-5-
nitrobenzoate
The solid methyl 4-amino-2,3-difluoro-5-nitrobenzoate (0.087 g, 3.7 x
10~ mol) was dissolved in ortho-toluidine (3 ml, 0.028 mol). The reaction
mixture was stirred at 200 °C for 35 minutes under a nitrogen
atmosphere.
The mixture was then partitioned between diethyl ether (150 ml) and 10
aqueous hydrochloric acid (150 ml). The ether phase was dried with
anhydrous magnesium sulfate and was concentrated in vacuo to a crude
solid. The crude product was dissolved in 5 ml of dichloromethane and was
filtered through a flash silica plug. Elution with dichloromethane afforded
0.0953 g of a yellow solid; 81 % yield; m.p. 164-168 °C;'H-NMR (400
MHz;
DMSO) 8 9.20 (s, 1 H), 8.52 (d, 1 H, J=1.7 Hz), 7.57 (s, 2H), 7.19 (d, 1 H,
J=7.3
Hz), 7.12-7.08 (m, 1 H), 7.02-6.98 (m, 1 H), 6.95-6.91 (m, 1 H), 3.78 (s, 3H),
2.21 (s, 3H); '9F-NMR (376 MHz; DMSO) b -141.13 (s); MS (APCI+) 320
(M+1, 100); (APCI-) 318 (M-1, 100); IR (KBr) 3467, 3346, 1690, 1305 cm-';
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Anal. calcd/found for: C,5H~4FN3O4' 0.21 H20 C, 55.77/55.97; H, 4.50/4.55; N,
13.01 /12.61; F, 5.88/5.95.
Step e: Preparation of methyl 4.5-diamino-3-fluoro-2-(2-methyl-
phenylamino)benzoate
To a mixture comprised of methyl 4-amino-3-fluoro-2-(2-methyl-
phenylamino)-5-nitrobenzoate (2.52 g, 0.00789 mol), tetrahydrofuran (50 ml),
methanol (50 ml) and washed Raney nickel (0.5 g) was initially applied 48.6
psi of hydrogen gas at 30.2 °C in a shaker for 4 hours 48 minutes. The
mixture was filtered and the filtrate concentrated in vacuo to afford 2.20 g
of a
salmon-colored amorphous solid; 96 % yield; ' H-NMR (400 MHz; DMSO) 8
7.84 (s, 1 H), 7.04 (d, 1 H, J=7.1 Hz), 6.98 (d, 1 H, J=1.2 Hz), 6.95-6.91 (m,
1 H),
6.68-6.64 (m, 1 H), 6.40-6.36 (m, 1 H), 5.39 (s, 2H), 4.73 (s, 2H), 3.66 (s,
3H),
2.21 (s, 3H); '9F-NMR (376 MHz; DMSO) 8 -139.66 (s).
Step f: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylate
A stirring solution comprised of methyl 4,5-diamino-3-fluoro-2-(2-
methyl-phenylamino)-benzoate (1.78 g, 0.00615 mol) in formic acid (Aldrich,
95-97 %, 100 ml, 2.5 mol) was brought to reflux for 3 hours followed by
concentration in vacuo to give a crude brown solid. The crude product was
triturated with chloroform (40 ml) and subsequently collected by vacuum
filtration. The solids were dried with suction to afford 1.09 g of a light-
lavender
powder. The filtrate was concentrated in vacuo to a crude solid which was
triturated with 10 ml of chloroform-dichloromethane. These solids were
collected by vacuum filtration, rinsed with dichloromethane, and were suction-
dried to give an additional 0.55 g of a light-lavender powder (total yield:
1.64
g); 87 % yield; m.p. 259-262 °C; ' H-NMR (400 MHz; DMSO) b 8.42 (s, 1
H),
8.03 (s, 1 H), 7.93 (broad s, 1 H), 7.12 (d, 1 H, J=7.0 Hz), 6.99-6.95 (m, 1
H),
6.75-6.71 (m, 1 H), 6.48-6.44 (m, 1 H), 3.81 (s, 3H), 2.30 (s, 3H); '9F-NMR
(376
MHz; DMSO) 8 -132.84 (s); MS (APCI+) 300 (M+1, 100); (APCI-) 298 (M-1,
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100); IR (KBr) 3322, 1689, 1437, 1326, 1218 cm-'; Anal. calcd/found for:
C~6H~4FN302' 0.32 H20 C, 62.99/63.01; H, 4.84/4.61; N, 13.77/13.70.
