U.S. patent number RE46,024 [Application Number 14/788,921] was granted by the patent office on 2016-06-07 for thyroid hormone analogs.
This patent grant is currently assigned to Hoffmann-La Roche Inc.. The grantee listed for this patent is Hoffmann-La Roche Inc.. Invention is credited to Nancy-Ellen Haynes, Denis J. Kertesz, Sherrie Lynn Pietranico-Cole, Yimin Qian, Nathan Robert Scott, Sung-Sau So, Kshitij Chhabilbhai Thakkar, Jefferson Wright Tilley.
United States Patent |
RE46,024 |
Haynes , et al. |
June 7, 2016 |
Thyroid hormone analogs
Abstract
Provided herein are compounds of the formula (I): ##STR00001##
as well as pharmaceutically acceptable salts thereof, wherein the
substituents are as those disclosed in the specification. These
compounds, and the pharmaceutical compositions containing them, are
useful for the treatment of diseases such as obesity,
hyperlipidemia, hypercholesterolemia and diabetes and other related
disorders and diseases, and may be useful for other diseases such
as NASH, atherosclerosis, cardiovascular diseases, hypothyroidism,
thyroid cancer and other disorders and diseases related
thereto.
Inventors: |
Haynes; Nancy-Ellen (Granford,
NJ), Kertesz; Denis J. (Mountain View, CA),
Pietranico-Cole; Sherrie Lynn (Montclair, NJ), Qian;
Yimin (Wayne, NJ), Scott; Nathan Robert (Livingston,
NJ), So; Sung-Sau (Nutley, NJ), Thakkar; Kshitij
Chhabilbhai (Clifton, NJ), Tilley; Jefferson Wright
(North Caldwell, NJ) |
Applicant: |
Name |
City |
State |
Country |
Type |
Hoffmann-La Roche Inc. |
Nutley |
NJ |
US |
|
|
Assignee: |
Hoffmann-La Roche Inc. (Nutley,
NJ)
|
Family
ID: |
37114368 |
Appl.
No.: |
14/788,921 |
Filed: |
July 1, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
|
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11488870 |
Nov 18, 2008 |
7452882 |
|
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|
60701215 |
Jul 21, 2005 |
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Reissue of: |
12194643 |
Aug 20, 2008 |
7807674 |
Oct 5, 2010 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P
3/10 (20180101); C07D 237/16 (20130101); C07D
403/12 (20130101); C07D 403/10 (20130101); A61P
9/00 (20180101); A61P 3/06 (20180101); C07D
237/18 (20130101); A61P 5/14 (20180101); A61P
3/04 (20180101); A61P 5/16 (20180101); A61P
1/16 (20180101); A61P 9/10 (20180101); A61P
3/00 (20180101); C07D 237/14 (20130101); A61P
35/00 (20180101) |
Current International
Class: |
C07D
237/14 (20060101); C07D 237/16 (20060101); C07D
403/10 (20060101); C07D 403/12 (20060101); A61P
3/06 (20060101); A61P 3/04 (20060101); A61K
31/53 (20060101); A61P 3/10 (20060101); A61P
9/10 (20060101); A61P 9/00 (20060101); A61P
35/00 (20060101); C07D 253/07 (20060101); C07D
237/18 (20060101); A61K 31/50 (20060101) |
Field of
Search: |
;514/247
;544/224,182 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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188351 |
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Mar 1991 |
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EP |
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728482 |
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Aug 1996 |
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EP |
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WO 9639388 |
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Dec 1996 |
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WO |
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WO 9702023 |
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Jan 1997 |
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WO |
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WO 9857919 |
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Dec 1998 |
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WO |
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WO 9900353 |
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Jan 1999 |
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WO |
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WO 00/17204 |
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Mar 2000 |
|
WO |
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WO 2005051298 |
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Jun 2005 |
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WO |
|
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|
Primary Examiner: Huang; Evelyn
Parent Case Text
PRIORITY TO RELATED APPLICATIONS
This application is the division of U.S. application Ser. No.
11/488,870, filed Jul. 18, 2006, which claims the benefit of U.S.
Provisional Application No. 60/701,215, filed Jul. 21, 2005, which
is hereby incorporated by reference in its entirety.
Claims
What is claimed is:
1. A compound of the formula (I): ##STR00120## wherein: A is O,
CH.sub.2, S, SO or SO.sub.2; X and Y are each independently
selected from the group consisting of Br, Cl and --CH.sub.3;
R.sup.1 is selected from the group consisting of
--(CH.sub.2).sub.nCOOH, --OCH.sub.2COOH, --NHC(.dbd.O)COOH,
--NHCH.sub.2COOH, --NHC(.dbd.O)COONH.sub.2 and ##STR00121## R.sup.2
is lower alkyl; R.sup.3 is H or lower alkyl; n is 1, 2 or 3; p is 1
or 2; or a pharmaceutically acceptable salt or ester thereof.
2. The compound according to claim 1, wherein X and Y are each
Br.
3. The compound according to claim 1, wherein X and Y are each
Cl.
4. The compound according to claim 1, wherein X and Y are each
--CH.sub.3.
5. The compound according to claim 1, wherein X is Cl, and Y is
--CH.sub.3.
6. The compound according to claim 1, wherein R.sup.1 is selected
from the group consisting of --(CH.sub.2).sub.nCOOH,
--OCH.sub.2COOH, --NHC(.dbd.O)COOH, and --NHCH.sub.2COOH; and
esters thereof.
7. The compound according to claim 6, wherein R.sup.1 is
--(CH.sub.2).sub.nCOOH.
8. The compound according to claim 7, wherein n is 1.
9. The compound according to claim 6, wherein R.sup.1 is
--NHC(.dbd.O)COOH.
10. The compound according to claim 1, wherein R.sup.1 is:
##STR00122##
11. The compound according to claim 1, wherein R.sup.2 is lower
alkyl having from 1 to 3 C atoms.
12. The compound according to claim 11, wherein R.sup.2 is lower
alkyl having 3 C atoms.
13. The compound according to claim 1, wherein R.sup.3 is
CH.sub.3.
14. The compound according to claim 1, which is ##STR00123## or a
pharmaceutically acceptable salt or ester thereof.
15. The compound according to claim 1, which is ##STR00124## or a
pharmaceutically acceptable salt thereof.
16. The compound according to claim 1, which is ##STR00125## or a
pharmaceutically acceptable salt or ester thereof.
17. The compound according to claim 1, which is ##STR00126## or a
pharmaceutically acceptable salt or ester thereof.
18. A pharmaceutical composition comprising a therapeutically
effective amount of a compound according to claim 1 or a
pharmaceutically acceptable salt thereof, and a pharmaceutically
acceptable carrier.
.[.19. A compound, which is: ##STR00127## or a pharmaceutically
acceptable salt thereof..].
Description
FIELD OF THE INVENTION
The present invention is directed to novel thyroid receptor
ligands, particularly to pyridazinone analogs. The invention is
also directed to methods of preparing such compounds which are
useful for treating metabolic diseases such as obesity,
hyperlipidemia, hypercholesterolemia and diabetes and may be useful
for other disorders and diseases such as NASH (nonalcoholic
steatohepatitis), liver steatosis, atherosclerosis, cardiovascular
diseases, hypothyroidism, thyroid cancer and related disorders and
diseases.
All documents cited or relied upon below are expressly incorporated
herein by reference.
BACKGROUND OF THE INVENTION
Thyroid hormones are critical for normal growth and development and
for maintaining metabolic homeostasis (Paul M. Yen Physiological
reviews, Vol. 81(3): pp. 1097-1126 (2001)). Circulating levels of
thyroid hormones are tightly regulated by feedback mechanisms in
the hypothalamus/pituitary/thyroid (HPT) axis. Thyroid dysfunction
leading to hypothyroidism or hyperthyroidism clearly demonstrates
that thyroid hormones exert profound effects on cardiac function,
body weight, metabolism, metabolic rate, body temperature,
cholesterol, bone, muscle and behavior.
Thyroid hormone is produced by the thyroid gland and secreted into
circulation as two distinct forms, 3,5,3',5'-tetra-iodo-L-thyronine
(T4) and 3,5,3'-tri-iodo-L-thyronine (T3). While T4 is the
predominant form secreted by the thyroid gland, T3 is the more
biologically active form. T4 is converted to T3 by tissue specific
deiodinases in all tissues but predominantly in the liver and
kidney. The biological activity of thyroid hormones is mediated by
thyroid hormone receptors (TRs) (M. A. Lazar Endocrine Reviews,
Vol. 14: pp. 348-399 (1993)) TRs belong to the superfamily known as
nuclear receptors. TRs form heterodimers with the retinoid receptor
that act as ligand-inducible transcription factors. TRs have a
ligand binding domain, a DNA binding domain, and an amino terminal
domain, and regulate gene expression through interactions with DNA
response elements and with various nuclear co-activators and
co-repressors. The thyroid hormone receptors are derived from two
separate genes, .alpha. and .beta.. These distinct gene products
produce multiple forms of their respective receptors through
differential RNA processing. The major thyroid receptor isoforms
are .alpha.1, .alpha.2, .beta.1 and .beta.2. Thyroid hormone
receptors .alpha.1, .beta.1 and .beta.2 bind thyroid hormone. It
has been shown that the thyroid hormone receptor subtypes can
differ in their contribution to particular biological responses.
Recent studies suggest that TR.beta.1 plays an important role in
regulating TRH (thyrotropin releasing hormone) and on regulating
thyroid hormone actions in the liver. TR.beta.2 plays an important
role in the regulation of TSH (thyroid stimulating hormone) (Abel
et. al. J. Clin. Invest., Vol 104: pp. 291-300 (1999)). TR.beta.1
plays an important role in regulating heart rate (B. Gloss et. al.
Endocrinology, Vol. 142: pp. 544-550 (2001); C. Johansson et. al.
Am. J. Physiol., Vol. 275: pp. R640-R646 (1998)).
Some of the effects of thyroid hormones may be therapeutically
beneficial if adverse effects can be minimized or eliminated (Paul
M. Yen Physiological Reviews, Vol. 81(3): pp. 1097-1126 (2001);
Paul Webb Expert Opin. Investig. Drugs, Vol. 13(5): pp. 489-500
(2004)). For example, thyroid hormones increase metabolic rate,
oxygen consumption and heat production and thereby reduce body
weight. Reducing body weight will have a beneficial effect in obese
patients by ameliorating the co-morbidities associated with
obesity, and may also have a beneficial effect on glycemic control
in obese patients with Type 2 diabetes.
Another therapeutically beneficial effect of thyroid hormone is the
lowering of serum low density lipoprotein (LDL) (Eugene Morkin et.
al. Journal of Molecular and Cellular Cardiology, Vol. 37: pp.
1137-1146 (2004)). It has been found that hyperthyroidism is
associated with low total serum cholesterol, which is attributed to
thyroid hormone increasing hepatic LDL receptor expression and
stimulating the metabolism of cholesterol to bile acids (J. J.
Abrams et. al. J. Lipid Res., Vol. 22: pp 323-38 (1981)).
Hypothyroidism, in turn, has been associated with
hypercholesterolemia and thyroid hormone replacement therapy is
known to lower total cholesterol (M. Aviram et. al. Clin. Biochem.,
Vol. 15: pp. 62-66 (1982); J. J. Abrams et. al. J. Lipid Res., Vol.
22: pp. 323-38 (1981)). Thyroid hormone has been shown in animal
models to have the beneficial effect of increasing HDL cholesterol
and improving the ratio LDL to HDL by increasing the expression of
apo A-1, one of the major apolipoproteins of HDL (Gene C. Ness et.
al. Biochemical Pharmacology, Vol. 56: pp. 121-129 (1998); G. J.
Grover et. al. Endocrinology, Vol. 145: pp. 1656-1661 (2004); G. J.
Grover et. al. Proc. Natl. Acad. Sci. USA, Vol. 100: pp.
10067-10072 (2003)). Through its effects on LDL and HDL
cholesterol, it is possible that thyroid hormones may also lower
the risk of atherosclerosis and other cardiovascular diseases. The
incidence of atherosclerotic vascular disease is directly related
to the level of LDL cholesterol. Additionally, there is evidence
that thyroid hormones lower Lipoprotein (a), an important risk
factor which is elevated in patients with atherosclerosis (Paul
Webb Expert Opin. Investig. Drugs, Vol. 13(5): pp. 489-500 (2004);
de Bruin et. al. J. Clin. Endo. Metab., Vol. 76: pp. 121-126
(1993)).
With the incidence of obesity and its co-morbidities, diabetes,
metabolic syndrome, and atherosclerotic vascular disease rising at
epidemic rates, the utility of compounds capable of treating these
diseases would be highly desirable. To date, the therapeutic uses
of the naturally occurring thyroid hormone have been limited by the
adverse side effects associated with hyperthyroidism, especially
cardiovascular toxicity.
Therefore, efforts have been made to synthesize thyroid hormone
analogs which exhibit increased thyroid hormone receptor beta
selectivity and/or tissue selective action. Such thyroid hormone
mimetics may yield desirable reductions in body weight, lipids,
cholesterol, and lipoproteins, with reduced impact on
cardiovascular function or normal function of the
hypothalamus/pituitary/thyroid axis (A. H. Underwood et al. Nature,
Vol. 324: pp. 425-429 (1986), G. J. Grover et. al. PNAS, Vol. 100:
pp. 10067-10072 (2003); G. J. Grover Endocrinology, Vol. 145: pp
1656-1661 (2004); Yi-lin Li et. al. PCT Int. Appl. WO 9900353
(1999); Thomas S. Scanlan et. al. PCT Int. Appl. WO 9857919 (1998);
Keith A. Walker et. al. U.S. Pat. No. 5,284,971 (1994); Mark D.
Erion et. al. PCT Int. Appl. WO 2005051298 (2005); Malm Johan
Current Pharmaceutical Design, Vol. 10(28): pp. 3525-3532 (2004);
Expert Opin. Ther. Patents, Vol. 14: pp 1169-1183 (2004); Thomas S.
Scalan Current Opinion in Drug Discovery & Development, Vol. 4
(5): pp. 614-622 (2001); Paul Webb Expert Opinion on
Investigational Drugs, Vol. 13 (5): pp 489-500 (2004)).
The development of thyroid hormone analogs which avoid the
undesirable effects of hyperthyroidism and hypothyroidism while
maintaining the beneficial effects of thyroid hormones would open
new avenues of treatment for patients with metabolic diseases such
as obesity, hyperlipidemia, hypercholesterolemia, diabetes and
other disorders and diseases such as liver steatosis and NASH,
atherosclerosis, cardiovascular diseases, hypothyroidism, thyroid
cancer, thyroid diseases, and related disorders and diseases.
Pyridazinone compounds that are structurally different from the
compounds of the present invention have been previously disclosed
(Teruomi et. al. Agricultural and Biological Chemistry, Vol.
38(6):1169-76 (1974); P. D. Leeson et. al. J. Med. Chem. Vol. 32:
pp. 320-326 (1989); Eur. Pat. Appl. EP 188351 (1986); Damien John
Dunnington PCT Int. Appl. WO 9702023 (1997); and Eur. Pat. Appl. EP
728482 (1996)).
Against this background there is still a need, therefore, for novel
thyroid hormone mimetics such as, for example, novel pyridazinone
thyroid hormone mimetics, that have the beneficial effects of
thyroid hormone while avoiding the undesirable effects.
SUMMARY OF THE INVENTION
In one embodiment of the present invention, provided is a compound
of the formula (I):
##STR00002## wherein: A is O, CH.sub.2, S, SO or SO.sub.2; X and Y
are each independently selected from the group consisting of Br, Cl
and --CH.sub.3; R.sup.1 is selected from the group consisting of
--(CH.sub.2).sub.nCOOH; --OCH.sub.2COOH; --NHC(.dbd.O)COOH;
--NHCH.sub.2COOH;
##STR00003## Z is H, or --C.dbd.CN; R.sup.2 is lower alkyl having
from 1 to 4 C atoms; R.sup.3 is H or lower alkyl; n is 1 or 2 p is
1 or 2; or a pharmaceutically acceptable salt or ester thereof.
In another embodiment of the present invention, provided is a
pharmaceutical composition comprising a therapeutically effective
amount of a compound according to formula (I) or a pharmaceutically
acceptable salt or ester thereof, and a pharmaceutically acceptable
carrier.
In a further embodiment of the present invention, provided is a
method for treating a metabolic disorder in a patient in need of
such treatment, comprising administering to the patient a
therapeutically effective amount of a compound according to formula
(I) or a pharmaceutically acceptable salt or ester thereof.
DETAILED DESCRIPTION OF THE INVENTION
It is to be understood that the terminology employed herein is for
the purpose of describing particular embodiments, and is not
intended to be limiting. Further, although any methods, devices and
materials similar or equivalent to those described herein can be
used in the practice or testing of the invention, the preferred
methods, devices and materials are now described.
As used herein, the term "alkyl" means, for example, a branched or
unbranched, cyclic or acyclic, saturated or unsaturated (e.g.
alkenyl or alkynyl)hydrocarbyl group which may be substituted or
unsubstituted. Where cyclic, the alkyl group is preferably C.sub.3
to C.sub.12, more preferably C.sub.4 to C.sub.10, more preferably
C.sub.4 to C.sub.7. Where acyclic, the alkyl group is preferably
C.sub.1 to C.sub.10, more preferably C.sub.1 to C.sub.6, more
preferably methyl, ethyl, propyl(n-propyl or isopropyl),
butyl(n-butyl, isobutyl or tertiary-butyl) or pentyl (including
n-pentyl and isopentyl), more preferably methyl. It will be
appreciated therefore that the term "alkyl" as used herein includes
alkyl (branched or unbranched), substituted alkyl (branched or
unbranched), alkenyl (branched or unbranched), substituted alkenyl
(branched or unbranched), alkynyl (branched or unbranched),
substituted alkynyl (branched or unbranched), cycloalkyl,
substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl,
cycloalkynyl and substituted cycloalkynyl.