Step g: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylate
A stirring mixture comprised of methyl 7-fluoro-6-(2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylate (0.2492 g, 8.326 x 10~ mol),
benzyltrimethylammonium dichloroiodinate (Aldrich, 95 %, 0.3934 g, 0.00113
mol), and zinc chloride (0.1899 g, 0.00139 mol) in glacial acetic acid (20 ml)
was brought to reflux for 15 minutes. The hot suspension was filtered to
isolate the precipitate which was dried in the vacuum oven (90 °C, ca.
10 mm
Hg) overnight to afford 0.2392 g of a green powder; 68 % yield; m.p. 219-220
°C DEC; ' H-NMR (400 MHz; DMSO) b 8.71 (s, 1 H), 8.02 (s, 1 H), 7.85
(broad
s, 1 H), 7.43 (d, 1 H, J=1.7 Hz), 7.24 (dd, 1 H, J=8.5, 2.2 Hz), 6.24 (dd, 1
H,
J=8.5, 5.4 Hz), 3.76 (s, 3H), 2.22 (s, 3H); '9F-NMR (376 MHz; DMSO) 8 -
132.86 (s); MS (APCI+) 426 (M+1, 48), 169 (100);
(APCI-) 424 (M-1, 100); IR (KBr) 1704, 1508, 1227 cm-'
Step h: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid
To a stirring solution comprised of methyl 7-fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylate (0.2035 g, 4.786 x 10~ mol)
in tetrahydrofuran (20 ml) was added solid potassium trimethylsilanolate
(0.315 g, 0.00246 mol). The reaction mixture was stirred at ambient
temperature under argon for 16 hours. An additional 0.082 g (6.39 x 10~ mol)
of potassium trimethylsilanolate was added and the mixture stirred 30
minutes. The reaction mixture was concentrated in vacuo to one-third volume
and was treated with diethyl ether (50 ml). The off-white precipitate formed
was collected by vacuum filtration, giving a hygroscopic solid. The wet solid
was dissolved in a 4:1 (v/v) ethyl acetate-methanol solution (500 ml). The
solution was washed with 0.84 M aqueous citric acid (50 ml), dried (MgS04),
and concentrated in vacuo to a yellow liquid. The liquid was redissolved in
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fresh ethyl acetate-methanol. The solution was washed with brine, dried
(MgS04), and concentrated in vacuo. The residue was redissolved in
chloroform and reconcentrated to afford 1.55 g of a viscous yellow residue
which was comprised mainly of citric acid; MS (APCI-) 191 (M-1, 100). The
residue was dissolved in water (50 ml). Insoluble material was extracted into
1:1 (v/v) ethyl acetate-diethyl ether (250 ml). Upon separation, the aqueous
phase remained strongly acidic (pH 0). The organic phase was washed with a
fresh portion of water (150 ml). Upon separation, this wash was only slightly
acidic (pH 4.5). The organic phase was dried (MgS04), concentrated in
vacuo, and chased with chloroform to give a tan semisolid. The product was
triturated with hexanes. Vacuum filtration and suction-drying afforded 0.0839
g of a tan powder. A portion of the product (0.050 g) was recrystallized from
boiling ethanol (1 ml). While cooling and moderate scratching, an off-white
solid formed. This product was isolated by vacuum filtration and dried under
high vacuum (23 °C) to afford 0.018 g of an off-white powder; 9 %
yield; m.p.
247-248 °C DEC; '9F-NMR (376 MHz; DMSO) 8 -132.87 (s); MS (APCI+) 412
(M+1, 100); (APCI-) 410 (M-1, 100); IR (KBr) 3322, 1689, 1437, 1326,
1218 cm-'; Anal. calcd/found for: C~5H»FIN302' 0.61 C2H60 ' 0.59 H20 (91.4
parent) C, 43.30/43.30; H, 3.55/3.34; N, 9.34/9.15.
EXAMPLE 2
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzoimidazole-
5-carboxylic acid cyclopropylmethoxy-amide (PD 254552) (APK IC5o < 10 nM
(n = 2); colon 26 cells, 1 hour pretreatment, ICSO = 20 nM)
Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid pentafluorophenyl ester (PD 254551 ) (APK
IC5o = 120 nM (n=2))
To a stirring suspension comprised of 7-fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylic acid (0.844 g, 2.05x10-3 mol) in
ethyl acetate (4 ml) was added a solution comprised of pentafluorophenol
(0.375 g, 2.04x10-3 mol) in N,N-dimethylformamide (10 ml). Solid
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dicyclohexylcarbo-diimide (0.415 g, 1.99x10-3 mol) was then added and the
reaction mixture was stirred for 22 hours. The reaction mixture was vacuum
filtered to remove the precipitate that had formed. The filtrate was diluted
with
ethyl acetate (400 ml), and that solution was washed with water (3x400 ml),
was dried (MgS04), and was concentrated in vacuo to afford 1.7 g of a yellow
foam. The crude product was purified by flash silica column chromatography.
Elution with a gradient (CHC13 to 0.5 % methanol in CHC13) afforded 0.69 g of
the yellow amorphous product; 60 % yield;'H-NMR (400 MHz; CDC13) 8 8.54
(s, 1 H), 8.28 (s, 1 H), 8.04 (s, 1 H), 7.49 (d, 1 H, J=1.7 Hz), 7.36 (dd, 1
H, J=8.2,
1.7 Hz), 6.57 (dd, 1 H, J=8.4, 6.5 Hz), 2.31 (s, 3H); '9F-NMR (376 MHz;
CDC13) b -132.02 (s), -152.35 (d, J=18.3 Hz), -157.26 (t, J=21.4 Hz), -161.96
(dd, J=21.3, 18.3 Hz); MS (APCI+) 578 (M+1, 57), 394 (100); (APCI-) 576 (M-
1, 44), 409 (100), 393 (95), 392 (82), 378 (55), 183 (97), 165 (68), 127 (53);
IR
(KBr) 1731 cm-' (C=O stretch).
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid cyclopropylmethoxy-amide
To a stirring solution comprised of 7-fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzoimidazole-5-carboxylic acid pentafluorophenyl ester
(0.63 g, 1.09x10'3 mol) in anhydrous tetrahydrofuran (5 ml) was added solid
cyclopropylmethoxylamine hydrochloride (0.14 g, 1.13x10-3 mol) and
diisopropylethylamine (0.6 ml, 3.4x10-3 mol). The reaction mixture was stirred
for one week. The solvent was removed and the evaporate was treated with
10 % aqueous hydrochloric acid (200 ml) and was extracted with diethyl ether
(200 ml). A biphasic suspension resulted, and the precipitate was isolated by
vacuum filtration. The crude product was recrystallized from absolute ethanol
to afford 0.18 g of a green-yellow powder; 35 % yield; mp 168-172 °C; '
H-
NMR (400 MHz; DMSO) 8 11.48 (s, 1 H), 8.37 (s, 1 H), 7.50 (broad s, 1 H), 7.45
(s, 1 H), 7.24 (s, 1 H), 7.07 (d, 1 H, J=8.4 Hz), 6.03-5.97 (m, 1 H), 3.38 (d,
2H,
J=6.5 Hz), 2.04 (s, 3H), 0.85-0.75 (m, 1 H), 0.30-0.22 (m, 2H), 0.00 (s, 2H);
'9F-NMR (376 MHz; DMSO) 8 -133.23 (s); MS (APCI+) 481 (M+1, 77), 409
(100); (APCI-) 480 (M, 22), 407 (100); IR (KBr) 1659, 1632, 1493 cm~'; Anal.
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calcd/found for: C~9H~8FIN402' 0.50 HCI (96.3 % parent) C, 45.78/45.74; H,
3.74/3.84; N, 11.24/10.88.
EXAMPLE 3
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzoimidazole-
5-carboxylic acid hydroxyamide
Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide
A solution comprised of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid, O-(tetrahydro-2H-pyran-2-yl)-
hydroxylamine (1.25 equiv.), benzotriazole-1-yl-oxy-Iris-pyrrolidino-
phosphonium hexafluorophosphate (1.25 equiv.), and diisopropylethylamine
(3 equiv.) in 1:1 v/v tetrahydrofuran-dichloromethane is stirred for 30
minutes.
The reaction mixture is concentrated in vacuo and the residue is purified by
flash chromatography; elution with dichloromethane affords the desired
product. The product may be recrystallized with an appropriate solvent like
methanol if further purification is necessary.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid hydroxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H
benzoimidazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide is
dissolved in an appropriate hydrogen chloride-saturated solvent like methanol
or ethanol. Once homogeneous, the solution is concentrated in vacuo to give
the desired product. The product may be triturated with an appropriate
solvent like chloroform or dichloromethane if further purification is
necessary.