As used herein, the term "lower alkyl" means, for example, a
branched or unbranched, cyclic or acyclic, saturated or unsaturated
(e.g. alkenyl or alkynyl)hydrocarbyl group wherein said cyclic
lower alkyl group is C.sub.5, C.sub.6 or C.sub.7, and wherein said
acyclic lower alkyl group is C.sub.1, C.sub.2, C.sub.3, C.sub.4,
C.sub.5, or C.sub.6, preferably from 1 to 4 carbon atoms. Typical
lower alkyl groups include methyl, ethyl, propyl(n-propyl or
isopropyl), butyl(n-butyl, isobutyl or tertiary-butyl), pentyl and
hexyl. It will be appreciated therefore that the term "lower alkyl"
as used herein includes, for example, lower alkyl (branched or
unbranched), lower alkenyl (branched or unbranched), lower alkynyl
(branched or unbranched), cycloloweralkyl, cycloloweralkenyl and
cycloloweralkynyl. When attached to another functional group, lower
alkyl as used herein may be divalent, e.g., -lower alkyl-COOH.
At position R.sup.2 of formula (I), lower alkyl has from 1 to 4
carbon atoms. A preferred R.sup.2 is lower alkyl having 3 carbon
atoms. More preferred is isopropyl.
As used herein, the term "aryl" means, for example, a substituted
or unsubstituted carbocyclic aromatic group, such as phenyl or
naphthyl, or a substituted or unsubstituted heteroaromatic group
containing one or more, preferably one, heteroatom, such as
pyridyl, pyrrolyl, furanyl, thienyl, thiazolyl, isothiazolyl,
oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl pyrazolyl,
imidazolyl, triazolyl, pyrimidinyl pyridazinyl, pyrazinyl,
triazinyl, indolyl, indazolyl, quinolyl, quinazolyl,
benzimidazolyl, benzothiazolyl, benzisoxazolyl and
benzisothiazolyl.
The alkyl and aryl groups may be substituted or unsubstituted.
Where substituted, there will generally be, for example, 1 to 3
substituents present, preferably 2 substituents. Substituents may
include, for example: carbon-containing groups such as alkyl, aryl,
arylalkyl (e.g. substituted and unsubstituted phenyl, substituted
and unsubstituted benzyl); halogen atoms and halogen-containing
groups such as haloalkyl (e.g. trifluoromethyl); oxygen-containing
groups such as alcohols (e.g. hydroxyl, hydroxyalkyl,
aryl(hydroxyl)alkyl), ethers (e.g. alkoxy, aryloxy, alkoxyalkyl,
aryloxyalkyl), aldehydes (e.g. carboxaldehyde), ketones (e.g.
alkylcarbonyl, alkylcarbonylalkyl, arylcarbonyl, arylalkylcarbonyl,
arycarbonylalkyl), acids (e.g. carboxy, carboxyalkyl), acid
derivatives such as esters (e.g. alkoxycarbonyl,
alkoxycarbonylalkyl, alkylcarbonyloxy, alkylcarbonyloxyalkyl),
amides (e.g. aminocarbonyl, mono- or di-alkylaminocarbonyl,
aminocarbonylalkyl, mono- or di-alkylaminocarbonylalkyl,
arylaminocarbonyl), carbamates (e.g. alkoxycarbonylamino,
arloxycarbonylamino, aminocarbonyloxy, mono- or
di-alkylaminocarbonyloxy, arylminocarbonloxy) and ureas (e.g. mono-
or di-alkylaminocarbonylamino or arylaminocarbonylamino);
nitrogen-containing groups such as amines (e.g. amino, mono- or
di-alkylamino, aminoalkyl, mono- or di-alkylaminoalkyl), azides,
nitriles (e.g. cyano, cyanoalkyl), nitro; sulfur-containing groups
such as thiols, thioethers, sulfoxides and sulfones (e.g.
alkylthio, alkylsulfinyl, alkylsulfonyl, alkylthioalkyl,
alkylsulfinylalkyl, alkylsulfonylalkyl, arylthio, arysulfinyl,
arysulfonyl, arythioalkyl, arylsulfinylalkyl, arylsulfonylalkyl);
and heterocyclic groups containing one or more, preferably one,
heteroatom, (e.g. thienyl, furanyl, pyrrolyl, imidazolyl,
pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, oxadiazolyl,
thiadiazolyl, aziridinyl, azetidinyl, pyrrolidinyl, pyrrolinyl,
imidazolidinyl, imidazolinyl, pyrazolidinyl, tetrahydrofuranyl,
pyranyl, pyronyl, pyridyl, pyrazinyl, pyridazinyl, piperidyl,
hexahydroazepinyl, piperazinyl, morpholinyl, thianaphthyl,
benzofuranyl, isobenzofuranyl, indolyl, oxyindolyl, isoindolyl,
indazolyl, indolinyl, 7-azaindolyl, benzopyranyl, coumarinyl,
isocoumarinyl, quinolinyl, isoquinolinyl, naphthridinyl,
cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl,
quinoxalinyl, chromenyl, chromanyl, isochromanyl, phthalazinyl and
carbolinyl).
The lower alkyl groups may be substituted or unsubstituted,
preferably unsubstituted. Where substituted, there will generally
be, for example, 1 to 3 substitutents present, preferably 2
substituents.
As used herein, the term "alkoxy" means, for example, alkyl-O-- and
"alkoyl" means, for example, alkyl-CO--. Alkoxy substituent groups
or alkoxy-containing substituent groups may be substituted by, for
example, one or more alkyl groups.
As used herein, the term "halogen" means, for example, a fluorine,
chlorine, bromine or iodine group, preferably a chlorine or bromine
group, and more preferably a chlorine group.
"Pharmaceutically acceptable," such as pharmaceutically acceptable
carrier, excipient, etc., means pharmacologically acceptable and
substantially non-toxic to the subject to whom the particular
compound is administered.
"Pharmaceutically acceptable salt" refers to conventional
acid-addition salts or base-addition salts that retain the
biological effectiveness and properties of the compounds of formula
I and are formed from suitable organic or inorganic acids or
organic or inorganic bases. Sample acid-addition salts include
those derived from inorganic acids such as hydrochloric acid,
hydrobromic acid, hydroiodic acid, sulfuric acid, sulfamic acid,
phosphoric acid and nitric acid, and those derived from organic
acids such as p-toluenesulfonic acid, salicylic acid,
methanesulfonic acid, oxalic acid, succinic acid, citric acid,
malic acid, lactic acid, fumaric acid, and the like. Sample
base-addition salts include those derived from ammonium, potassium,
sodium and, quaternary ammonium hydroxides, such as for example,
tetramethylammonium hydroxide. The chemical modification of a
pharmaceutical compound (i.e. drug) into a salt is a well known
technique which is used in attempting to improve properties
involving physical or chemical stability, e.g., hygroscopicity,
flowability or solubility of compounds. See, e.g., H. Ansel et.
al., Pharmaceutical Dosage Forms and Drug Delivery Systems (6th Ed.
1995) at pp. 196 and 1456-1457.
"Pharmaceutically acceptable ester" refers to a conventionally
esterified compound of formula I having a carboxyl group, which
esters retain the biological effectiveness and properties of the
compounds of formula I and are cleaved in vivo (in the organism) to
the corresponding active carboxylic acid. In the present invention,
esters may be present, for example, where R.sup.1 is
--(CH.sub.2).sub.nCOOH, --OCH.sub.2COOH, --NHC(.dbd.O)COOH, or
--NHCH.sub.2COOH. Examples of ester groups which are cleaved (in
this case hydrolyzed) in vivo to the corresponding carboxylic acids
are those in which the hydrogen is replaced with--lower alkyl which
is optionally substituted, e.g., with heterocycle, cycloalkyl, etc.
Examples of substituted lower alkyl esters are those in
which--lower alkyl is substituted with pyrrolidine, piperidine,
morpholine, N-methylpiperazine, etc. The group which is cleaved in
vivo may be, for example, ethyl, morpholino ethyl, and diethylamino
ethyl. In connection with the present invention, --CONH.sub.2 is
also considered an ester, as the --NH.sub.2 may be cleaved in vivo
and replaced with a hydroxy group, to form the corresponding
carboxylic acid.
Further information concerning examples of and the use of esters
for the delivery of pharmaceutical compounds is available in Design
of Prodrugs. Bundgaard H. ed. (Elsevier, 1985). See also, H. Ansel
et. al., Pharmaceutical Dosage Forms and Drug Delivery Systems (6th
Ed. 1995) at pp. 108-109; Krogsgaard-Larsen, et. al., Textbook of
Drug Design and Development (2d Ed. 1996) at pp. 152-191.
Compounds of the present invention can be prepared beginning with
commercially available starting materials and utilizing general
synthetic techniques and procedures known to those skilled in the
art. Chromatography supplies and equipment may be purchased from
such companies as for example AnaLogix, Inc, Burlington, Wis.;
Analytical Sales and Services, Inc., Pompton Plains, N.J.; Teledyne
Isco, Lincoln, Nebr.; VWR International, Bridgeport, N.J.; and
Rainin Instrument Company, Woburn, Mass. Chemicals may be purchased
from companies such as for example Aldrich, Argonaut Technologies,
VWR and Lancaster. Outlined below are reaction schemes suitable for
preparing such compounds. Further exemplification is found in the
specific Examples detailed below.
##STR00004##
##STR00005##
##STR00006##
Compounds 10a and 10b were synthesized following a series of
reactions outlined in schemes 1-3. The starting material for the
synthesis of 10a was 4-hydroxy-2,6-dimethyl phenyl acetic acid
methyl ester, 6a, which was synthesized in five steps from
commercially available 2,6-dimethyl phenol as shown in scheme 1
using the procedure described by Gardner, P. D., et. al in J. Amer.
Chem. Soc., 1959, 81, 3364. 3,6-dichloropyridazine, was alkylated
to give compound 7 (scheme 2) (see for example, Organic
Preparations and Procedures International, 1988, 20(1-2), 117-121).
Compound 6a was condensed with compound 7 using a base and a
catalytic amount of an organometallic halide at elevated
temperatures to afford 8a (see for example, Yuhpyng L. C., et. al.
PCT Int. Appl. (1996) WO 9639388). Base hydrolysis of the methyl
ester of 8a was performed via conventional procedures to afford
compound 9a. Conversion of the chloropyridazine 9a to pyridizinone
10a was performed under acidic conditions (see for example, J.
Chem. Soc. Perkin Trans. 1: Org. and Bioorg. Chem., 1988, 12,
3103-3111). Compound 10b was synthesized in the same manner as 10a
starting from compound 6b. Compound 6b was synthesized in an
analogous manner as 6a (scheme 1) starting from compound 1b.
##STR00007##
Compound 14 was synthesized following a series of reactions
outlined in scheme 4. The starting material for the synthesis of 14
was 2-(4-hydroxyphenyl)ethanol which was converted to 11 under
conventional bromination conditions (see for example, J. Amer.
Chem. Soc., 1989, 111(24), 8912-8914). Condensation of compound 7
and compound 11 was accomplished with potassium tert-butoxide in
N,N-dimethyl acetamide at high temperatures to produce compound 12.
Compound 12 was oxidized to produce compound 13 using a similar
procedure as described by Anelli, P. L., et. al, J. Org. Chem.,
1987, 52(12), 2559-2562. The chloropyridazine 13 was converted to
the pyridizinone 14 under conditions as described previously.
##STR00008##
Compound 19 was synthesized following a series of reactions
outlined in scheme 5. The starting material for the synthesis of 19
was methyl 4-hydroxyphenyl acetate which was converted to compound
15 under conventional chlorination conditions (see for example,
Maeda, R., et. al., Chem. Pharm. Bull., 1983, 31(10), 3424-3445).
Compound 15 was reduced to compound 16 using lithium aluminum
hydride in tetrahydrofuran at low temperature and short reaction
times. Compound 16 was converted in three steps to pyridizinone 19
under conditions which were described previously.
##STR00009## ##STR00010##
Compound 23 was synthesized following a series of reactions
outlined in scheme 6. Bromination of 2-(4-hydroxyphenyl)ethanol to
produce compound 20 was performed under conditions previously
described. Compound 20 was condensed with compound 7 using
potassium tert-butoxide in N,N-dimethylacetamide at high
temperatures to produce compound 21. Compound 21 was oxidized to
compound 22 by Jones oxidation (see for example, Bowden, K., et.
al., J. Chem. Soc., 1946, 39-45). The chloropyridazine analog 22
was then converted to the pyridazinone 23 under conditions
previously described. The 3,5-dichlorophenyl analog of 23, the
3,5-dimethylphenyl analog of 23, the 3-chloro-5-methylphenyl analog
of 23, 3-bromo-5-methylphenyl analog of 23, and the
3-bromo-5-chlorophenyl analog of 23 can be synthesized in a similar
manner.
##STR00011##
Compound 26 was synthesized following a series of reactions
outlined in scheme 7. The starting material,
4-amino-2,6-dichlorophenol, was condensed with compound 7 to
produce compound 24 by the procedure described previously. The
chloropyridazine 24 was converted to the pyridazinone by the
procedure described previously. Hydrolysis of the resulting
acetamide was performed under standard aqueous basic conditions to
produce compound 25. Compound 25 was then converted to compound 26
via reductive amination using cyanoborohydride on resin
(MP-cynanoborohydride, commercially available from Argonaut
Technologies) and glyoxylic acid. The 3,5-dibromophenyl analog of
26, the 3,5-dimethylphenyl analog of 26, the
3-chloro-5-methylphenyl analog of 26, 3-bromo-5-methylphenyl analog
of 26, and the 3-bromo-5-chlorophenyl analog of 26 can be
synthesized in a similar manner.
##STR00012##
Compound 29 was synthesized following a series of reactions
outlined in scheme 8. Compound 24 was converted to compound 27 via
acylation of the amine with methyl oxalyl chloride (see for
example, Sellstedt. J. H., et. al, J. Med. Chem., 1975, 18(9),
926-933). Compound 27 was converted to compound 28 by hydrolysis of
the .alpha.-keto ester to the .alpha.-keto acid using standard
conditions (see for example, Minisci, F., et. al., J. Org. Chem.,
1995, 60, 5430-5433). The chloropyridazine 28 was converted to the
pyridazinone 29 by the procedure described previously. The
3,5-dibromophenyl analog of 29, the 3,5-dimethylphenyl analog of
29, the 3-chloro-5-methylphenyl analog of 29,
3-bromo-5-methylphenyl analog of 29, and the 3-bromo-5-chlorophenyl
analog of 29 can be synthesized in a similar manner.
##STR00013##
Compound 31 was synthesized following a series of reactions
outlined in scheme 9. Compound 25 was converted to the
cyanoacetylurethane 30 following the procedure of Carroll, R. D.,
et. al., J. Med. Chem., 1983, 26, 96-100. Cyclization of the
urethane of compound 30 to produce cyano-azauracil 31 was carried
out following the procedure of Carroll, R. D., et. al., J. Med.
Chem., 1983, 26, 96-100. The 3,5-dibromophenyl analog of 31, the
3,5-dimethylphenyl analog of 31, the 3-chloro-5-methylphenyl analog
of 31, 3-bromo-5-methylphenyl analog of 31, and the
3-bromo-5-chlorophenyl analog of 31 can be synthesized in a similar
manner.
##STR00014##
Compound 33 was synthesized following a series of reactions
outlined in scheme 10. Compound 31 was converted to compound 32 by
hydrolyzing the cyano group to a carboxylic acid via the procedure
described by Carroll, R. D., et. al., J. Med. Chem., 1983, 26,
96-100. Compound 32 was then converted to compound 33 by
decarboxylation following the procedure described by Carroll, R.
D., et. al., J. Med. Chem., 1983, 26, 96-100. The 3,5-dibromophenyl
analog of 33, the 3,5-dimethylphenyl analog of 33, the
3-chloro-5-methylphenyl analog of 33, 3-bromo-5-methylphenyl analog
of 33, and the 3-bromo-5-chlorophenyl analog of 33 can be
synthesized in a similar manner.
##STR00015## ##STR00016##
Compound 37 was synthesized following a series of reactions
outlined in scheme 11. The chloropyridazine 17 was converted to the
pyridazinone 34 by the procedure described previously. The
pyridazinone 34 was alkylated using base and methyl iodide to
produce compound 35 following the procedure similar to that
described in J. Med. Chem., 1989, 32(10), 2381-2388. The acetate 35
was hydrolyzed to the alcohol 36 under standard basic conditions
(see for example, Hauser, C. R., et. al., J. Amer. Chem. Soc.,
1945, 67, 409-412). Compound 36 was then oxidized to compound 37 by
Jones oxidation. The 3,5-dibromophenyl analog of 37, the
3,5-dimethylphenyl analog of 37, the 3-chloro-5-methylphenyl analog
of 37, 3-bromo-5-methylphenyl analog of 37, and the
3-bromo-5-chlorophenyl analog of 37 can be synthesized in a similar
manner.
##STR00017## ##STR00018##
Compound 42 was synthesized following a series of reactions
outlined in scheme 12. Compound 11 was converted to compound 42 in
five steps using previously described conditions. The
3,5-dichlorophenyl analog of 42, the 3,5-dimethylphenyl analog of
42, the 3-chloro-5-methylphenyl analog of 42,
3-bromo-5-methylphenyl analog of 42, and the 3-bromo-5-chlorophenyl
analog of 42 can be synthesized in a similar manner.