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EXAMPLE 4
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzoimidazole-
5-carboxylic acid cyclopropylmethoxy-amide
Step a: Preparation of O-cyclopropylmethylhydroxylamine hydrochloride
Step i: Preparation of 2-cylcopropylmethoxy-isoindole-1,3-dione
To a stirring solution/suspension comprised of N-hydroxyphthalimide (Aldrich,
57.15 g, 339.8 mmol), cyclopropanemethanol (Aldrich, 25.10 g, 341.1 mmol),
and triphenylphosphine ("DEAD," Aldrich, 91.0 g, 344 mmol) in 1.00 L of
tetrahydrofuran under a nitrogen atmosphere and cooled to 6 °C
(internal
mixture temperature) with an ice-water bath was added diethyl
azodicarboxylate (Aldrich, 56 ml, 356 mmol) dropwise over 20 minutes via
addition funnel. The reaction mixture temperature was kept below 20 °C
during the addition. Following addition of the DEAD, the cold bath was
removed and the reaction mixture was stirred for 15 hours. The mixture was
concentrated to a paste under reduced pressure. Chloroform (ca. 300 ml)
was added and the mixture swirled to loosen all solids. Vacuum filtration
removed the insolubles. The filtrate was likewise filtered to remove white
precipitate that formed and to give a clear filtrate. Concentration under
reduced pressure afforded a clear oil. Flash filtration through silica gel
(100
chloroform) gave filtrates containing unseparated product. These filtrates
were combined and concentrated under reduced pressure to afford 127.4 g of
a clear oil. The oil was dissolved in absolute ethanol (400 ml) and the
solution
was refrigerated for two hours. A white crystalline solid had precipitated and
was subsequently collected by vacuum filtration. The product was dried in the
vacuum oven (60 °C) to afford 42.66 g (58 %) of the desired material;
m.p. 71-
77 °C;'H-NMR (400 MHz; CDC13 signal offset to 8 6.96) 8 7.54-7.43 (m,
4H),
3.74 (d, 2H, J=7.6 Hz), 1.02-0.95 (m, 1 H), 0.34-0.30 (m, 1 H), 0.04-0.00 (m,
1 H).
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Step ii: Preparation of O-cyclopropylmethylhydroxylamine hydrochloride
To a stirring solution comprised of 2-cyclopropylmethoxy-isoindole-1,3-dione
(42.64 g, 196.3 mmol) in 150 ml of dichloromethane under ambient conditions
was carefully added methylhydrazine (Aldrich, 10.7 ml, 197 mmol). A white
precipitate began to form almost instantly. After 15 minutes of vigorous
stirring, the suspension was vacuum filtered. The filtrate was likewise
filtered
to remove additional precipitate. The resulting clear filtrate was
concentrated
carefully (volatile product) under reduced pressure to afford a clear
liquid/solid
mixture. The white solids were removed when an ether (200 ml) solution of
the product was made and vacuum filtered. The filtrate was acidified with
gaseous hydrogen chloride, affording instantly a white precipitate. Collection
of the solid by vacuum filtration and vacuum-oven drying (55 °C)
afforded 18.7
g (77 %) of the white powder product; m.p. 165-168 °C;'H-NMR (400 MHz;
DMSO) 8 10.77 (broad s, 2H), 3.57 (d, 2H, J=7.3 Hz), 0.84-0.74 (m, 1 H), 0.31-
0.25 (m, 2H), 0.04-0.00 (m, 1 H); '3C-NMR (100 MHz; DMSO) s 75.39, 5.52,
0.00.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenYlamino)-1 H-
benzoimidazole-5-carboxylic acid cyclopropylmethoxy-amide
A solution comprised of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzoimidazole-5-carboxylic acid, O-cyclopropylmethylhydroxylamine
hydrochloride (1.25 equiv.), benzotriazole-1-yl-oxy-tris-pyrrolidino-
phosphonium hexafluorophosphate (1.25 equiv.), and diisopropylethylamine
(3 equiv.) in 1:1 v/v tetrahydrofuran-dichloromethane is stirred for 30
minutes.
The reaction mixture is concentrated in vacuo and the residue is taken up into
diethyl ether. The ether phase is washed with dilute aqueous hydrochloric
acid, saturated aqueous sodium bicarbonate, and brine, is dried (MgS04), and
is concentrated in vacuo to afford the desired product. The product may be
recrystallized with an appropriate solvent like methanol or chloroform if
further
purification is necessary.
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EXAMPLE 5
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzooxazole-5-
carboxylic acid
Step a: Preparation of 5-nitro-2 3 4-trifluorobenzoic acid
Same as for Example 1, Step a.