##STR00019## ##STR00020##
Compound 48 was synthesized following a series of reactions
outlined in scheme 13. The starting material,
1,2,3-trichloro-5-nitrobenzene, was converted to compound 43 by
selectively displacing the chloro at the 2-position with the anion
formed from tert-butyl cyanoacetate (see for example, Salturo, F.,
et. al., PCT WO 00/17204). The nitro group of compound 43 was
reduced to an amine using standard conditions. The tert-butyl ester
of 43 was hydrolyzed and decarboxylated to give compound 44 (see
for example, Salturo, F., et. al., PCT WO 00/17204). The anion of
compound 44 was condensed with compound 7 to give compound 45 using
a similar procedure described by Salturo, F., et. al., PCT WO
00/17204. The chloropyridazine 45 was converted to the pyridazinone
46 under acidic conditions at high temperatures. Under these
conditions, the cyano group was hydrolyzed to the carboxylic acid
and then decarboxylated using the procedure described by Carroll,
R. D., et. al., J. Med. Chem., 1983, 26, 96-100. Compound 46 was
converted to compound 48 in two steps which were previously
described. The 3,5-dibromophenyl analog of 48, the
3,5-dimethylphenyl analog of 48, the 3-chloro-5-methylphenyl analog
of 48, 3-bromo-5-methylphenyl analog of 48, and the
3-bromo-5-chlorophenyl analog of 48 can be synthesized in a similar
manner.
##STR00021##
Compound 50 was synthesized following a series of reactions
outlined in scheme 14. Compound 50 was obtained from compound 48 in
two steps that were described previously. The 3,5-dibromophenyl
analog of 50, the 3,5-dimethylphenyl analog of 50, the
3-chloro-5-methylphenyl analog of 50, 3-bromo-5-methylphenyl analog
of 50, and the 3-bromo-5-chlorophenyl analog of 50 can be
synthesized in a similar manner.
##STR00022## ##STR00023##
Compound 56 was synthesized following a series of reactions
outlined in scheme 15. The amine of compound 46 was converted to
the bromide of compound 51 using standard conditions (see for
example, Doyle, M. P., et. al., J. Org. Chem., 1977, 42(14),
2426-2431). The bromide 51 was converted to the methyl ester 52 by
palladium catalyzed carbonylation in methanol (see for example,
Takatori, K., et. al., Tetrahedron, 1998, 54, 15861-15869).
Compound 52 was reduced to compound 53 by standard reduction
conditions, treatment with diisobutylaluminum hydride in
tetrahydrofuran (see for example, Yoon, N. M., et. al., J. Org.
Chem., 1985, 50, 2443-2450). The alcohol 53 was converted to
bromide 54 using standard conditions (see for example, Lan, Aj. J.
Y, et. al., J. Amer. Chem. Soc., 1987, 109, 2738-2745). The bromide
54 was displaced with sodium cyanide to produce nitrile 55 using
the procedure described by Law, H., et. al. J. Med. Chem., 1998,
41, 2243-2251. The nitrile 55 was hydrolyzed to acid 56 by a
conventional procedure to hydrolyze a nitrile to a carboxylic acid
under aqueous acidic conditions (see for example, Wenner, O., Org.
Synth.; Coll. Vol. IV, 1963, 760). The 3,5-dimethylphenyl analog of
56, the 3-chloro-5-methylphenyl analog of 56,
3-bromo-5-methylphenyl analog of 56, and the 3-bromo-5-chlorophenyl
analog of 56 can be synthesized in a similar manner.
##STR00024## ##STR00025##
Compound 64 was synthesized following a series of reactions
outlined in scheme 16. The starting material, methyl
3,5-dibromo-4-methylbenzoate, was brominated to produce compound 57
under standard bromination conditions (see for example, Buu-Hoy,
P., et. al., J. Org. Chem., 1953, 18, 649-652). Compound 57 was
converted to compound 59 in two steps that were described
previously. The alcohol 59 was converted to ether 60 by a
conventional procedure to convert an alcohol to a tetrahydropyranyl
ether (see for example, Miyashita, M., et. al., J. Org. Chem.,
1977, 42, 3882-3774). Condensation of compound 7 and compound 60
was accomplished with sodium hydride in N,N-dimethylformamide at
high temperatures to produce compound 61 using a similar procedure
as described by Salturo, F., et. al., PCT WO 00/17204. Compound 61
was treated under acidic conditions at elevated temperatures. These
reaction conditions resulted in the conversion of the
chloropyridazine to the pyridzinone, hydrolysis of the nitrile
moiety to the carboxylic acid followed by decarboxylation of the
acid, and conversion of the tetrahydropyranyl protected alcohol to
the benzyl chloride of compound 62. The chloride of compound 62 was
displaced with sodium cyanide to produce nitrile 63 using a
procedure similar to that of Law, H., et. al. J. Med. Chem., 1998,
41, 2243-2251. Compound 64 was synthesized from compound 63
following a procedure that was described previously.
##STR00026## ##STR00027##
Compound 69 was synthesized following a series of reactions
outlined in scheme 17. The starting material,
4-amino-2,6-dichloro-phenol, was condensed with compound 7 using
potassium tert-butoxide in N,N-dimethylacetamide at elevated
temperature. The resulting intermediate was then treated with
phthalic anhydride at elevated temperatures to form the
phthalimide. The resulting phthalimide was heated in glacial acetic
acid with sodium acetate to produce compound 65. The pyridizinone
nitrogen of compound 65 was methylated under the conditions similar
to those found in Sotelo, E., et. al., Synth. Commun., 2002,
32(11), 1675-1680 to form compound 66. The phthalimide protecting
group of compound 66 was removed using butylamine in methanol at
elevated temperatures to afford compound 67. Compound 67 was then
converted to compound 68 using previously described methods.
Compound 68 was converted to compound 69 using potassium acetate in
N,N-dimethylacetamide at elevated temperatures. The
3,5-dibromophenyl analog of 69, the 3,5-dimethylphenyl analog of
69, the 3-chloro-5-methylphenyl analog of 69,
3-bromo-5-methylphenyl analog of 69, and the 3-bromo-5-chlorophenyl
analog of 69 can be synthesized in a similar manner.
##STR00028## ##STR00029##
Compound 76 was synthesized following a series of reactions
outlined in scheme 18. The alcohol 16 was protected with a
tert-butyldiphenylsilyl ether to produce compound 70 following a
standard procedure as described in Chaudhary, S. K., et. al., Tet.
Lett., 1979, 20(2), 99-102. The phenol 70 was converted to the
thiophenol 73 using a three step procedure as described by D. M.
Springer, et. al., Bioorg Med. Chem., 2003, 11, 265-279.
Condensation of compound 7 and compound 73 was accomplished with
potassium carbonate in dimethyl sulfoxide at high temperatures to
produce compound 74. Compound 74 was converted to the pyridizinone
acid 76 in two steps that have been previously described. The
3,5-dibromophenyl analog of 76, the 3,5-dimethylphenyl analog of
76, the 3-chloro-5-methylphenyl analog of 76,
3-bromo-5-methylphenyl analog of 76, and the 3-bromo-5-chlorophenyl
analog of 76 can be synthesized in a similar manner.
##STR00030##
Compounds 77 and 78 were synthesized through a series of reactions
outlined in scheme 19. The thioether 76 was oxidized using formic
acid and hydrogen peroxide to form a separable mixture of the
sulfoxide 77 and the sulfonyl 78 (see for example, Cragoe, E. J.,
et. al. U.S. Pat. No. 4,582,842 Apr. 15, 1986). The
3,5-dibromophenyl analog of 77 and 78, the 3,5-dimethylphenyl
analog of 77 and 78, the 3-chloro-5-methylphenyl analog of 77 and
78, 3-bromo-5-methylphenyl analog of 77 and 78, and the
3-bromo-5-chlorophenyl analog of 77 and 78 can be synthesized in a
similar manner.
##STR00031## ##STR00032## ##STR00033##
Compound 88 was synthesized following a series of reactions
outlined in scheme 20. The starting material,
2,6-dichloro-4-nitrophenol, was converted to compound 79 under
conditions that have been previously described. Compound 79 was
converted to compound 80 under conditions that have been previously
described. The nitro group of compound 80 was reduced to the amine
81 using standard reaction conditions, iron powder in an acidic
medium (see for example, Org. Syn. Coll. Vol. 2, 1943, 471).
Compound 81 was then converted to compound 88 under conditions that
have been previously described. The 3,5-dibromophenyl analog of 88,
the 3,5-dimethylphenyl analog of 88, the 3-chloro-5-methylphenyl
analog of 88, 3-bromo-5-methylphenyl analog of 88, and the
3-bromo-5-chlorophenyl analog of 88 can be synthesized in a similar
manner.
##STR00034##
Compound 91 was synthesized following a series of reactions
outlined in scheme 21. Compound 87 was converted to compound 91
under conditions that have been previously described. The
3,5-dibromophenyl analog of 91, the 3,5-dimethylphenyl analog of
91, the 3-chloro-5-methylphenyl analog of 91,
3-bromo-5-methylphenyl analog of 91, and the 3-bromo-5-chlorophenyl
analog of 91 can be synthesized in a similar manner.
##STR00035## ##STR00036##
Compound 96 was synthesized following a series of reactions
outlined in scheme 22. The amine 46 was treated with phthalic
anhydride under acidic conditions at elevated temperatures to form
the phthalimide 92 (see for example, Vera, L. M. S., et. al.,
Farmaco, 2003, 58(12), 1283-1288). The pyridizinone nitrogen of
compound 92 was methylated under the conditions outlined in Sotelo,
E., et. al., Synth. Commun., 2002, 32(11), 1675-1680. The
phthalimide protecting group of compound 93 was removed under
acidic conditions at elevated temperatures to afford compound 94.
Compound 94 was converted to compound 96 under conditions that have
been previously described. The 3,5-dibromophenyl analog of 96, the
3,5-dimethylphenyl analog of 96, the 3-chloro-5-methylphenyl analog
of 96, 3-bromo-5-methylphenyl analog of 96, and the
3-bromo-5-chlorophenyl analog of 96 can be synthesized in a similar
manner.
Proposed Syntheses
##STR00037## ##STR00038##
The alcohol 17 may be converted to bromide 97 using standard
conditions (see for example, Lan, Aj. J. Y, et. al., J. Amer. Chem.
Soc., 1987, 109, 2738-2745). The bromide of compound 97 can be
displaced with sodium cyanide to produce nitrile 98 using a
procedure similar to that of Law, H., et. al. J. Med. Chem., 1998,
41, 2243-2251. Nitriles such as 98 can be converted to tetrazoles
like 98 using known procedures, for example by treatment of 98 with
ammonium chloride and sodium azide at elevated temperature (see for
example, Synthesis, 1998, 6, 910-914). Conversion of the
chloropyridazine 99 to pyridizinone 100 may be accomplished under
acidic conditions (see for example, J. Chem. Soc. Perkin Trans. 1:
Org. and Bioorg. Chem., 1988, 12, 3103-3111). The 3,5-dibromophenyl
analog of 100, the 3,5-dimethylphenyl analog of 100, the
3-chloro-5-methylphenyl analog of 100, 3-bromo-5-methylphenyl
analog of 100, and the 3-bromo-5-chlorophenyl analog of 100 can be
synthesized in a similar manner.
##STR00039## ##STR00040##
The ester of methyl 3,5-dichloro-4-hydroxybenzoate can be reduced
to afford alcohol 101 using a reducing agent such as lithium
aluminum hydride at low temperature (see for example, J. Org.
Chem., 1998, 63, 5658-5661). Compound 101 may then be condensed
with compound 7 using a base at elevated temperatures to afford 102
(see for example, Yuhpyng L. C., et. al. PCT Int. Appl. (1996) WO
9639388). The alcohol 102 can be converted to compound 106 using
conditions previously described. The 3,5-dibromophenyl analog of
106, the 3,5-dimethylphenyl analog of 106, the
3-chloro-5-methylphenyl analog of 106, 3-bromo-5-methylphenyl
analog of 106, and the 3-bromo-5-chlorophenyl analog of 106 can be
synthesized in a similar manner.
##STR00041## ##STR00042##
The bromide 54 may be homologated to ester 107 using conditions as
illustrated in Can. J. Chem. 2001, 79(5-6), 752-759. Esters such as
107 can be converted to alcohols like 108 using known procedures,
for example by treatment of 107 with diisobutylammonium hydride in
tetrahydrofuran (see for example, Yoon, N. M., et. al., J. Org.
Chem., 1985, 50, 2443-2450). Conversion of the alcohol 108 to
tetrazole 111 may be accomplished under conditions as previously
described. The 3,5-dibromophenyl analog of 111, the
3,5-dimethylphenyl analog of 111, the 3-chloro-5-methylphenyl
analog of 111, 3-bromo-5-methylphenyl analog of 111, and the
3-bromo-5-chlorophenyl analog of 111 can be synthesized in a
similar manner.
##STR00043##
Conversion of the nitrile 55 to tetrazole 112 may be accomplished
using conditions as previously described. The 3,5-dibromophenyl
analog of 112, the 3,5-dimethylphenyl analog of 112, the
3-chloro-5-methylphenyl analog of 112, 3-bromo-5-methylphenyl
analog of 112, and the 3-bromo-5-chlorophenyl analog of 112 can be
synthesized in a similar manner.
##STR00044## ##STR00045##
Conversion of compound 74 to compound 116 may be accomplished using
conditions as previously described. The 3,5-dibromophenyl analog of
116, the 3,5-dimethylphenyl analog of 116, the
3-chloro-5-methylphenyl analog of 116, 3-bromo-5-methylphenyl
analog of 116, and the 3-bromo-5-chlorophenyl analog of 116 can be
synthesized in a similar manner.
##STR00046##
Conversion of compound 116 to compounds 117 and 118 may be
accomplished using conditions as previously described. The
3,5-dibromophenyl analog of 117 and 118, the 3,5-dimethylphenyl
analog of 117 and 118, the 3-chloro-5-methylphenyl analog of 117
and 118, 3-bromo-5-methylphenyl analog of 117 and 118, and the
3-bromo-5-chlorophenyl analog of 117 and 118 can be synthesized in
a similar manner.
##STR00047## ##STR00048##
Conversion of compound 101 to compound 127 may be accomplished
using conditions as previously described. The 3,5-dibromophenyl
analog of 127, the 3,5-dimethylphenyl analog of 127, the
3-chloro-5-methylphenyl analog of 127, 3-bromo-5-methylphenyl
analog of 127, and the 3-bromo-5-chlorophenyl analog of 127 can be
synthesized in a similar manner.
##STR00049##
Conversion of compound 127 to compounds 128 and 129 may be
accomplished using conditions as previously described. The
3,5-dibromophenyl analog of 128 and 129, the 3,5-dimethylphenyl
analog of 128 and 129, the 3-chloro-5-methylphenyl analog of 128
and 129, 3-bromo-5-methylphenyl analog of 128 and 129, and the
3-bromo-5-chlorophenyl analog of 128 and 129 can be synthesized in
a similar manner.
##STR00050## ##STR00051##
The starting material, 2,6-dichloro-benzene-1,4-diol, may be
treated with bromo-acetic acid methyl ester and base at elevated
temperatures to produce compound 130 using a procedure similar to
that of J. Het. Chem., 1994, 31(6), 1439-43. Conversion of compound
130 to compound 131 may be accomplished using conditions as
previously described. Hydrolysis of ester 131 to compound 132 may
be accomplished using standard aqueous basic conditions. Conversion
of compound 132 to compound 133 may be accomplished using
conditions as previously described. The 3,5-dibromophenyl analog of
133, the 3,5-dimethylphenyl analog of 133, the
3-chloro-5-methylphenyl analog of 133, 3-bromo-5-methylphenyl
analog of 133, and the 3-bromo-5-chlorophenyl analog of 133 can be
synthesized in a similar manner.
##STR00052##
Compound 46 may be converted to compound 134 under similar
conditions as illustrated in Gattermann, L. (1914), Die Praxis des
organischen Chemikers (12), 228. Conversion of compound 134 to
compound 136 may be accomplished using conditions as previously
described. The 3,5-dibromophenyl analog of 136, the
3,5-dimethylphenyl analog of 136, the 3-chloro-5-methylphenyl
analog of 136, 3-bromo-5-methylphenyl analog of 136, and the
3-bromo-5-chlorophenyl analog of 136 can be synthesized in a
similar manner.
##STR00053## ##STR00054##
Conversion of compound 130 to compound 141 may be accomplished
using conditions as previously described. The 3,5-dibromophenyl
analog of 141, the 3,5-dimethylphenyl analog of 141, the
3-chloro-5-methylphenyl analog of 141, 3-bromo-5-methylphenyl
analog of 141, and the 3-bromo-5-chlorophenyl analog of 141 can be
synthesized in a similar manner.
##STR00055##
Conversion of compound 141 to compounds 142 and 143 may be
accomplished using conditions as previously described. The
3,5-dibromophenyl analog of 142 and 143, the 3,5-dimethylphenyl
analog of 142 and 143, the 3-chloro-5-methylphenyl analog of 142
and 143, 3-bromo-5-methylphenyl analog of 142 and 143, and the
3-bromo-5-chlorophenyl analog of 142 and 143 can be synthesized in
a similar manner.
##STR00056## ##STR00057##
Conversion of compound 94 to compound 149 may be accomplished using
conditions as previously described. The 3,5-dibromophenyl analog of
149, the 3,5-dimethylphenyl analog of 149, the
3-chloro-5-methylphenyl analog of 149, 3-bromo-5-methylphenyl
analog of 149, and the 3-bromo-5-chlorophenyl analog of 149 can be
synthesized in a similar manner.
##STR00058## ##STR00059##
Conversion of compound 85 to compound 154 may be accomplished using
conditions as previously described. The 3,5-dibromophenyl analog of
154, the 3,5-dimethylphenyl analog of 154, the
3-chloro-5-methylphenyl analog of 154, 3-bromo-5-methylphenyl
analog of 154, and the 3-bromo-5-chlorophenyl analog of 154 can be
synthesized in a similar manner.
##STR00060##
Conversion of compound 154 to compounds 155 and 156 may be
accomplished using conditions as previously described. The
3,5-dibromophenyl analog of 155 and 156, the 3,5-dimethylphenyl
analog of 155 and 156, the 3-chloro-5-methylphenyl analog of 155
and 156, 3-bromo-5-methylphenyl analog of 155 and 156, and the
3-bromo-5-chlorophenyl analog of 155 and 156 can be synthesized in
a similar manner.
##STR00061## ##STR00062##
Conversion of compound 152 to compound 162 may be accomplished
using conditions as previously described. The 3,5-dibromophenyl
analog of 162, the 3,5-dimethylphenyl analog of 162, the
3-chloro-5-methylphenyl analog of 162, 3-bromo-5-methylphenyl
analog of 162, and the 3-bromo-5-chlorophenyl analog of 162 can be
synthesized in a similar manner.
##STR00063##
Conversion of compound 162 to compounds 163 and 164 may be
accomplished using conditions as previously described. The
3,5-dibromophenyl analog of 163 and 164, the 3,5-dimethylphenyl
analog of 163 and 164, the 3-chloro-5-methylphenyl analog of 163
and 164, 3-bromo-5-methylphenyl analog of 163 and 164, and the
3-bromo-5-chlorophenyl analog of 163 and 164 can be synthesized in
a similar manner.
In the practice of the method of the present invention, an
effective amount of any one of the compounds of this invention or a
combination of any of the compounds of this invention or a
pharmaceutically acceptable salt or ester thereof, is administered
via any of the usual and acceptable methods known in the art,
either singly or in combination. The compounds or compositions can
thus be administered orally (e.g., buccal cavity), sublingually,
parenterally (e.g., intramuscularly, intravenously, or
subcutaneously), rectally (e.g., by suppositories or washings),
transdermally (e.g., skin electroporation) or by inhalation (e.g.,
by aerosol), and in the form or solid, liquid or gaseous dosages,
including tablets and suspensions. The administration can be
conducted in a single unit dosage form with continuous therapy or
in a single dose therapy ad libitum. The therapeutic composition
can also be in the form of an oil emulsion or dispersion in
conjunction with a lipophilic salt such as pamoic acid, or in the
form of a biodegradable sustained-release composition for
subcutaneous or intramuscular administration.
Useful pharmaceutical carriers for the preparation of the
compositions hereof, can be solids, liquids or gases; thus, the
compositions can take the form of tablets, pills, capsules,
suppositories, powders, enterically coated or other protected
formulations (e.g. binding on ion-exchange resins or packaging in
lipid-protein vesicles), sustained release formulations, solutions,
suspensions, elixirs, aerosols, and the like. The carrier can be
selected from the various oils including those of petroleum,
animal, vegetable or synthetic origin, e.g., peanut oil, soybean
oil, mineral oil, sesame oil, and the like. Water, saline, aqueous
dextrose, and glycols are preferred liquid carriers, particularly
(when isotonic with the blood) for injectable solutions. For
example, formulations for intravenous administration comprise
sterile aqueous solutions of the active ingredient(s) which are
prepared by dissolving solid active ingredient(s) in water to
produce an aqueous solution, and rendering the solution sterile.
Suitable pharmaceutical excipients include starch, cellulose, talc,
glucose, lactose, talc, gelatin, malt, rice, flour, chalk, silica,
magnesium stearate, sodium stearate, glycerol monostearate, sodium
chloride, dried skim milk, glycerol, propylene glycol, water,
ethanol, and the like. The compositions may be subjected to
conventional pharmaceutical additives such as preservatives,
stabilizing agents, wetting or emulsifying agents, salts for
adjusting osmotic pressure, buffers and the like. Suitable
pharmaceutical carriers and their formulation are described in
Remington's Pharmaceutical Sciences by E. W. Martin. Such
compositions will, in any event, contain an effective amount of the
active compound together with a suitable carrier so as to prepare
the proper dosage form for proper administration to the
recipient.
The pharmaceutical preparations can also contain preserving agents,
solubilizing agents, stabilizing agents, wetting agents,
emulsifying agents, sweetening agents, coloring agents, flavoring
agents, salts for varying the osmotic pressure, buffers, coating
agents or antioxidants. They can also contain other therapeutically
valuable substances, including additional active ingredients other
than those of formula I.
The compounds of the present invention are useful as medicaments
for the treatment of metabolic diseases such as obesity,
hyperlipidemia, hypercholesterolemia and diabetes, and may be
useful for other diseases such as NASH, atherosclerosis,
cardiovascular diseases, hypothyroidism, thyroid cancer and related
disorders and diseases. An obese patient is a human with a body
mass index of 25 or greater.
The therapeutically effective amount or dosage of a compound
according to this invention can vary within wide limits and may be
determined in a manner known in the art. Such dosage will be
adjusted to the individual requirements in each particular case
including the specific compound(s) being administered, the route of
administration, the condition being treated, as well as the patient
being treated. In general, in the case of oral or parenteral
administration to adult humans weighing approximately 70 kg, a
daily dosage of from about 0.01 mg/kg to about 50 mg/kg should be
appropriate, although the upper limit may be exceeded when
indicated. The dosage is preferably from about 0.3 mg/kg to about
10 mg/kg per day. A preferred dosage may be from about 0.70 mg/kg
to about 3.5 mg/kg per day. The daily dosage can be administered as
a single dose or in divided doses, or for parenteral administration
it may be given as continuous infusion.
The compounds of formula (I) are thyroid hormone analogs. The
TR/RXR/GRIP Assay was used to test compounds of formula (I), as
shown in the Examples below. Thus, the tested compounds are thyroid
hormone receptor agonists, having EC50 of 1000 .mu.M, or less, with
a preferred EC50 of 100 .mu.M or less.
The invention will now be further described in the Examples below,
which are intended as an illustration only and do not limit the
scope of the invention.
EXAMPLES
##STR00064##
##STR00065##
##STR00066##
Example 1
Synthesis of
[4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-
-acetic acid (10a)
##STR00067##
Step 1: Preparation of 4-dimethylaminomethyl-2,6-dimethyl-phenol
(2a)
A solution of 2,6-dimethyl phenol (20 g, 0.163 mol) in ethanol (100
mL) at room temperature was treated with dimethylamine (19 mL of a
40% solution of dimethylamine in water, 0.163 mol) followed by
formaldehyde (13.5 mL of a 37% solution of formaldehyde in water,
0.163 mol). The reaction was heated to reflux for 12 h. The
reaction mixture was cooled to 0.degree. C. in an ice bath, diluted
with water (100 mL) and brought to pH=5 with a 1N aqueous
hydrochloric acid solution. The water layer was extracted with
diethyl ether (2.times.150 mL). The aqueous layer was brought to
pH=8 by the addition of a 1N aqueous sodium hydroxide solution,
cooled to 0.degree. C. in an ice bath and was extracted with
diethyl ether (3.times.200 mL). The organics were washed with a
saturated aqueous sodium chloride solution, dried with magnesium
sulfate, filtered and concentrated under vacuum. The resulting oil
was dried under high vacuum overnight to afford
4-dimethylaminomethyl-2,6-dimethyl-phenol as an off-white solid
(13.42 g, 46%); EI(+)-HRMS m/z calcd for C.sub.11H.sub.17NO
(M.sup.+) 179.1310, found 179.1308. Molecular Weight=179.2642;
Exact Mass=179.1310
Step 2: Preparation of 2,6-dimethyl-4-trimethylaminomethylphenol
(3a)
A suspension of 4-dimethylaminomethyl-2,6-dimethylphenol (2a) (3.0
g, 0.016 mol) in diethyl ether (25 mL) was treated with methyl
iodide (2.08 mL, 0.033 mol) at room temperature. Additional diethyl
ether (20 mL) was added to the reaction. The reaction mixture was
stirred at room temperature overnight. The resulting solids were
filtered and washed with diethyl ether. NMR and LCMS indicated the
presence of starting material. The solids were resubmitted to the
reaction conditions. The reaction was stirred at room temperature
for 2 d. At this time, additional methyl iodide (2.08 mL, 0.033
mol) was added. The reaction was stirred at room temperature
overnight. At this time, the resulting solids were filtered and
washed with diethyl ether. The solids were dried under high vacuum
to afford 2,6-dimethyl-4-trimethylaminomethylphenol (3a) (5.1 g,
crude) as a white solid which was used without further
purification. Molecular Weight=195.3073; Exact Mass=195.1623
Step 3: Preparation of (3,5-dimethyl-4-hydroxy-phenyl)-acetonitrile
(4a)
A solution of 2,6-dimethyl-4-trimethylaminomethylphenol (3a)
(theoretically, 0.016 mol) in ethanol (50 mL) under argon was
treated with sodium cyanide (976 mg, 0.019 mol) at room
temperature. The reaction mixture was heated to reflux overnight.
The reaction mixture was then diluted with water and was acidified
by the addition of a 1N aqueous hydrochloric acid solution. The
mixture was extracted with ethyl acetate (3.times.200 mL). The
combined organics were washed with water (1.times.200 mL) and a
saturated aqueous sodium chloride solution (1.times.200 mL), dried
with magnesium sulfate, filtered and concentrated under vacuum to
afford a brown solid. The solids were slurried in diethyl ether (50
mL) and collected by filtration. The solid was purified by column
chromatography using silica gel eluted with 10-15% ethyl acetate in
petroleum ether to afford
3,5-dimethyl-4-hydroxyphenyl)-acetonitrile (4a) as a yellow solid
(1.61 g, 60% yield for the two steps); EI(+)-HRMS m/z calcd for
C.sub.10H.sub.11NO (M.sup.+) 161.0841, found 161.0841. Molecular
Weight=161.2053; Exact Mass=161.0841
Step 4: Preparation of (3,5-dimethyl-4-hydroxy-phenyl)-acetic acid
(5a)
A suspension of (3,5-dimethyl-4-hydroxy-phenyl)-acetonitrile (4a)
(550 mg, 0.003 mol) in water (0.98 mL) was treated with ethylene
glycol dimethyl ether (6.6 mL, 0.063 mol) followed by potassium
hydroxide (1.34 g, 0.024 mol). The reaction mixture was heated to
reflux for 2 d. The reaction mixture was concentrated under vacuum.
The resulting solid was diluted with water (100 mL) and extracted
with ethyl acetate (2.times.75 mL). The organic layers were
discarded. The aqueous layer was acidified to pH=2 by the addition
of a 1N aqueous hydrochloric acid solution and was extracted with
ethyl acetate (2.times.100 mL). The organic layers were combined,
washed with a saturated aqueous sodium chloride solution
(2.times.50 mL), dried with magnesium sulfate, filtered and
concentrated under vacuum. The solid was purified by column
chromatography using silica gel eluted with 15-25% ethyl acetate in
petroleum ether to afford (3,5-dimethyl-4-hydroxy-phenyl)-acetic
acid (5a) (30 mg, 5%) as a yellow solid; EI(+)-HRMS m/z calcd for
C.sub.10H.sub.12O.sub.3 (M.sup.+) 180.0786, found 180.0782.
Molecular Weight=180.2053; Exact Mass=180.0786
Step 5: Preparation of (3,5-dimethyl-4-hydroxy-phenyl)-acetic acid
methyl ester (6a)
A solution of (3,5-dimethyl-4-hydroxy-phenyl)-acetic acid (5a)
(12.4 g, 0.069 mmol) in methanol (300 mL) was treated with
concentrated sulfuric acid (6.25 mL). The resulting solution was
heated to 70.degree. C. overnight. At this time, the reaction was
cooled to room temperature. The reaction was concentrated under
vacuum. The residue was diluted with ethyl acetate (700 mL) and was
washed with water (2.times.250 mL). The organics were dried over
magnesium sulfate, filtered and concentrated under vacuum to give
an orange solid. The solid was slurried in 10% ethyl
acetate/petroleum ether and was stirred for 1 h. The solid was
collected by filtration, washed with 10% ethyl acetate/petroleum
ether, and dried under vacuum overnight. The residue was purified
by column chromatography using silica gel eluted with 10-15% ethyl
acetate in petroleum ether to afford
(3,5-dimethyl-4-hydroxy-phenyl)-acetic acid methyl ester (6a) (1.93
g, 86%) as an off-white solid; EI(+)-HRMS m/z calcd for
C.sub.11H.sub.14O.sub.3 (M.sup.+) 194.0943, found 191.0942.
Molecular Weight=194.2324; Exact Mass=194.0943
Step 6: Preparation of 3,6-dichloro-4-isopropyl pyridazine (7)
A solution of 3,6-dichloropyridazine (22.5 g, 0.15 mol) in
acetonitrile (35 mL), tetramethylene sulfone (107 mL) and water
(245 mL) at room temperature was treated with isobutyric acid (14
mL, 0.151 mol) followed by silver nitrate (13 g, 0.075 mol). The
reaction mixture was heated to 55.degree. C. A solution of
concentrated sulfuric acid (24 mL) in water (75 mL) was added in
one portion followed by the dropwise addition over 35 min of a
solution of ammonium persulfate (51.5 g, 0.22 mol) in water (75
mL). The reaction mixture was heated to 70.degree. C. for 20 min
and then cooled to room temperature and stirred for 24 h. At this
time, the reaction was cooled to 0.degree. C. and 28-30% ammonium
hydroxide (100 mL) was added slowly dropwise to bring the reaction
to pH=8. The reaction was diluted with water (500 mL) and was
filtered over celite and washed well with ethyl acetate (500 mL).
The water layer and the organic layer were separated. The organic
layer was saved. The aqueous layer was extracted with ethyl acetate
(2.times.500 mL). The combined organics were washed with water
(1.times.400 mL) and a saturated aqueous sodium chloride solution
(1.times.400 mL), dried with magnesium sulfate, filtered and
concentrated under vacuum. The resulting oil was purified by column
chromatography using silica gel eluted with petroleum ether
followed by 10% ethyl acetate in petroleum ether to afford
3,6-dichloro-4-isopropyl pyridazine (7) (19.26 g, 67%) as an oil;
(+)-HRMS m/z calcd for C.sub.7H.sub.8Cl.sub.2N.sub.2 (M.sup.+)
190.0065, found 190.0059. Molecular Weight=191.0612; Exact
Mass=190.0065
Step 7: Preparation of
[4-(6-Chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid methyl ester (8a)
A solution of (3,5-dimethyl-4-hydroxy-phenyl)-acetic acid methyl
ester (6a) (12.5 g, 0.064 mol) in anhydrous dimethyl sulfoxide (256
mL) under argon at room temperature was treated with
3,6-dichloro-4-isopropyl pyridazine (7) (18.43 g, 0.096 mol)
followed by anhydrous potassium carbonate (17.69 g, 0.12 mol) and
copper (I) iodide (6.09 g, 0.032 mol). The reaction mixture was
heated to 90.degree. C. for 24 h. At this time, the reaction was
cooled to room temperature and was poured onto a 1N aqueous
hydrochloric acid solution (200 mL) and ice. The aqueous layer was
diluted with water (100 mL) and extracted with ethyl acetate
(2.times.500 mL). The aqueous layer was made basic (pH=8) by the
addition of a 1N aqueous sodium hydroxide solution. The water layer
was extracted again with ethyl acetate (1.times.500 mL). The
combined organics were washed with a saturated aqueous sodium
chloride solution, dried with magnesium sulfate, filtered and
concentrated under vacuum. The resulting residue was purified by
column chromatography using silica gel eluted with 5-15% ethyl
acetate in petroleum ether to afford
[4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid methyl ester (8a) (15 g, 67%) and a minor amount of isomer as
a white solid; EI(+)-HRMS m/z calcd for
C.sub.18H.sub.21ClN.sub.2O.sub.3 (M.sup.+) 348.1241, found
348.1237. Molecular Weight=348.8327; Exact Mass=348.1241
Step 8: Preparation of
[4-(6-Chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid (9a)
A solution of
[4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid methyl ester (8a) (1.66 g, 4.75 mmol, contains a minor amount
of isomer) in methanol (50 mL) at room temperature was treated
dropwise with a 1N aqueous sodium hydroxide solution (9.5 mL, 9.50
mmol). The reaction was stirred at room temperature for 24 h. At
this time, the reaction mixture was concentrated under vacuum. The
resulting solid was diluted with water (200 mL) and extracted with
ethyl acetate (200 mL). The ethyl acetate layer was discarded. The
water layer was acidified to pH=4 by the addition of a 1N aqueous
hydrochloric acid solution and was extracted with ethyl acetate
(2.times.200 mL). The organic layers were combined, washed with a
saturated aqueous sodium chloride solution, dried with magnesium
sulfate, filtered and concentrated under vacuum to afford a solid.
The solid was dried overnight under high vacuum to afford
[4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid (9a) (1.58 g, 100%, contains a minor amount of isomer) as an
off-white solid; EI(+)-HRMS m/z calcd for
C.sub.17H.sub.19ClN.sub.2O.sub.3 (M.sup.+) 334.1084, found
334.1083. Molecular Weight=334.8056; Exact Mass=334.1084
Step 9: Preparation of
[4-(5-Isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-
-acetic acid (10a)
A solution of
[4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-acetic
acid (9a) (0.10 g, 0.29 mol) in glacial acetic acid (3 mL) was
treated with sodium acetate (54 mg, 0.65 mol) at room temperature.
The reaction mixture was heated to 100.degree. C. for 24 h. At this
time, the reaction mixture was cooled to room temperature and was
concentrated under vacuum. The resulting residue was diluted with
water (100 mL), made basic by the addition of a 1N aqueous sodium
hydroxide solution and was extracted with ethyl acetate (200 mL).
The ethyl acetate layer was discarded. The water layer was
acidified to pH=4 by the addition of a 1N aqueous hydrochloric acid
solution and was extracted with ethyl acetate (2.times.200 mL). The
organic layers were combined, washed with a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, filtered
and concentrated under vacuum to afford a solid. The solid was
dissolved in ethyl acetate by adding methanol to form a solution
and it was absorbed onto silica and concentrated under vacuum. The
preabsorbed solid was purified by column chromatography using
silica gel eluted with 20% ethyl acetate in hexanes containing 2%
glacial acetic acid. The desired fractions were concentrated as
several separate batches and placed under high vacuum for 1 h.