Step b: Preparation of 2 3-difluoro-4-hydroxy-5-nitrobenzoic acid
The solid 5-nitro-2,3,4-trifluorobenzoic acid (1.00 g, 0.00452 mol) was
dissolved in 10 wt. % aqueous sodium hydroxide solution. The mixture was
clear deep orange. After standing under ambient conditions for several
minutes, the mixture was quenched with concentrated aqueous hydrochloric
acid until strongly acidic (pH 0). A white solid precipitated which was
isolated
by vacuum filtration and dried with suction to afford 0.40 g of an off-white
solid. This solid was recrystallized from chloroform (20 ml) to afford 0.22 g
of
an off-white crystalline powder; 22 % yield; MS (APCI-) 218 (M-1, 100).
Step c: Preparation of methyl 2 3-difluoro-4-hydroxy-5-nitrobenzoate
Anhydrous hydrogen chloride gas was dissolved in anhydrous
methanol (50 ml) until the solution was warm. The microcrystalline solid 2,3
difluoro-4-hydroxy-5-nitrobenzoic acid 0.22 g, 0.00100 mol) was dissolved in
the methanolic hydrogen chloride solution. The stirring reaction mixture was
brought to reflux under nitrogen for 16 hours. The mixture was concentrated
in vacuo to give a white solid. The product was dried under high vacuum to
afford 0.213 g of a white powder; 91 % yield; m.p. 108-109.5 °C;'H-NMR
(400 MHz; DMSO) 8 8.25 (dd, 1 H, J=7.7, 2.2 Hz), 3.83 (s, 3H); (CDC13) 8
10.83 (s, 1 H), 8.66 (dd, 1 H, J=7.0, 2.2 Hz), 3.98 (s, 3H); '9F-NMR (376 MHz;
DMSO) 8-127.85 (s), -154.32 (d, J=19.8 Hz); (CDC13) 8 -118.31 to -118.37
(m), -152.38 (d, J=18.3 Hz); MS (APCI-) 232 (M-1, 100); IR (KBr) 3264, 1731,
1640, 1546, 1307, 1286, 1160 cm-' .
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Step d: Preparation of 1-adamantyl 4-carboxymethyl-2,3-difluoro-6-nitrophenyl
carbonate
To a solution comprised of 1-adamantyl fluoroformate (2.0 M) and
pyridine (2.0 M) in tetrahydrofuran is added a stirred solution comprised of
methyl 2,3-difluoro-4-hydroxy-5-nitrobenzoate (0.96 equiv., 0.384 M) in
anhydrous tetrahydrofuran at ambient temperature. The reaction mixture is
stirred for 6 hours and the solvent is removed in vacuo. The residue is
dissolved in dichloromethane. The organic solution is washed with dilute
aqueous hydrochloric acid, dilute aqueous sodium carbonate, and water, is
dried (MgS04), and is concentrated in vacuo to give the desired product.
Step e: Preparation of 1-adamantyl 4-carboxymethyl-2-fluoro-3-f2-methyl-
phenylamino)-6-nitrophenyl carbonate
The compound 1-adamantyl 4-carboxymethyl-2,3-difluoro-6-nitrophenyl
carbonate is dissolved in excess ortho-toluidine. The reaction mixture is
stirred at 200 °C for 6 hours. The mixture is allowed to cool and is
dissolved
in diethyl ether. The organic phase is washed with dilute aqueous
hydrochloric acid, saturated aqueous sodium bicarbonate, and brine, is dried
(MgS04), and is concentrated in vacuo to afford the desired product. The
product is purified by flash chromatography as necessary.
Step f: Preparation of methyl 3-fluoro-4-hydroxy-2-(2-methyl-phenylamino)-5-
nitrobenzoate
The compound 1-adamantyl 4-carboxymethyl-2-fluoro-3-(2-methyl
phenylamino)-6-nitrophenyl carbonate is dissolved in excess trifluoroacetic
acid at ambient temperature. The mixture is stirred for 20 minutes. The TFA
is removed under reduced pressure. The residue is subjected to vacuum
pump to remove adamantan-1-of to give the desired product.
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Step g: Preparation of methyl 5-amino-3-fluoro-4-hydroxy-2-(2-methyl-
phenylamino)-benzoate
The compound methyl 3-fluoro-4-hydroxy-2-(2-methyl-phenylamino)-5-
nitrobenzoate is treated as in Step e, Example 1.
Step h: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylate
The compound 5-amino-3-fluoro-4-hydroxy-2-(2-methyl-phenylamino)-
benzoate is treated as in Step f, Example 1. The product may be
recrystallized with an appropriate solvent like chloroform or ethanol if
further
purification is necessary.
Step i: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carbox 1y ate
A stirring mixture comprised of methyl 7-fluoro-6-(2-methyl-
phenylamino)-1 H-benzooxazole-5-carboxylate (0.042 M),
benzyltrimethylammonium dichloroiodinate (Aldrich, 95 %, 0.057 M, 1.36
equiv.), and zinc chloride (0.070 M, 1.67 equiv.) in glacial acetic acid is
brought to reflux for 15 minutes. The mixture is concentrated in vacuo and the
residue taken up into diethyl ether. The ether solution is washed with dilute
aqueous hydrochloric acid, water, and brine, is dried (MgS04), and is
concentrated in vacuo to obtain the desired product. The product may be
purified by recrystallization with an appropriate solvent like ethanol.
Step j: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid
To a stirring solution comprised of methyl 7-fluoro-6-(4-iodo-2-methyl-
phenylamino)-1 H-benzooxazole-5-carboxylate (0.024 M) in tetrahydrofuran is
added solid potassium trimethylsilanolate (5.14 equiv.). The reaction mixture
is stirred at ambient temperature under argon for 16 hours. An additional
equivalent of potassium trimethylsilanolate is added and the mixture stirred
30
minutes. The reaction mixture is concentrated in vacuo to give a residue that
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is then taken up into 1:1 (v/v) ethyl acetate-diethyl ether. The organic phase
is washed with dilute aqueous hydrochloric acid, water, and brine, is dried
(MgS04), is concentrated in vacuo, and chased with chloroform to give a
crude product. Recrystallization from an appropriate solvent like ethanol
gives
the purified desired product.
EXAMPLE 6
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzooxazole-5-
carboxylic acid hydroxyamide
Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid hydrox~ramide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide is
treated as in Step b, Example 2.
EXAMPLE 7
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzooxazole-5-
carboxylic acid cyctopropylmethox~ amide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzooxazole-5-carboxylic acid is treated as in Step b, Example 3.
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EXAMPLE 8
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzothiazole-5-
carboxylic acid
Step a: Preparation of 5-vitro-2 3 4-trifluorobenzoic acid
Same as for Example 1, Step a.
Step b: Preparation of 2 3-difluoro-4-hydroxy-5-nitrobenzoic acid
Same as for Example 4, Step b.
Step c: Preparation of methyl 2 3-difluoro-4-hydroxy-5-nitrobenzoate
Same as for Example 4, Step c.
Step d: Preparation of 4-dimethylthiocarbamoyloxy-2,3-difluoro-5-nitro-
benzoic acid methyl ester
A solution of methyl 2,3-difluoro-4-hydroxy-5-nitrobenzoate in N,N-
dimethylformamide is treated with one molar equivalent of cesium carbonate
and warmed to 85 °C for 30 minutes. The stirring mixture is then
treated
dropwise rapidly with a solution comprised of a slight excess of N,N-
dimethylthiocarbamoyl chloride in N,N-dimethylformamide. The reaction
mixture is stirred at room temperature for one hour, or may be warmed over a
steam bath for one hour. The mixture is then poured into water and extracted
with ethyl acetate. The organic phase is washed with 5 % aqueous sodium
hydroxide, water, and brine, and is then dried with a drying agent like
magnesium sulfate of sodium sulfate. The solvent is then removed in vacuo
to give a crude product. The compound is purified by ordinary methods such
as chromatography or crystallization from an appropriate solvent.
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Step e: Preparation of 4-Dimethylthiocarbamoyloxy-3-fluoro-5-nitro-2-o-
tolylamino-benzoic acid methyl ester
The compound 4-dimethylthiocarbamoyloxy-2,3-difluoro-5-nitro-benzoic
acid methyl ester is dissolved in excess o-toluidine. The stirring mixture is
brought to 200 °C for one hour. The mixture is then poured into 5 %
aqueous
hydrochloric acid. The aqueous mixture is extracted with diethyl ether. The
organic phase is washed with water and brine, is dried over magnesium
sulfate, and is concentrated in vacuo. The crude product is purified by
ordinary methods such as chromatography or crystallization from an
appropriate solvent.
Step f: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylate
The compound methyl 5-amino-3-fluoro-4-mercapto-2-(2-methyl-
phenylamino)-benzoate is treated as in Step h, Example 4.
Step g: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylate
The compound methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylate is treated as in Step i, Example 4.
Step h: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylic acid
The compound methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylate is treated as in Step j, Example 4.