.sup.1H NMRs were obtained to determine if batches contained only
the desired isomer. The best batches were combined, diluted with a
1:1 methylene chloride:hexanes solution. This process was performed
three times. The solid was dried under high vacuum overnight and
then slurried in acetonitrile and filtered. The resulting solid was
dried in the vacuum oven at 60.degree. C. for 24 h to afford
[4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-3,5-dimethyl-phenyl]-
-acetic acid (10a) (61 mg, 65%) as a white solid; EI(+)-HRMS m/z
calcd for C.sub.17H.sub.20N.sub.2O.sub.4 (M.sup.+) 316.1423, found
316.1427. Molecular Weight=316.3599; Exact Mass=316.1423
Example 2
Preparation of
[3-Chloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-5-methyl-ph-
enyl]-acetic acid (10b)
##STR00068##
Step 1: Preparation of
2-Chloro-4-dimethylaminomethyl-6-methyl-phenol (2b)
A solution of 2-chloro-6-methyl phenol (5.0 g, 0.035 mol) in
ethanol (25 mL) at room temperature was treated with dimethyl amine
(3.95 mL of a 40% solution of dimethylamine in water, 0.035 mol)
followed by formaldehyde (2.85 mL of a 37% solution of formaldehyde
in water, 0.035 mol). The reaction was heated to reflux for 24 h.
At this time, the reaction mixture was cooled to room temperature
and concentrated under vacuum. The resulting oil was diluted with
water (200 mL) and acidified to pH=2 by addition of a 1N aqueous
hydrochloric acid solution. The water layer was extracted with
ethyl acetate (300 mL). The ethyl acetate layer was discarded. The
water layer was made basic to pH=10 by the addition of a 1N aqueous
sodium hydroxide solution and was extracted with ethyl acetate
(2.times.350 mL). The organics were washed with a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, filtered
and concentrated under vacuum. The resulting oil was dried under
high vacuum overnight to afford
2-chloro-4-dimethylaminomethyl-6-methyl-phenol (2b) (4.62 g, 66%)
as an off-white solid; EI(+)-HRMS m/z calcd for
C.sub.10H.sub.14ClNO (M.sup.+) 199.0764, found 199.0767. Molecular
Weight=199.6822; Exact Mass=199.0764
Step 2: Preparation of
2-Chloro-6-methyl-4-trimethylaminomethylphenol (3b)
A suspension of 2-chloro-4-dimethylaminomethyl-6-methyl-phenol (2b)
(65.5 g, 0.328 mol) in ether (1.5 L) under argon was treated with
methyl iodide (40.83 mL, 0.65 mol) at room temperature. The
reaction mixture was stirred at room temperature overnight. At this
time, the resulting solids were filtered and washed with ether. The
solids were dried under high vacuum to afford
2-chloro-6-methyl-4-trimethylaminomethylphenol (3b) as an off-white
solid which was used as is for the next step. Molecular
Weight=215.7252; Exact Mass=215.1077
Step 3: Preparation of
(3-Chloro-4-hydroxy-5-methyl-phenyl)-acetonitrile (4b)
A suspension of 2-chloro-6-methyl-4-trimethylaminomethylphenol (3b)
(theoretically 0.32 mol) in ethanol (2 L) under argon was treated
with sodium cyanide (16.01 g, 0.32 mol) at room temperature. The
reaction mixture was heated to reflux for 2 d. At this time, the
reaction mixture was concentrated under vacuum. The resulting
residue was diluted with water and was acidified to pH=2 by the
addition of a 1N aqueous hydrochloric acid solution. The water
layer was extracted with ethyl acetate (3.times.600 mL). The
organic layers were combined, washed with a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, filtered
and concentrated under vacuum to afford a brown solid. The solid
was dissolved in carbon tetrachloride (250 mL) and the remaining
dark gums were removed via filtration. Precipitates formed in the
filtrate upon standing at room temperature. The solids were
filtered, rinsed with hexanes, and dried under high vacuum to
afford (3-chloro-4-hydroxy-5-methyl-phenyl)-acetonitrile (4b) (30.3
g) as a yellow solid. The filtrate was concentrated and the
resulting residue was purified by column chromatography using
silica gel eluted with 5-10% ethyl acetate in petroleum ether to
afford (3-chloro-4-hydroxy-5-methyl-phenyl)-acetonitrile (4b) (2.28
g, 55% combined yield for 2 steps) as a yellow solid; EI(+)-HRMS
m/z calcd for C.sub.9H.sub.8ClNO (M.sup.+) 181.0294, found
181.0295. Molecular Weight=181.6232; Exact Mass=181.0294
Step 4: Preparation of (3-Chloro-4-hydroxy-5-methyl-phenyl)-acetic
acid (5b)
A suspension of (3-chloro-4-hydroxy-5-methyl-phenyl)-acetonitrile
(4b) (1.3 g, 0.0071 mol) in water (2.06 mL) was treated with
ethylene glycol dimethyl ether (13.93 mL, 0.133 mol) followed by
potassium hydroxide (2.8 g, 0.0071 mol). The reaction mixture was
heated to reflux for 24 h. At this time, the reaction mixture was
concentrated under vacuum. The resulting solid was diluted with
water (100 mL) and extracted with ethyl acetate (2.times.50 mL).
The organic layers were discarded. The water layer was acidified to
pH=2 by the addition of a 1N aqueous hydrochloric acid solution and
was extracted with ethyl acetate (2.times.50 mL). The organic
layers were combined, washed with a saturated aqueous sodium
chloride solution (50 mL), dried with magnesium sulfate, filtered
and concentrated under vacuum. The resulting solid was dried under
high vacuum overnight to afford
(3-chloro-4-hydroxy-5-methyl-phenyl)-acetic acid (5b) (1.4 g, 97%)
as a white solid; EI(+)-HRMS m/z calcd for C.sub.9H.sub.9ClO.sub.3
(M.sup.+) 200.0240, found 200.0247. Molecular Weight=200.6233;
Exact Mass=200.0240
Step 5: Preparation of (3-Chloro-4-hydroxy-5-methyl-phenyl)-acetic
acid methyl ester (6b)
A solution of (3-chloro-4-hydroxy-5-methyl-phenyl)-acetic acid (5b)
(1.4 g, 6.98 mmol) in methanol (60 mL) was treated with
concentrated sulfuric acid (0.5 mL) at room temperature under
argon. The reaction mixture was heated to 70.degree. C. for 24 h.
At this time, the reaction mixture was concentrated under vacuum.
The resulting oil was diluted with ethyl acetate (100 mL). The
organics were washed with water (2.times.50 mL), dried with
magnesium sulfate, filtered and concentrated. The resulting residue
was purified by column chromatography using silica gel eluted with
10% ethyl acetate in petroleum ether to afford
3-chloro-4-hydroxy-5-methyl-phenyl)-acetic acid methyl ester (6b)
(1.36 g, 91%) as a white solid; EI(+)-HRMS m/z calcd for
C.sub.10H.sub.11ClO.sub.3 (M.sup.+) 214.0397, found 214.0400.
Molecular Weight=214.6504; Exact Mass=214.0397
Step 6: Preparation of
[3-Chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid methyl ester (8b)
A solution of 3-chloro-4-hydroxy-5-methyl-phenyl)-acetic acid
methyl ester (6b) (909 mg, 0.004 mol) in anhydrous dimethyl
sulfoxide (8 mL) under argon at room temperature was treated with
3,6-dichloro-4-isopropyl pyridazine (7) (1.2 g, 0.006 mol),
anhydrous potassium carbonate (1.15 g, 0.008 mol) and copper (I)
iodide (239 mg, 0.001 mol). The reaction mixture was heated to
90.degree. C. overnight. At this time, the reaction mixture was
cooled to room temperature, poured onto a 1N aqueous hydrochloric
acid solution and was extracted with ethyl acetate. The organics
were washed with a saturated aqueous sodium chloride solution,
dried with magnesium sulfate, filtered and concentrated under
vacuum. The resulting residue was purified by column chromatography
using silica gel eluted with 10% ethyl acetate in petroleum ether
to afford
[3-chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid methyl ester (8b) (1.2 g, 78%) as a yellow solid;
EI(+)-HRMS m/z calcd for C.sub.17H.sub.18Cl.sub.2N.sub.2O.sub.3
(M.sup.+) 369.0767, found 369.0766. Molecular Weight=369.2506;
Exact Mass=368.0694
Step 7: Preparation of
[3-Chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid (9b)
A solution of
[3-chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid methyl ester (8b) (1.1 g, 2.97 mmol) in methanol (30 mL)
at room temperature was treated dropwise with a 1N aqueous sodium
hydroxide solution (5.9 mL, 5.9 mmol). The reaction was stirred at
room temperature for 4 d. At this time, the reaction mixture was
concentrated under vacuum. The resulting solid was diluted with
water and ethyl acetate. The ethyl acetate layer was discarded. The
water layer was acidified by the addition of a 1N aqueous
hydrochloric acid solution and was extracted with ethyl acetate.
The organic layers were washed with a saturated aqueous sodium
chloride solution, dried with magnesium sulfate, filtered and
concentrated under vacuum to afford
[3-chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid (9b) (1.05 g, 100%) as a white solid. This material was
used without further purification; EI(+)-HRMS m/z calcd for
C.sub.16H.sub.16Cl.sub.2N.sub.2O.sub.3 (M.sup.+) 355.0611, found
355.0610. Molecular Weight=355.2235; Exact Mass=354.0538
Step 8: Preparation of
[3-Chloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-5-methyl-ph-
enyl]-acetic acid (10b)
A solution of
[3-chloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-5-methyl-phenyl]-ace-
tic acid (9b) (1.0 g, 2.81 mmol) in glacial acetic acid (30 mL) was
treated with sodium acetate (808 mg, 9.83 mmol) at room
temperature. The reaction mixture was heated to 105.degree. C. for
24 h. At this time, the reaction mixture was cooled to room
temperature and concentrated under vacuum. The resulting residue
was diluted with water, brought to pH=10 by the addition of a 1N
aqueous sodium hydroxide solution and was extracted with ethyl
acetate. The ethyl acetate layer was discarded. The water layer was
acidified by the addition of a 3N aqueous hydrochloric acid
solution and was extracted with ethyl acetate. The organic layers
were combined, washed with water and a saturated aqueous sodium
chloride solution, dried with magnesium sulfate, filtered and
concentrated under vacuum. The resulting solid was dissolved in
methylene chloride and methanol and then was absorbed onto silica.
The preabsorbed solid was purified by column chromatography using
silica gel eluted with 20% ethyl acetate in hexanes containing 2%
glacial acetic acid. The desired fractions were concentrated as
several separate batches and placed under high vacuum for 15 min.
The solid was diluted with a 1:1 methylene chloride:hexanes
solution. This process was performed three times. The solid was
dried under high vacuum overnight. The solid was then slurried in
acetonitrile, filtered, and dried in the vacuum oven at 80.degree.
C. overnight to afford
[3-chloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-5-methyl-ph-
enyl]-acetic acid (10b) (300 mg, 32%) as a white solid; EI(+)-HRMS
m/z calcd for C.sub.16H.sub.17ClN.sub.2O.sub.4 (M+H).sup.+
337.0950, found 337.0949. Molecular Weight=336.7779; Exact
Mass=336.0877
##STR00069##
Example 3
Synthesis of
[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
acetic acid (14)
##STR00070##
Step 1: Preparation of 2,6-Dibromo-4-(2-hydroxy-ethyl)-phenol
(11)
A solution of 4-(2-hydroxy-ethyl)-phenol (50 g, 0.35 mol) in
glacial acetic acid (400 mL) was treated with a solution of bromine
(41 mL, 0.78 mol) in glacial acetic acid (40 mL). The reaction
mixture was stirred at room temperature overnight. At this time,
the solvent was removed under vacuum. The resulting residue was
diluted with toluene (200 mL) and the solvent was again
concentrated under vacuum. The resulting residue was dissolved in
tetrahydrofuran (250 mL) and a 4N aqueous sodium hydroxide solution
(350 mL) was added followed by water (150 mL). The reaction was
stirred at room temperature for 3 h. The reaction was then brought
to pH=5 by the addition of concentrated hydrochloric acid (80 mL).
The layers were separated (the bottom layer is product). The
aqueous layer was extracted with methyl tert-butyl ether (500 mL).
The organic layers were combined, dried with sodium sulfate,
filtered and concentrated under vacuum to about 100 mL. Heptane
(100 mL) was added and the solvent was concentrated under vacuum to
about 100 mL. The solid was filtered and washed with 10% methyl
tert-butyl ether in heptane (150 mL). The solid was dried under
high vacuum overnight to afford
2,6-dibromo-4-(2-hydroxy-ethyl)-phenol (11) (86.8 g, 82%) as a
white solid; LRMS for C.sub.8H.sub.8Br.sub.2O.sub.2 (M-H) m/z=295.
Molecular Weight=295.9598; Exact Mass=293.8891
Step 2: Preparation of
2-[3,5-Dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethanol
(12)
A mixture of 2,6-dibromo-4-(2-hydroxy-ethyl)-phenol (11) (183.3 g,
0.619 mol) in N,N-dimethyl acetamide (300 mL) was treated with
potassium tert-butoxide (70.5 g, 0.59 mol) under nitrogen at room
temperature. The suspension was heated to 10.degree. C. and stirred
until a solution formed. At this time, 3,6-dichloro-4-isopropyl
pyridazine (7) (100 g, 0.50 mol) was added to the solution and the
reaction mixture was stirred at 135.degree. C. for 24 h. The
reaction was cooled to room temperature, diluted with water (350
mL) and extracted with tert-butyl methyl ether (1.times.600 mL)
followed by isopropyl acetate (1.times.600 mL). The organic layers
were combined and washed with a 1N aqueous sodium hydroxide
solution (1.times.256 mL) and water (2.times.200 mL). The organic
layer was distilled to about 300 mL. The residue was treated with
heptane (300 mL). The mixture was stirred under reflux for 30 min
and then was cooled to room temperature. The resulting solid was
filtered, washed with a 2:1 mixture of tert-butyl methyl ether:
heptane (240 mL) and dried overnight to afford
2-[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethanol
(12) (112 g, 49%) as an off-white solid; LRMS for
C.sub.15H.sub.15Br.sub.2ClN.sub.2O.sub.2 (M.sup.+) at m/z=450.
Molecular Weight=450.5600; Exact Mass=447.9189
Step 3: Preparation of
[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
acetic acid (14)
A solution of
2-[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethanol
(12) (111.4 g, 247 mmol) in acetonitrile (223 mL), methylene
chloride (343 mL) and sodium phosphate buffer (343 mL of a 0.67 M
solution, pH=6.7) was treated with
2,2,6,6-tetramethyl-1-piperidinyloxy free group (2.35 g, 14.83
mmol) at room temperature. A solution of sodium chlorite (44.71 g,
395.5 mmol) in water (135 mL) and a solution of sodium hypochlorite
(28 mL, 24.5 mmol) in water (50 mL) were then added simultaneously
to the reaction mixture over 45 min. The temperature of the
reaction mixture rose to 40.degree. C. during the addition and a
cold water bath was used to prevent the temperature from going any
higher. The brown reaction mixture was stirred for 30 min. While
cooling with cold water, a solution of sodium bisulfite (28.42 g,
297 mmol) in water (86 mL) was added dropwise to the reaction
mixture over 5 min. The yellow solution was diluted with methylene
chloride (223 mL) and extracted. The layers were separated and the
organic layer was extracted with water (2.times.200 mL), dried with
sodium sulfate, filtered and concentrated under vacuum to afford
[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-acetic
acid (13). Glacial acetic acid (549 mL) and sodium acetate (40.56
g, 494 mmol) were added to
[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-acetic
acid (13). The reaction mixture was heated to 120.degree. C. for 24
h. At this time, the reaction mixture was cooled to room
temperature, diluted with toluene (200 mL) and concentrated under
vacuum two times. The resulting residue was diluted with
tetrahydrofuran (223 mL) and isopropyl acetate (892 mL) and was
washed with water (3.times.150 mL). The organic layer was separated
and concentrated under vacuum to about 600 mL. Isopropyl acetate
(180 mL) was added and the mixture was concentrated under vacuum to
about 450 mL. Heptane (300 mL) was added in portions over 10 min.