EXAMPLE 9
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzothiazole-5-
carboxylic acid hydroxyamide
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Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazoie-5-carboxylic acid hydroxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide is
treated as in Step b, Example 2.
EXAMPLE 10
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzothiazole-5-
carboxylic acid cyclopropylmethoxy=amide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzothiazole-5-carboxylic acid is treated as in Step b, Example 3.
EXAMPLE 11
Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-puinoxaline-6-
carboxylic acid
Step a: Preparation of 8-fluoro-7-(2-methyl-phenylamino)-puinoxaline-6-
carboxylic acid
The compound methyl 4,5-diamino-3-fluoro-2-(2-methyl-phenylamino)-
benzoate (from Step e, Example 1 ) is dissolved in 2:1:1.2 v/v/v of 2.0 M
acetic
acid-4.0 M sodium acetate-methanol. The suspension is warmed to 65 °C
(or
until homogeneous) and the clear solution is poured into a 0.078 M aqueous
sodium glyoxal bisulfite (Aldrich, monohydrate, 1.05 equiv.) solution which is
warmed to 70 °C. The reaction mixture is stirred gently between 55-75
°C for
one hour, and is then cooled to 12 °C with an ice-water bath.
Pulverized
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sodium hydroxide pellets (27 equiv.) are added to the cold solution. The
mixture is gently warmed to 30 °C and stirred for 45 minutes. The
temperature is raised to 70 °C for 15 minutes. The mixture is allowed
to cool
and is treated with ethyl acetate. The biphasic mixture is treated with
concentrated aqueous hydrochloric acid to achieve pH 0 in the aqueous
phase. The organic phase is separated, dried (MgS04), and concentrated in
vacuo to give the desired product. The product may be triturated with an
appropriate solvent like dichloromethane or recrystallized from a solvent like
ethanol for further purification as necessary.
Step b: Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-4uinoxaline-
6-carboxylic acid
The compound 8-fluoro-7-(2-methyl-phenylamino)-quinoxaline-6-
carboxylic acid is treated as in Step i, Example 4.
EXAMPLE 12
Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-guinoxaline-6-
carboxylic acid hydroxyamide
Step a: Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-ctuinoxaline-
6-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide
The compound 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-
6-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-auinoxaline-
6-carboxylic acid hydroxyamide
The compound 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-
6-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide is treated as in Step
b, Example 2.
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EXAMPLE 13
Preparation of 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-auinoxaline-6-
carboxylic acid cyclopropylmethoxy-amide
The compound 8-fluoro-7-(4-iodo-2-methyl-phenylamino)-quinoxaline-
6-carboxylic acid is treated as in Step b, Example 3.
EXAMPLE 14
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51thiadiazole-5-carboxylic acid
Step a: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzof 1,2,51thiadiazole-5-carboxylate
To a stirring solution comprised of methyl 4,5-diamino-3-fluoro-2-(2-methyl-
phenylamino)-benzoate (from Step e, Example 1 ) and diisopropylethylamine
(2 equiv.) in an appropriate solvent like diethyl ether or toluene is added a
reagent like N-thioaniline or thionyl chloride (1.35 equiv.). The reaction
mixture is brought to reflux for one hour. The mixture is quenched with dilute
aqueous hydrochloric acid. The organic phase is washed with saturated
aqueous sodium bicarbonate and brine, is dried (MgS04), and is concentrated
in vacuo to afford the desired product. The product may be recrystallized with
an appropriate solvent like chloroform or ethanol, or may be chromatographed
if further purification is necessary.
Alternative method: The compound methyl 4,5-diamino-3-fluoro-2-(2-methyl-
phenylamino)-benzoate is added to a stirring solution of sulfur monochloride
(6 equiv.) in N,N-dimethylformamide and the mixture is gradually heated to
75-80 °C. After 5 hours the mixture is cooled to 10 °C, water is
slowly added.
The mixture is extracted with a solvent like diethyl ether or dichloromethane.
The organic extract is dried (MgS04) and is concentrated in vacuo to afford
the desired product. The product may be recrystallized with an appropriate
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solvent like chloroform or ethanol, or may be chromatographed if further
purification is necessary.
Step b: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51thiadiazole-5-carboxylate
The compound methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzo[1,2,5]thiadiazole-5-carboxylate is treated as in Step i, Example 4.
Step c: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51thiadiazole-5-carboxylic acid
The compound methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]thiadiazole-5-carboxylate is treated as in Step j, Example 4.