The mixture was stirred under reflux for 15 min and then was cooled
to room temperature. The resulting solid was filtered, washed with
50% isopropyl acetate in heptane (200 mL), and dried under high
vacuum overnight to afford
[3,5-dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
acetic acid (14) (80.4 g, 73%) as a white solid; LRMS for
C.sub.15H.sub.14Br.sub.2N.sub.2O.sub.4 (M+H) m/z=446. Molecular
Weight=446.0978; Exact Mass=443.9320
##STR00071## ##STR00072##
Example 4
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-
-acetic acid (19)
##STR00073##
Step 1: Preparation of (3,5-Dichloro-4-hydroxy-phenyl)-acetic acid
methyl ester (15)
A solution of (4-hydroxy-phenyl)-acetic acid methyl ester (25 g,
0.150 mol) in toluene (600 mL) under argon at room temperature was
treated with diisobutyl amine (2.8 mL, 0.015 mol). The reaction
mixture was heated to 70.degree. C. and a solution of sulfuryl
chloride (24 mL, 0.30 mol) in toluene (75 mL) was added slowly
dropwise over 30 min. The reaction mixture was heated 1 h at
70.degree. C. At this time, the reaction mixture was cooled to
0.degree. C. and then poured onto ice water. The resulting white
solid was filtered and washed well with water. The filtrate was
saved. The solid was dissolved in ethyl acetate (500 mL) and washed
with water (1.times.125 mL). The organic layer was separated, dried
with magnesium sulfate, filtered and concentrated under vacuum to
give a white solid. The solid was dried overnight under high vacuum
to afford (3,5-dichloro-4-hydroxy-phenyl)-acetic acid methyl ester
(15) (18.81 g) as a white solid. The filtrate containing water and
toluene were separated. The toluene layer was washed with water
(150 mL), dried with magnesium sulfate, filtered and concentrated
under vacuum to give a pale yellow solid. The solid was slurried in
10% ethyl acetate in hexanes, filtered and washed well with
hexanes. The solid was dried overnight under high vacuum to afford
(3,5-dichloro-4-hydroxy-phenyl)-acetic acid methyl ester (15) (10.5
g) as a white solid (83% for the two crops); EI(+)-HRMS m/z calcd
for C.sub.9H.sub.8Cl.sub.2O.sub.3 (M.sup.+) 233.9850, found
233.9839. Molecular Weight=235.0683; Exact Mass=233.9850
Step 2: Preparation of 2,6-Dichloro-4-(2-hydroxy-ethyl)-phenol
(16)
A solution of (3,5-dichloro-4-hydroxy-phenyl)-acetic acid methyl
ester (15) (15.7 g, 67.09 mmol) in anhydrous tetrahydrofuran (600
mL) under argon cooled to -10.degree. C. was treated slowly
dropwise with a 1 M solution of lithium aluminum hydride in
tetrahydrofuran (67.1 mL, 67.1 mmol). The reaction mixture was
stirred for 5 min after the addition was complete and was quenched
at -10.degree. C. by the dropwise addition of a 10% aqueous
Rochelle's Salt solution (80 mL). The reaction suspension was
stirred 10 min. Ethyl acetate (100 mL) was then added to the
suspension. The mixture was then filtered. The solids were rinsed
with ethyl acetate (2.times.500 mL). The organic layer was
separated. The aqueous layer was extracted with ethyl acetate
(2.times.200 mL). The organic layers were combined and washed with
a 0.1N aqueous hydrochloric acid solution, water, and a saturated
aqueous sodium chloride solution. The organic layer was dried with
magnesium sulfate, filtered and concentrated to afford the first
crop of solid. The resulting solid was placed under high vacuum
overnight. The aqueous layer was acidified to pH=5 by the addition
of a 1N aqueous hydrochloric acid solution and was then
re-extracted with ethyl acetate (3.times.200 mL). The organic
layers were combined and washed with water and a saturated aqueous
sodium chloride solution. The organic layer was dried with
magnesium sulfate, filtered and concentrated. The resulting solid
was placed under high vacuum overnight to afford
2,6-dichloro-4-(2-hydroxy-ethyl)-phenol (16) (13.9 g, 100%); LRMS
for C.sub.8H.sub.8Cl.sub.2O.sub.2 (M.sup.+) m/z=207 Molecular
Weight=207.0578; Exact Mass=205.9901
Step 3: Preparation of
2-[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17)
A mixture of 2,6-dichloro-4-(2-hydroxy-ethyl)-phenol (16) (12.5 g,
60.2 mmol) in N,N-dimethyl acetamide (28 mL) was treated with
potassium tert-butoxide (6.5 g, 57.8 mmol) under nitrogen at room
temperature. The suspension was heated to 100.degree. C. and
stirred until a solution formed. The reaction was then treated with
3,6-dichloro-4-isopropyl pyridazine (7) (9.2 g, 48 mmol) followed
by a rinse with N,N-dimethyl acetamide (3 mL). The reaction mixture
was stirred at 135.degree. C. for 24 h. The reaction was cooled to
room temperature, diluted with water (30 mL) and extracted with
tert-butyl methyl ether (1.times.60 mL) followed by isopropyl
acetate (1.times.60 mL). The organic layers were combined and were
washed with a 1N aqueous sodium hydroxide solution (1.times.25 mL)
and water (2.times.25 mL). The organic layer was separated and
distilled to a volume of approximately 30 mL. This solution was
treated with heptane (30 mL). The mixture was stirred under reflux
for 30 min and then cooled to room temperature. The resulting solid
was filtered, washed with ether and dried overnight to afford
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17) (7.5 g, 44%) as a white solid; LRMS for
C.sub.15H.sub.15Cl.sub.3N.sub.2O.sub.2 (M+H) m/z=363. Molecular
Weight=361.6580; Exact Mass=360.0199
Step 4: Preparation of
[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-acetic
acid (18)
A solution of
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17) (8.4 g, 23.2 mmol) in acetone (270 mL) was treated with
Jones Reagent (34.8 mL of a 2.7 M solution, prepared via standard
method) slowly dropwise at -4.degree. C. The resulting red reaction
mixture was stirred for 1 h at -3 to 0.degree. C. The red reaction
mixture was quenched with isopropanol. The resulting green
suspension was filtered through celite and the celite was washed
well with ethyl acetate (600 mL). The filtrate was washed with
water (600 mL) and a saturated aqueous sodium chloride solution
(300 mL). The organic layer was separated, dried with magnesium
sulfate, filtered, and concentrated. The resulting solid was dried
under vacuum overnight to afford
[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-acetic
acid (18) (8.3 g, 95%) as a white solid; LRMS for
C.sub.15H.sub.15Cl.sub.3N.sub.2O.sub.3 (M.sup.+) m/z=377. Molecular
Weight=377.6574; Exact Mass=376.0148
Step 5: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-
-acetic acid (19)
A mixture of glacial acetic acid (200 mL), sodium acetate (6.1 g,
74.4 mmol) and
[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl-
]-acetic acid (18) (8.3 g, 21.96 mmol) was heated to 125.degree. C.
for 24 h. The reaction mixture was cooled to room temperature and
concentrated. The resulting residue was diluted with methylene
chloride (200 mL) and was washed with water (150 mL). The organic
layer was separated. Hexanes (3.times.200 mL) were added to the
residue in portions and then subsequently concentrated under
vacuum. The resulting semi-solid was diluted with a minimum amount
of ether, scratched, and slurried. The resulting white solid was
filtered, washed with cold ether and dried under high vacuum
overnight to afford
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-
-acetic acid (19) (5.1 g, 65%) as a white solid; LRMS for
C.sub.15H.sub.14Cl.sub.2N.sub.2O.sub.4 (M.sup.+) m/z=357. Molecular
Weight=357.1958; Exact Mass=356.0331
##STR00074## ##STR00075##
Example 5
Synthesis of
3-[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-proprionic acid (23)
##STR00076##
Step 1: Preparation of 2,6-Dibromo-4-(3-hydroxy-propyl)-phenol
(20)
A solution of 4-(3-hydroxy-propyl)-phenol (10 g, 65.7 mmol) in
glacial acetic acid (73.8 mL) was treated with a solution of
bromine (7.4 mL, 144.5 mmol) in glacial acetic acid (7.3 mL). The
reaction mixture was stirred at room temperature overnight. The
solvent was removed under vacuum. The resulting residue was diluted
with toluene (37 mL) and the solvent was again concentrated under
vacuum. The resulting residue was dissolved in tetrahydrofuran (46
mL) and a 4N aqueous sodium hydroxide solution (64 mL) was added
followed by water (27.5 mL). The reaction was stirred at room
temperature for 2.5 h and the pH was adjusted to 5 by the addition
of concentrated hydrochloric acid (14 mL). The layers were
separated (the bottom layer is product). The aqueous layer was
extracted with methyl tert-butyl ether (3.times.100 mL). The
organic layers were combined, dried with sodium sulfate, filtered
and concentrated under vacuum. The resulting residue was purified
by column chromatography using silica gel eluted with a 1:1 ethyl
acetate:hexanes solution. The desired fractions were collected and
concentrated under vacuum. The resulting oil was dried under high
vacuum overnight to afford 6-dibromo-4-(3-hydroxy-propyl)-phenol
(20) (20.2 g, 99%) as a pink oil; LRMS for
C.sub.9H.sub.10Br.sub.2O.sub.2 (M-H) m/z=309. Molecular
Weight=309.9869; Exact Mass=307.9048
Step 2: Preparation of
3-[3,5-Dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-propan--
1-ol (21)
A mixture of 2,6-dibromo-4-(3-hydroxy-propyl)-phenol (20) (5.0 g,
16.3 mmol) in N,N-dimethyl acetamide (8 mL) was treated with
potassium tert-butoxide (1.74 g, 15.48 mmol) under nitrogen at room
temperature. The suspension was heated to 100.degree. C. for 15 min
and turned brown in color. 3,6-Dichloro-4-isopropyl pyridazine (7)
(2.47 g, 12.9 mmol) was added to the suspension and the reaction
was stirred at 140.degree. C. for 24 h. The reaction was cooled to
room temperature, diluted with water (180 mL) and extracted with
ethyl acetate (3.times.100 mL). The organic layers were combined
and were washed with a 1N aqueous sodium hydroxide solution
(1.times.150 mL), followed by a saturated aqueous sodium chloride
solution (1.times.150 mL). The organic layer was dried with sodium
sulfate, filtered and concentrated. The resulting residue was
purified by flash chromatography (Biotage 40M) eluted with 15%
ethyl acetate in hexanes, followed by 25% ethyl acetate in hexanes,
followed by 50% ethyl acetate in hexanes. The desired fractions
were collected and concentrated under vacuum. The resulting solid
was slurried in cold acetonitrile and filtered. The solid was then
diluted with a 1:1 mixture of isopropyl acetate:methyl tert-butyl
ether (20 mL). The mixture was heated to reflux and then cooled to
room temperature. The solvent was decanted. The solid was slurried
in heptane, filtered and dried under high vacuum overnight. The
filtrate was concentrated. The resulting solid was then diluted
with a 1:1 mixture of isopropyl acetate: methyl tert-butyl ether
(20 mL). The mixture was heated to reflux and then cooled to room
temperature. The solvent was decanted and the solid was slurried in
heptane and filtered to afford a second crop of solid which was
dried under high vacuum overnight. The solids were combined to
afford
3-[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-propan--
1-ol (21) (1.48 g, 20%) as a white solid; LRMS for
C.sub.16H.sub.17Br.sub.2ClN.sub.2O.sub.2 (M+H) m/z=465. Molecular
Weight=464.5871; Exact Mass=461.9345
Step 3: Preparation of
3-[3,5-Dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-propion-
ic acid (22)
This compound was prepared by a similar method to that described in
Example 4, Step 4 except that
3-[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-propano-
l (21) was used in place of
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17) to afford
3-[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]proprion-
ic acid (22) (95%) as a white solid; EI(+)-HRMS m/z calcd for
C.sub.16H.sub.15Br.sub.2ClN.sub.2O.sub.3 (M+H) 476.9211, found
476.9210. Molecular Weight=478.5706; Exact Mass=475.9138
Step 4: Preparation of
3-[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-propionic acid (23)
A mixture of glacial acetic acid (16.7 mL), sodium acetate (480 mg,
5.85 mmol) and
[3,5-dibromo-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-
-proprionic acid (22) (800 mg, 1.67 mmol) was heated to 120.degree.
C. for 24 h. The reaction mixture was cooled to room temperature
and concentrated. The resulting residue was diluted with water (200
mL) and was made basic to pH=9 by the addition of a 1N aqueous
sodium hydroxide solution. This solution was extracted with ethyl
acetate (1.times.100 mL) and the ethyl acetate was discarded. The
water layer was acidified to pH=3 by the addition of a 1N aqueous
hydrochloric acid solution. The water layer was extracted with
ethyl acetate (3.times.100 mL). The organic layers were combined,
dried with sodium sulfate, filtered and concentrated under vacuum.
The resulting solid was slurried in ethyl acetate (20 mL). Heptane
(20 mL) was added to this mixture. The solvents were then
concentrated under vacuum to a volume of approximately 20 mL. The
solids were filtered and washed with a 1:1 ethyl acetate:heptane
solution (2.times.10 mL). The solid was dried under high vacuum to
afford
[3,5-dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
proprionic acid (23) (553 mg, 72%) as an off-white solid; LRMS for
C.sub.16H.sub.16Br.sub.2N.sub.2O.sub.4 (M+H) m/z=461. Molecular
Weight=460.1249; Exact Mass=457.9477
##STR00077##
Example 6
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyla-
mino]-acetic acid (26)
##STR00078##
Step 1: Preparation of
5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylamine
(24)
A solution of 3,6-dichloro-4-isopropyl pyridazine (7) (5.4 g, 28
mmol) in anhydrous dimethyl sulfoxide (20 mL) under argon at room
temperature was treated with 4-amino-2,6-dichlorophenol (5.0 g, 28
mmol), anhydrous potassium carbonate (15.6 g, 112 mmol) and copper
(I) iodide (3.2 g, 16.8 mmol). The reaction mixture was heated to
90.degree. C. for 24 h. The reaction mixture was then cooled to
room temperature and poured onto water (1 L). The solution was
brought to pH=8 with a 1N aqueous hydrochloric acid solution. The
water layer was diluted with ethyl acetate (1.times.500 mL), and
the two phases were filtered over celite. The organic layer was
separated. The celite was washed with ethyl acetate. The water
layer was extracted again with ethyl acetate (1.times.500 mL). The
combined organics were then washed with a saturated aqueous sodium
chloride solution (1.times.400 mL), dried with magnesium sulfate,
filtered and concentrated under vacuum. The resulting residue was
dissolved in chloroform and purified by column chromatography using
silica gel eluted with 10-15% ethyl acetate in petroleum ether. The
desired fractions were collected and concentrated under vacuum. The
resulting solid was slurried in ether, filtered and rinsed with
cold ether. The filtrate was concentrated under vacuum. The solid
was diluted with ether and filtered to obtain a second crop. This
contained a trace of the undesired isomer. It was re-slurried in
ether and filtered and shown to be the pure desired isomer by
.sup.1H NMR. The filtrate was concentrated and a third crop was
obtained in the same manner. The filtrate was concentrated and was
diluted with 60% ether in petroleum ether. Petroleum ether was
added with scratching and a solid crystallized. The solid was
collected and rinsed with 60% ether in petroleum ether. The four
crops of solid were shown to be the pure desired isomer by .sup.1H
NMR. The four pure crops of solid were combined and dried under
high vacuum overnight to afford
5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylamine
(24) (4.7 g, 50%) as an off-white solid; EI(+)-HRMS m/z calcd for
C.sub.13H.sub.12Cl.sub.3N.sub.3O (M-H) 331.0046, found 331.0056.
Molecular Weight=332.6191; Exact Mass=331.0046
Step 2:
6-(4-Amino-2,6-dichloro-peony)-4-isopropyl-2H-pyridazin-3-one
(25)
A mixture of glacial acetic acid (30 mL), sodium acetate (860 mg,
10.48 mmol) and
5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylami- ne
(24) (1.0 g, 3.0 mmol) was heated to 10.degree. C. for 24 h. The
reaction mixture was cooled to room temperature, stirred for 2 d
and then was concentrated. The resulting residue was diluted with
water (200 mL) and was made basic to pH=9 by the addition of a 1N
aqueous sodium hydroxide solution. This suspension was extracted
with ethyl acetate (1.times.250 mL). The water layer was acidified
to pH=5 by the addition of concentrated hydrochloric acid. The
water layer was extracted with ethyl acetate (1.times.250 mL). The
organic layers were combined, dried with magnesium sulfate,
filtered and concentrated under vacuum. The resulting oil was
diluted with methanol (20 mL) and was treated with a 1N aqueous
sodium hydroxide solution (20 mL, 20 mmol). The reaction mixture
was heated to 120.degree. C. for 24 h. The reaction mixture was
cooled to room temperature and the solvent was concentrated under
vacuum. The residue was diluted with water (100 mL) and was
extracted with ethyl acetate (200 mL). The ethyl acetate layer was
washed with water containing a 1N aqueous hydrochloric acid
solution (to pH=5) and a saturated aqueous sodium chloride
solution, dried with magnesium sulfate, filtered and concentrated
under vacuum. The residue was dissolved in chloroform and purified
by flash chromatography (Biotage 40 L) using silica gel eluted with
a 1:1 ethyl acetate:hexanes solution with 0.5% glacial acetic acid.
The desired fractions were collected and concentrated under vacuum
and dried under high vacuum at 37.degree. C. The solid was slurried
in diethyl ether (.about.10 mL) and petroleum ether (10 mL). The
solid was stirred 20 min at room temperature, was filtered and
rinsed well with petroleum ether. The solid was dried under high
vacuum to afford
6-(4-amino-2,6-dichloro-peony)-4-isopropyl-2H-pyridazin-3-one (25)
(538 mg, 57%) as an off-white solid. LRMS for
C.sub.13H.sub.13Cl.sub.2N.sub.3O.sub.2 (M.sup.+) m/z=314. Molecular
Weight=314.1735; Exact Mass=313.0385
Step 3: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyla-
mino]-acetic acid (26)
A solution of
6-(4-amino-2,6-dichloro-peony)-4-isopropyl-2H-pyridazin-3-one (25)
(500 mg, 1.59 mmol) in methylene chloride (22 mL) and methanol (22
mL) at room temperature was treated with glyoxylic acid monohydrate
(292 mg, 3.17 mmol), glacial acetic acid (0.10 mL, 1.74 mmol), a
small spatula tip of magnesium sulfate, and resin bound
cyanoborohydride (Argonaut Technologies Inc. MP-(CN) BH.sub.3, 0.97
g, 2.39 mmol). The reaction mixture was heated to 50.degree. C. for
24 h. The reaction mixture was then cooled to room temperature,
filtered through celite, and rinsed with chloroform. A 1N aqueous
hydrochloric acid solution (150 mL) was added to the filtrate. The
resulting mixture was stirred for 30 min. The organic layer was
separated. The water layer was extracted with chloroform (100 mL).