EXAMPLE 15
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof1 2,51thiadiazole-5-carboxylic acid hydroxyamide
Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1 2 5lthiadiazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-
oxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]thiadiazole-5-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1 2 5lthiadiazole-5-carboxylic acid hydroxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)
benzo[1,2,5]thiadiazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)
oxyamide is treated as in Step b, Example 2.
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EXAMPLE 16
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2.51thiadiazole-5-carboxylic acid cyclopropylmethoxy-amide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]thiadiazole-5-carboxylic acid is treated as in Step b, Example 3.
EXAMPLE 17
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1.2.51oxadiazole-5-carboxylic acid
Step a: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzof1,2,51oxadiazole-5-carboxylate 2-oxide
1 S See Takakis, I. M.; Hadjimihalakis, P. M., J. Heterocyclic Chem., 27,
177 (1990).
A mixture comprised of methyl 4-amino-3-fluoro-2-(2-methyl-
phenylamino)-5-nitrobenzoate (from Step d, Example 1 ) and
iodosobenzenediacetate (1.76 equiv.) in benzene is stirred at ambient
temperature for 5 hours. The mixture is concentrated in vacuo and the
residue purified by column chromatography to give the desired product.
Alternative method: A solution comprised of methyl 4-amino-3-fluoro-2-(2-
methyl-phenylamino)-5-nitrobenzoate (0.86 M) in tetrahydrofuran is diazotized
and the diazonium salt is treated in situ with sodium azide as described by
Smith, P. A. S.; Boyer, J. H., Org. Synth., 31, 14 (1951) and references 4 and
8 cited therein. Thermolysis of this intermediate in ethylene glycol at 110-
120
°C for one hour affords the desired product.
Step b: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzof 1.2.51oxadiazole-5-carboxylate
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A solution comprised of methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylate 2-oxide and sodium azide (1.38 equiv.)
in ethylene glycol is heated to 140-150 °C for 30 minutes to obtain,
after
column chromatography, the desired product.
Step c: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo(1.2.51oxadiazole-5-carboxylate
The compound methyl 7-fluoro-6-(2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylate is treated as in Step i, Example 4.
Step d: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51oxadiazole-5-carboxylic acid
The compound methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylate is treated as in Step j, Example 4.
EXAMPLE 18
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phen~rlamino)-
benzoj1.2,51oxadiazole-5-carboxylic acid hydroxyamide
Step a: Preparation of 7-fluoro-6-(4-iodo-2-meth~rl-phenylamino)-
benzof 1,2.51oxadiazole-5-carboxylic acid O-(tetrahydro-2H pYran-2-y~
oxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51oxadiazole-5-carboxylic acid hydroxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-
oxyamide is treated as in Step b, Example 2.
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EXAMPLE 19
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzof 1,2,51oxadiazole-5-carboxylic acid cyclopropylmethoxy-amide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-
benzo[1,2,5]oxadiazole-5-carboxylic acid is treated as in Step b, Example 3.
EXAMPLE 20
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzotriazole-5-
carboxylic acid
Step a: Preparation of methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylate
The compound methyl 4,5-diamino-3-fluoro-2-(2-methyl-phenylamino)-
benzoate (from Step e, Example 1 ) is diazotized by ordinary methods.
Workup gives the desired product.
Step b: Preparation of methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylate
The compound methyl 7-fluoro-6-(2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylate is treated as in Step i, Example 4.
Step c: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid
The compound methyl 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylate is treated as in Step j, Example 4.
EXAMPLE 21
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzotriazole-5-
carboxylic acid hydroxyamide
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Step a: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid is treated as in Step a, Example 2.
Step b: Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid hydroxyamide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid O-(tetrahydro-2H-pyran-2-yl)-oxyamide is
treated as in Step b, Example 2.
EXAMPLE 22
Preparation of 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-benzotriazole-5-
carboxylic acid cyclopropylmethoxy-amide
The compound 7-fluoro-6-(4-iodo-2-methyl-phenylamino)-1 H-
benzotriazole-5-carboxylic acid is treated as in Step b, Example 3.
F. OTHER EMBODIMENTS
From the above disclosure and examples, and from the claims below,
the essential features of the invention are readily apparent. The scope of the
invention also encompasses various modifications and adaptations within the
knowledge of a person of ordinary skill. Examples include a disclosed
compound modified by addition or removal of a protecting group, or an ester,
pharmaceutical salt, hydrate, acid, or amide of a disclosed compound.
Publications cited herein are hereby incorporated by reference in their
entirety.
What is claimed is:
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