The organic layers were combined, methanol (10 mL) was added to
dissolve insoluble material and the organic layer was dried with
magnesium sulfate, filtered and concentrated under vacuum. The
resulting solid was slurried with ether (3 mL) and diluted with
petroleum ether (10 mL). The mixture was stirred for 45 min. The
solids were collected by filtration and rinsed with petroleum
ether. A small amount of higher Rf impurity was present by TLC. The
solid was purified by flash chromatography (Biotage 40 L) using
silica gel eluted with 100% ethyl acetate followed by 0.4% glacial
acetic acid in ethyl acetate. The desired fractions were collected
and concentrated under vacuum. The solid was diluted with 1:1
methylene chloride: hexanes and concentrated under vacuum. The
solid was slurried in petroleum ether, filtered, rinsed with
petroleum ether and dried under high vacuum overnight to afford
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyla-
mino]-acetic acid (26) (98.7 mg, 17%) as a white solid; LRMS for
C.sub.15H.sub.15Cl.sub.2N.sub.3O.sub.4 (M.sup.+) m/z=372. Molecular
Weight=372.2105; Exact Mass=371.0440
##STR00079##
Example 7
Synthesis of
N-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-oxalamic acid (29)
##STR00080##
Step 1: Preparation of
N-[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-oxalam-
ic acid methyl ester (27)
A solution of
5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylamine
(24) (2.0 g, 6 mmol) in anhydrous tetrahydrofuran (28 mL) at room
temperature was treated with N,N-diisopropylethylamine (2.5 mL, 144
mmol). The reaction mixture was cooled to 0.degree. C. and was then
treated with methyl oxalyl chloride (0.66 mL, 72 mmol). The
reaction mixture was warmed to room temperature and was stirred for
3 h. The reaction mixture was poured onto water (1 L) and was
diluted with ethyl acetate (300 mL) and a saturated aqueous sodium
chloride solution (100 mL). The organic layer was separated. The
water layer was re-extracted with ethyl acetate (2.times.300 mL).
The organic layers were combined and washed with a 1N aqueous
hydrochloric acid solution (1.times.150 mL) and a saturated aqueous
sodium chloride solution (1.times.150 mL), dried with magnesium
sulfate, filtered and concentrated under vacuum. The resulting
solid was dissolved in ethyl acetate with methanol and was purified
by column chromatography on silica eluted with 15-30% ethyl acetate
in petroleum ether. The desired fractions were concentrated under
vacuum. The resulting solid was dried under high vacuum to afford
N-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-oxalam-
ic acid methyl ester (27) (1.91 g, 76%) as a white solid;
EI(+)-HRMS m/z calcd for C.sub.16H.sub.14Cl.sub.3N.sub.3O.sub.4
(M+H) 418.0123, found 418.0123. Molecular Weight=418.6667; Exact
Mass=417.0050
Step 2: Preparation of
N-[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-oxalam-
ic acid (28)
A suspension of
N-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-oxalam-
ic acid methyl ester (27) (1.87 g, 4.47 mmol) in methanol (35 mL)
at 0.degree. C. was treated with a 1N aqueous sodium hydroxide
solution (8.95 mL, 8.95 mmol). The reaction mixture was warmed to
room temperature and stirred for 2 h. The solvent was concentrated
under vacuum. The residue was diluted with water (500 mL), and the
mixture was made acidic to pH=1-2 by the addition of a 1N aqueous
hydrochloric acid solution. The mixture was extracted with ethyl
acetate (2.times.250 mL). The organic layer was separated, dried
with magnesium sulfate, filtered and concentrated. The resulting
solid was dried on high vacuum overnight to afford
N-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-
-oxalamic acid (28) (1.77 g, 98%) as a white solid; EI(+)-HRMS m/z
calcd for C.sub.15H.sub.12Cl.sub.3N.sub.3O.sub.4 (M+H) 403.9966,
found 403.9968. Molecular Weight=404.6396; Exact Mass=402.9893
Step 3: Preparation of
N-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-oxalamic acid (29)
A mixture of glacial acetic acid (55 mL), sodium acetate (1.2 g,
14.7 mmol) and
N-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phen-
yl]-oxalamic acid (28) (1.7 g, 4.2 mmol) was heated to 100.degree.
C. for 24 h. The reaction mixture was cooled to room temperature
and was concentrated under vacuum. The resulting solid was diluted
with water (50 mL). The mixture was made acidic to pH=3 by the
addition of glacial acetic acid. The solids were filtered and
rinsed well with water and dried on the funnel. The solid was
slurried in 1:1:2 isopropyl acetate: methyl tert-butyl ether:
hexanes (3 mL) and then was heated to reflux. The mixture was
cooled and filtered. A slight amount of impurity was detected by
TLC. The solid was slurried in isopropyl acetate (10 mL) and
filtered. The resulting pure solid was dried under high vacuum for
24 h and then dried in a vacuum oven at 80.degree. C. for 24 h to
afford
N-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-oxalamic acid (29) (0.91 g, 56%) as a white solid; EI(+)-HRMS
m/z calcd for C.sub.15H.sub.13Cl.sub.2N.sub.3O.sub.5 (M+H)
386.0305, found 386.0308. Molecular Weight=386.1940; Exact
Mass=385.0232
##STR00081##
Example 8
Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(31)
##STR00082##
Step 1: Preparation of
2-Cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester (30)
A suspension of
5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylamine
(24) (134 mg, 0.42 mmol) in water (5.6 mL) was treated with
concentrated hydrochloric acid (2.8 mL). The reaction mixture was
cooled to 0.degree. C. and then was treated with a solution of
sodium nitrate (36.5 mg, 0.529 mmol) in water (0.2 mL) under the
surface of the reaction mixture followed by a water (0.2 mL) rinse.
The reaction mixture was stirred at 0.degree. C. for 30 min, and a
solution formed. In a separate flask, equipped with a magnetic
stirrer, was added N-cyanoacetylurethane (73 mg, 0.46 mol), water
(9.4 mL) and pyridine (2.8 mL). This reaction mixture was cooled to
0.degree. C. and the solution from the first reaction was quickly
filtered and poured into the second reaction mixture. An orange
precipitate formed and the suspension was stirred at 0.degree. C.
for 30 min. The solid was filtered and rinsed with water followed
by petroleum ether. The solid was dried in a vacuum oven overnight
at 80.degree. C. to afford
2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazi-
n-3-yloxy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(30) (156 mg, 76%) as an orange solid; EI(+)-HRMS m/z calcd for
C.sub.19H.sub.18Cl.sub.2N.sub.6O.sub.5 (M+H) 481.0789, found
481.0790. Molecular Weight=481.2985; Exact Mass=480.0716
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(31)
A mixture of
2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester (30) (3.49
g, 7.17 mmol) in glacial acetic acid (72 mL) was treated with
sodium acetate (2.94 g, 35.8 mmol) at room temperature. The
reaction mixture was heated to 120.degree. C. for 1.5 h. At this
time, the reaction was cooled to 0.degree. C., diluted with water
(220 mL) and stirred for 30 min. The resulting solid was filtered
and rinsed with water (3.times.100 mL) followed by petroleum ether
(3.times.100 mL). The solid was air dried for 30 min. The solid was
then diluted with hot acetonitrile (250 mL). The resulting red
mixture was treated with neutral decolorizing carbon, filtered
through celite and rinsed with acetonitrile (1 L) until no UV
active material eluted. The yellow filtrate was concentrated under
reduced pressure. The resulting solid was triturated with hot
acetonitrile (50 mL), cooled for 15 min, diluted with water (100
mL) and filtered. The solid was triturated again with hot
acetonitrile (10 mL), filtered and rinsed with acetonitrile, water,
and petroleum ether. The solids were collected and dried under high
vacuum overnight and then dried in a vacuum oven at 80.degree. C.
overnight to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile (31)
(1.91 g, 61%) as a yellow solid; EI(+)-HRMS m/z calcd for
C.sub.17H.sub.12Cl.sub.2N.sub.6O.sub.4 (M+H).sup.+ 435.0370, found
435.0368. Molecular Weight=435.2290; Exact Mass=434.0297
##STR00083##
Example 9
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-2H-[1,2,4]triazine-3,5-dione (33)
##STR00084##
Step 1: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic acid
(32)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile (31)
(136 mg, 0.312 mmol) in glacial acetic acid (3.0 mL) was treated
with concentrated hydrochloric acid (0.345 mL). The reaction
mixture was heated to 120.degree. C. for 24 h. Starting material
was still present by LC/MS. Additional concentrated hydrochloric
acid (0.34 mL) was added. The reaction mixture was heated to
120.degree. C. for another 24 h. The reaction mixture was cooled to
room temperature and was diluted with water (50 mL). At this time,
the reaction was made basic by the addition of a 1N aqueous sodium
hydroxide solution and was extracted with ether (100 mL). The
organic layer was discarded. The aqueous layer was acidified by the
addition of a 1N aqueous hydrochloric acid solution and was
extracted with ethyl acetate (2.times.100 mL). The organic layers
were combined, washed with a saturated aqueous sodium chloride
solution, dried with magnesium sulfate, filtered and concentrated
under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (32) (111 mg, 78%) as an orange solid which was used without
further purification; LRMS for
C.sub.17H.sub.13Cl.sub.2N.sub.5O.sub.6 (M m/z=454. Molecular
Weight=454.2291; Exact Mass=453.0243
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-2H-[1,2,4]triazine-3,5-dione (33)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic acid
(32) (102 mg, 0.22 mmol) and mercaptoacetic acid (2.2 mL) was
heated to 170.degree. C. for 1 h. At this time, the reaction
mixture was cooled to room temperature and was diluted with water
(120 mL) and was extracted with ethyl acetate (100 mL). The organic
layer was separated and washed with a saturated aqueous sodium
bicarbonate solution (2.times.100 mL) and a saturated aqueous
sodium chloride solution (100 mL), dried with magnesium sulfate,
filtered and concentrated under vacuum. The resulting solid was
dissolved in methylene chloride and was purified by flash
chromatography (Biotage 40S) using silica gel eluted with 100%
ethyl acetate to elute the impurity followed by 0.2% glacial acetic
acid in ethyl acetate to elute the desired product. The desired
fractions were collected and concentrated under vacuum. The
resulting solid was slurried in hot methanol (2 mL), filtered and
rinsed with petroleum ether. The solids were dried in a vacuum oven
at 80.degree. C. to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-2H-[1,2,4]triazine-3,5-dione (33) (21.4 mg, 23%) as a yellow
solid; EI(+)-HRMS m/z calcd for
C.sub.16H.sub.13Cl.sub.2N.sub.5O.sub.4 (M.sup.+) 410.0418, found
410.0419. Molecular Weight=410.2191; Exact Mass=409.0345
##STR00085## ##STR00086##
Example 10
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-acetic acid (37)
##STR00087##
Step 1: Preparation of Acetic acid
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-ethyl ester (34)
A solution of
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17) (500 mg, 1.38 mmol) in glacial acetic acid (5 mL) was
treated with sodium acetate (230 mg, 2.8 mmol) and heated to
114.degree. C. for 24 h. At this time, the reaction mixture was
cooled to room temperature and was concentrated under vacuum. The
resulting residue was diluted with methylene chloride (25 mL) and
was washed with water (10 mL). The organic layer was separated and
was washed with a saturated aqueous sodium bicarbonate solution (10
mL). The waters layers were combined and extracted with methylene
chloride (10 mL). The organic layers were combined and dried with
sodium sulfate, filtered, concentrated and dried under high vacuum
overnight to afford acetic acid
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-ethyl ester (34) (531 mg, 100%) as a white solid; LRMS for
C.sub.17H.sub.18Cl.sub.2N.sub.2O.sub.4 (M.sup.+) m/z=385 Molecular
Weight=385.2500; Exact Mass=384.0644
Step 2: Preparation of acetic acid
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-ethyl ester (35)
A mixture of acetic acid
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-ethyl ester (34) (531 mg, 1.38 mmol), potassium carbonate (280
mg, 2.03 mmol) and methyl iodide (2 mL, 32.1 mmol) was warmed to
40.degree. C. for 2 h. Additional methyl iodide was added (1 mL,
16.05 mmol) followed by potassium carbonate (140 mg, 1.01 mmol).
The mixture was heated at 40.degree. C. for 2 h and then was
stirred at room temperature for 24 h. The reaction mixture was
concentrated and the resulting residue was partitioned between
ethyl acetate (50 mL) and water (25 mL). The water layer was
re-extracted with ethyl acetate (25 mL). The organic layers were
combined, dried with sodium sulfate, filtered, and concentrated.
The resulting residue was dissolved in 25% ethyl acetate in hexanes
and was purified by flash chromatography (Biotage 40S) using silica
gel eluted with 25-50% ethyl acetate in hexanes. The desired
fractions were collected and concentrated under vacuum. The
resulting solid was dried under high vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]ethyl ester (35) (395 mg, 72%) as a clear oil; LRMS for
C.sub.18H.sub.20Cl.sub.2N.sub.2O.sub.4 (M.sup.+) m/z=399. Molecular
Weight=399.2771; Exact Mass=398.0800
Step 3: Preparation of
6-[2,6-Dichloro-4-(2-hydroxyethyl)-phenoxy]-4-isopropyl-2-methyl-pyridazi-
n-3-one (36)
A solution of
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-ethyl ester (35) (390 mg, 0.97 mmol) in methanol (4 mL)
was treated with a 1N aqueous sodium hydroxide solution (1.0 mL,
1.0 mmol) at room temperature. The reaction mixture was stirred for
24 h and was then concentrated under vacuum. The resulting residue
was diluted with methylene chloride (35 mL) and was washed with a
saturated aqueous sodium chloride solution (20 mL). The aqueous
layer was separated and was re-extracted with methylene chloride
(25 mL). The organic layers were combined and dried with sodium
sulfate, filtered, and concentrated. The resulting solid was dried
under high vacuum to afford
6-[2,6-dichloro-4-(2-hydroxy-ethyl)-phenoxy]-4-isopropyl-2-methyl-pyridaz-
in-3-one (36) (350 mg, 97%) as a white solid that was used without
further purification. Molecular Weight=357.2395; Exact
Mass=356.0694
Step 4: Preparation of
[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-acetic acid (37)
A solution of
6-[2,6-dichloro-4-(2-hydroxy-ethyl)-phenoxy]-4-isopropyl-2-methyl-pyridaz-
in-3-one (36) (330 mg, 0.92 mmol) in acetone (5 mL) was treated
with Jones Reagent (0.51 mL of a 2.7M solution) slowly dropwise at
10.degree. C. The resulting red reaction mixture was stirred at
11.degree. C. for 30 min. The reaction mixture was then
concentrated under vacuum. The resulting residue was diluted with
ethyl acetate (25 mL) and water (20 mL) and was treated with sodium
bisulfite (100 mg). The resulting mixture was shaken and turned
from red to green. The aqueous layer was separated and was
re-extracted with ethyl acetate (25 mL). The organic layers were
combined and washed with a saturated aqueous sodium chloride
solution. The organic layer was separated, dried with sodium
sulfate, filtered, and concentrated under vacuum. The resulting
solid was recrystallized from ethyl acetate and dried under vacuum
overnight to afford
[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-acetic acid (37) (70 mg, 19%) as a white solid;
EI(+)-HRMS) m/z calcd for C.sub.16H.sub.16Cl.sub.2N.sub.2O.sub.4
(M+H).sup.+ 371.0560, found 371.0561. Molecular Weight=371.2229;
Exact Mass=370.0487
##STR00088## ##STR00089##
Example 11
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-acetic acid (42)
##STR00090##
Step 1: Preparation of
2-[3,5-Dibromo-4-(6-chloro-pyridazin-3-yloxy)-phenyl]-ethanol
(38)
This compound was prepared by a similar method to that described in
Example 3, Step 2 except that 3,6-dichloro-pyridazine was used in
place of 3,6-dichloro-4-isopropyl pyridazine (7). The reaction was
heated at 140.degree. C. for 3.5 h. The work up was different than
Example 3, Step 2. The reaction mixture was added slowly with
stirring to a saturated aqueous sodium chloride solution. The
mixture was extracted with methylene chloride. The organic layers
were combined and washed with a saturated aqueous sodium chloride
solution, dried over sodium sulfate, filtered, and concentrated.
The resulting oil was dissolved in methylene chloride and was
purified by flash chromatography (Biotage) using silica gel eluted
with 10-40% ethyl acetate in hexanes. The desired fractions were
collected and concentrated under vacuum to afford an oil which was
dried under high vacuum to afford
2-[3,5-dibromo-4-(6-chloro-pyridazin-3-yloxy)-phenyl]-ethanol (38)
(51%) as an amber oil; LRMS for
C.sub.12H.sub.9Br.sub.2ClN.sub.2O.sub.2 (M.sup.+) m/z=409.
Molecular Weight=408.4787; Exact Mass=405.8719
Step 2: Preparation of Acetic acid
2-[3,5-dibromo-4-(6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-ethyl
ester (39)
This compound was prepared by a similar method to that described in
Example 10, Step 1 except that
2-[3,5-dibromo-4-(6-chloro-pyridazin-3-yloxy)-phenyl]-ethanol (38)
was used in place of
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenyl]-ethano-
l (17) to afford acetic acid
2-[3,5-dibromo-4-(6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-ethyl
ester (39) (92%) as a colorless oil; LRMS for
C.sub.14H.sub.12Br.sub.2N.sub.2O.sub.4 (M+H) m/z=433. Molecular
Weight=432.0707; Exact Mass=429.9164
Step 3: Preparation of acetic acid
2-[3,5-dibromo-4-(1-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-ethyl ester (40)
This compound was prepared by a similar method to that described in
Example 10, Step 2 except that isopropyl iodide was used in place
of methyl iodide and acetic acid
2-[3,5-dibromo-4-(6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-ethyl
ester (39) was used in place of acetic acid
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-ethyl ester (34). The method was similar except that the
reaction was heated to 50.degree. C. for 24 h. The product was
purified by column chromatography using silica gel eluted with 10%
ethyl acetate in hexanes to 100% ethyl acetate to afford
2-[3,5-dibromo-4-(1-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-ethyl ester (40) (79%) as a white solid; LRMS for
C.sub.17H.sub.18Br.sub.2N.sub.2O.sub.4 (M+CH.sub.3CN+H) m/z=516,
Molecular Weight=474.1520; Exact Mass=471.9633
Step 4: Preparation of
6-[2,6-Dibromo-4-(2-hydroxy-ethyl)-phenoxy]-2-isopropyl-pyridazin-3-one
(41)
A solution of
2-[3,5-dibromo-4-(1-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl-
]-ethyl ester (40) (85 mg, 0.18 mmol) in methanol (0.5 mL) was
treated with a 2N aqueous potassium hydroxide solution (90 .mu.L,
0.18 mmol) at room temperature. The reaction mixture was heated to
55.degree. C., stirred for 1 h and then was concentrated under
reduced pressure. The resulting residue was diluted with methylene
chloride (5 mL) and was washed with water (2.times.3 mL). The
aqueous layers were re-extracted with methylene chloride (5 mL).
The organic layers were combined and dried with sodium sulfate,
filtered, and concentrated. The resulting solid was dried under
high vacuum to afford
6-[2,6-dibromo-4-(2-hydroxy-ethyl)-phenoxy]-2-isopropyl-pyridazin-3-one
(41) (77 mg, 100%) as a white solid that was used without further
purification; LRMS for C.sub.15H.sub.16Br.sub.2N.sub.2O.sub.3
(M+CH.sub.3CN+H) m/z=474. Molecular Weight=432.1144; Exact
Mass=429.9528
Step 5: Preparation of
[3,5-Dibromo-4-(1-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
acetic acid (42)
This compound was prepared by a similar method to that described in
Example 10, Step 4 except that
6-[2,6-dibromo-4-(2-hydroxy-ethyl)-phenoxy]-2-isopropyl-pyridazin-3-one
(41) was used in place of
6-[2,6-dichloro-4-(2-hydroxy-ethyl)-phenoxy]-4-isopropyl-2-methyl-pyridaz-
in-3-one (36). The product was purified by flash chromatography
(Biotage) using silica gel eluted with 50% ethyl acetate in
hexanes. The desired fractions were collected and concentrated
under vacuum to afford a solid which was dried under high vacuum to
afford
[3,5-dibromo-4-(1-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-phenyl]--
acetic acid (42) as an off-white solid (40 mg, 52%); EI(+)-HRMS m/z
calcd for C.sub.15H.sub.14Br.sub.2N.sub.2O.sub.4 (M+H).sup.+
444.9393, found 444.9392. Molecular Weight=446.0978; Exact
Mass=443.9320
##STR00091## ##STR00092##
Example 12
Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(48)
##STR00093##
Step 1: Preparation of Cyano-(2,6-dichloro-4-nitrophenyl)-acetic
acid tert-butyl ester (43)
A solution of 1,2,3-trichloro-5-nitrobenzene (50.33 g, 222.26 mmol)
in N,N-dimethylformamide (220 mL) at 25.degree. C. was treated with
2-butylcyanoacetate (33 mL, 230.72 mmol) and potassium carbonate
(61.44 g, 444.54 mmol). The reaction flask was fitted with a reflux
condenser. It was then heated to 50.degree. C. for 18 h. At this
time, the reaction was allowed to cool to 25.degree. C. and then
was concentrated under vacuum. The reaction mixture was dissolved
in ethyl acetate (1.2 L) and water (300 mL). This bilayer was
neutralized by the addition of a 1N aqueous hydrochloric acid
solution (250 mL) and a 3N aqueous hydrochloric acid solution (300
mL). The resulting layers were separated. The organics were washed
with water (3.times.500 mL) and a saturated aqueous sodium chloride
solution (250 mL), dried with magnesium sulfate, filtered, and
concentrated under vacuum to give
cyano-(2,6-dichloro-4-nitro-phenyl)-acetic acid tert-butyl ester
(43) (78.69 g) as a golden brown oil. The compound was used without
further purification. Exact Mass=330.0174; Molecular
Weight=331.16.
Step 2: Preparation of (4-amino-2,6-dichloro-phenyl)-acetonitrile
(44)
A solution of cyano-(2,6-dichloro-4-nitro-phenyl)-acetic acid
tert-butyl ester (43) (78.69 g, crude) in ethanol (320 mL) at
25.degree. C. was treated with concentrated hydrochloric acid (160
mL). The reaction was fitted with a reflux condenser and then was
heated to 75.degree. C. At this time, the resulting homogeneous
solution was treated portion wise with tin(II) chloride dihydrate
(225.64 g, 1.0 mol). Upon complete addition of the tin(II) chloride
dihydrate, the reaction was warmed to 110-115.degree. C. where it
was stirred for 3 h. At this time, the reaction was cooled to
25.degree. C. and then was diluted with ethyl acetate (1.5 L). The
resulting solution was carefully neutralized with a saturated
aqueous sodium carbonate solution (1.0 L). The resulting thick
mixture was filtered through filter paper via vacuum filtration and
was washed with ethyl acetate until no material was detected in the
filtrate. The filtrate was then transferred to a separatory funnel
where the resulting layers were separated. The organics were washed
consecutively with a 1N aqueous hydrochloric acid solution
(1.times.250 mL), a saturated aqueous sodium carbonate solution
(1.times.250 mL), a 1N aqueous hydrochloric acid solution
(1.times.250 mL), and a saturated aqueous sodium chloride solution
(1.times.250 mL). The organics were then dried with magnesium
sulfate, filtered, and concentrated under vacuum. Flash
chromatography (Merck Silica gel 60, 230-400 mesh, 80/20
hexanes/ethyl acetate) afforded
(4-amino-2,6-dichloro-phenyl)-acetonitrile (44) (25.95 g, 58%) as a
yellow solid; EL-HRMS m/e calcd for C.sub.8H.sub.6Cl.sub.2N.sub.2
(M.sup.+) 199.9908, found 199.9906. Exact Mass=199.9908; Molecular
Weight=201.06.
Step 3: Preparation of
(4-Amino-2,6-dichloro-phenyl)-(6-chloro-5-isopropyl-pyridazin-3-yl)-aceto-
nitrile (45)
A solution of 3,6-dichloro-4-isopropyl pyridazine (7) (5.54 g, 29.0
mmol) in tetrahydrofuran (116 mL) in a 500 mL round bottom flask
(caution: use an extra large flask) was treated with
(4-amino-2,6-dichloro-phenyl)-acetonitrile (44) (5.81 g, 28.9
mmol). The reaction mixture was equipped with a cold water
condenser and heated to 60.degree. C. The flask was then raised out
of the oil bath and potassium tert-butoxide (6.85 g, 57.99 mmol)
was added. The mixture was heated to 60.degree. C. for 45 min. The
reaction was cooled to room temperature, transferred to a
separatory funnel, diluted with ethyl acetate (500 mL) and was
washed with a saturated aqueous sodium chloride solution
(2.times.250 mL). The organic layer was separated, dried with
magnesium sulfate, and was filtered. Silica gel 60 (70-230 mesh)
was added to the filtrate and the solvent was concentrated under
vacuum. The resulting mixture was purified by flash chromatography
(Biotage 75 L) using silica gel eluted with 15%-30% ethyl acetate
in hexanes. The desired fractions were collected and concentrated
under vacuum to afford a solid which was dried under high vacuum to
afford
(4-amino-2,6-dichloro-phenyl)-(6-chloro-5-isopropyl-pyridazin-3-yl)-aceto-
nitrile (45) (7.87 g, 77%) as an orange foam; LRMS for
C.sub.15H.sub.13Cl.sub.3N.sub.4 (M+H) m/z=355. This compound was
used without further purification. Molecular Weight=355.6567; Exact
Mass=354.0206
Step 4: Preparation of
6-(4-Amino-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(46)
A mixture of
(4-amino-2,6-dichloro-phenyl)-(6-chloro-5-isopropyl-pyridazin-3-yl)-aceto-
nitrile (45) (6.98 g, 19.63 mmol), water (30 mL), concentrated
hydrochloric acid (120 mL) and glacial acetic acid (30 mL) was
heated to 120.degree. C. for 24 h. At this time, the reaction was
cooled to room temperature and the mixture was poured onto water
(250 mL). The pH was made neutral (pH=7) by the addition of a 4N
aqueous sodium hydroxide solution. The suspension was placed in the
freezer for 15 min. The resulting solids were filtered and washed
with water and petroleum ether. The solids were collected and
dissolved in hot ethyl acetate. Silica gel 60 (70-230 mesh) was
added and the solvent was concentrated under vacuum. The resulting
mixture was purified by flash chromatography (Biotage 75S) using
silica gel eluted with 40% ethyl acetate in hexanes to 50% ethyl
acetate in hexanes containing 0.5% glacial acetic acid. The desired
fractions were collected and concentrated under vacuum to afford a
solid which was dried under high vacuum to afford
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one (46)
(4.65 g, 76%) as an off-white solid; LRMS for
C.sub.14H.sub.15Cl.sub.2N.sub.3O (M+H) m/z=312. Molecular
Weight=312.2012; Exact Mass=311.0592
Step 5: Preparation of
(2-Cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yl-
methyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(47)
This compound was prepared by a similar method to that described in
Example 8, Step 1 except that
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one (46)
was used in place of
5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-yloxy)-phenylamine
(25) to afford
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridaz-
in-3-yl-methyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl
ester (47) as an orange solid (3.44 g, 75%); EI(+)-HRMS m/z calcd
for C.sub.20H.sub.20Cl.sub.2N.sub.6O.sub.4 (M+H)1+ 479.0996, found
479.0997. Molecular Weight=479.3262; Exact Mass=478.0923
Step 6: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(48)
This compound was prepared by a similar method to that described in
Example 8, Step 2 except that
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yl-
-methyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester (47)
was used in place of
2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester (30) to
afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(48) as an off-white solid (1.21 g, 39%); EI(+)-HRMS m/z calcd for
C.sub.18H.sub.14Cl.sub.2N.sub.6O.sub.3 (M+H)1+ 433.0577, found
433.0577. Molecular Weight=433.2567; Exact Mass=432.0504
##STR00094##
Example 13
Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-[1,2,4]triazine-3,5-dione (50)
##STR00095##
Step 1: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (49)
This compound was prepared by a similar method to that described in
Example 9, Step 1 except that
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(48) was used in place of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(31). In addition, when the reaction was complete, the reaction
mixture was diluted with water and the resulting solids were
filtered and rinsed well with water followed by petroleum ether.
The solids were dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (49) (81%) as an off-white solid; LRMS for
C.sub.18H.sub.15Cl.sub.2N.sub.5O.sub.5 (M+H) m/z=452. Molecular
Weight=452.2568; Exact Mass=451.0450
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-[1,2,4]triazine-3,5-dione (50)
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (49) (101.5 mg, 0.224 mmol) was treated with mercaptoacetic
acid (2.2 mL). The reaction mixture was stirred with a magnetic
stirrer and was heated to 155.degree. C. for 24 h. The reaction
mixture was then cooled to room temperature, diluted with water (25
mL) and extracted with ethyl acetate (25 mL). The organic layer was
separated and washed with a 1N aqueous sodium hydroxide solution
(3.times.10 mL). The aqueous layers were combined and acidified to
pH=4. The resulting solids were filtered and rinsed well with water
followed by petroleum ether. The solids were dried under vacuum to
afford
dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-phenyl]-[-
1,2,4]triazine-3,5-dione (50) (36 mg, 40%) as a tan solid;
EI(+)-HRMS m/z calcd for C.sub.17H.sub.15Cl.sub.2N.sub.5O.sub.3
(M+H)1+ 408.0625, found 408.0626. Molecular Weight=408.2468; Exact
Mass=407.0552
##STR00096## ##STR00097##
Example 14
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-phen-
yl]-acetic acid (56)
##STR00098##
Step 1: Preparation of
6-(4-bromo-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(51)
A solution of
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one (46)
(0.9 g, 2.88 mmol) in glacial acetic acid (16 mL) at room
temperature was treated with concentrated sulfuric acid (4 mL). A
solution of sodium nitrite (480 mg, 6.96 mmol) in water (5 mL) was
added below the surface of the reaction slowly over 10 min. The
reaction mixture was heated to 60.degree. C. for 1 h. The reaction
was cooled to room temperature and a mixture of copper(I) bromide
(450 mg, 3.14 mmol) and 48% hydrogen bromide in water (2 mL, 17.68
mmol) was added dropwise. The reaction was heated to 100.degree. C.
where vigorous gas evolution occurred. After 1 h, the reaction was
cooled to room temperature, poured onto ice water (100 mL) and was
extracted with ether (3.times.75 mL). The ether layer was
cautiously washed with a saturated aqueous sodium bicarbonate
solution (150 mL). The organic layer was separated, dried with
magnesium sulfate, filtered and concentrated under vacuum. The
resulting solid was purified by flash chromatography using silica
gel eluted with a gradient of 4:1 to 2:1 ethyl acetate:hexanes. The
desired fractions were collected and concentrated under vacuum to
afford a solid which was dried under high vacuum to afford
6-(4-bromo-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one (51)
(553 mg, 51%) as an off-white solid; LRMS-ES(+) for
C.sub.14H.sub.13BrCl.sub.2N.sub.2O (M+H) m/z=375. MW=376.0825,
Exact Mass=373.9588
Step 2: Preparation of
3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-benzo-
ic acid methyl ester (52)
A solution of
6-(4-bromo-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one (51)
(194 mg, 0.516 mmol) in acetonitrile (10 mL) and methanol (2 mL) at
room temperature was treated with palladium(II) acetate (23.7 mg,
0.106 mmol), 1,3-bis(diphenylphosphino)propane (45.7 mg, 0.111
mmol) and triethylamine (360 .mu.L, 2.58 mmol). The sealed tube was
then pressurized to 45 psi with carbon monoxide and heated to
90.degree. C. for 1 h. The reaction was cooled to room temperature,
the pressure was released and a TLC was taken of the reaction
mixture which indicated that starting material was still present.
Additional palladium(II) acetate (10 mg) and
1,3-bis(diphenylphosphino)propane (20 mg) were added to the
reaction mixture. The sealed tube was then pressurized to 45 psi
with carbon monoxide and heated to 90.degree. C. for another 1.5 h.
The reaction was cooled to room temperature, the pressure was
released and the reaction mixture was diluted with ethyl acetate
(100 mL) and was washed with 1:1 solution of water (30 mL) and a
saturated aqueous sodium chloride solution (30 mL). The organic
layer was separated, dried with magnesium sulfate, filtered and
concentrated. The resulting residue was purified by flash
chromatography using silica gel eluted with a gradient of 4:1 to
1:1 ethyl acetate:hexanes. The desired fractions were collected and
concentrated under vacuum to afford a solid which was dried under
high vacuum to afford
3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-benzo-
ic acid methyl ester (52) (183 mg, 90%) as an off-white solid;
LRMS-ES(+) for C.sub.16H.sub.16Cl.sub.2N.sub.2O.sub.3 (M+H)
m/z=355. MW=355.2235, Exact Mass=354.0538
Step 3: Preparation of
6-(2,6-Dichloro-4-hydroxymethyl-benzyl)-4-isopropyl-pyridazin-3-one
(53)
A mixture of
3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-benzo-
ic acid methyl ester (52) (160 mg, 0.45 mmol) in tetrahydrofuran (5
mL) at 25.degree. C. was treated with a 1M solution of
diisobutylaluminum hydride in tetrahydrofuran (2.7 mL, 2.7 mmol).
The reaction was stirred at room temperature for 24 h. A TLC of the
reaction indicated that starting material was still present. An
additional amount of the 1 M solution of diisobutylaluminum hydride
in tetrahydrofuran (1.0 mL) was added to the reaction. After
stirring 30 min at room temperature, the reaction was cautiously
quenched by addition to a 1:1 mixture of a saturated aqueous sodium
chloride solution (30 mL) and a 2N aqueous hydrochloric acid
solution (30 mL). The mixture was extracted with ethyl acetate
(3.times.50 mL). The organic layers were combined, washed with a
saturated aqueous sodium chloride solution (20 mL), dried with
magnesium sulfate, filtered and concentrated under vacuum. The
resulting solid was dried under high vacuum to afford
6-(2,6-dichloro-4-hydroxymethyl-benzyl)-4-isopropyl-pyridazin-3-one
(53) (168 mg) as a brown solid that was used without further
purification; LRMS-ES(+) for C.sub.15H.sub.16Cl.sub.2N.sub.2O.sub.2
(M+H) m/z=327. MW=327.2130, Exact Mass=326.0589
Step 4: Preparation of
6-(4-Bromomethyl-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(54)
A solution of
6-(2,6-dichloro-4-hydroxymethyl-benzyl)-4-isopropyl-pyridazin-3-one
(53)
(165 mg, theoretically 0.45 mmol) in methylene chloride (2 mL) at
0.degree. C. was treated with carbon tetrabromide (187 mg, 0.56
mmol) and a solution of triphenylphosphine (178 mg, 0.68 mmol) in
methylene chloride (2 mL). The reaction mixture was stirred at
0.degree. C. for 30 min and then was quenched with water. The
reaction was diluted with water (20 mL) and extracted with ethyl
acetate (3.times.50 mL). The organic layers were combined, dried
with magnesium sulfate, filtered and concentrated under vacuum. The
resulting oil was purified by flash chromatography using silica gel
eluted with 20%-45% ethyl acetate in hexanes. The desired fractions
were collected and concentrated under vacuum to afford a yellow
solid which was dried under high vacuum to afford
6-(4-bromomethyl-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(54) (70 mg, 36%) as a yellow solid; LRMS-ES(+) for
C.sub.15H.sub.15BrCl.sub.2N.sub.2O (M+H) m/z=389. MW=390.1096,
Exact Mass=387.9745
Step 5: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-phen-
yl]-acetonitrile (55)
A suspension of sodium cyanide (270 mg, 5.51 mmol) in dimethyl
sulfoxide (2 mL) at room temperature was treated with concentrated
sulfuric acid (0.1 mL, 1.88 mmol) and a solution of
6-(4-bromomethyl-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(54) (70 mg, 0.180 mmol) in dimethyl sulfoxide (2 mL). The reaction
mixture was stirred at room temperature for 30 min and then at
50.degree. C. for 1 h. The reaction was cooled to room temperature,
poured into a saturated aqueous sodium bicarbonate solution (50
mL), was diluted with water (20 mL) and then extracted with ethyl
acetate (3.times.75 mL). The organic layers were combined and
washed with a saturated aqueous sodium chloride solution (50 mL),
dried over magnesium sulfate, filtered, and concentrated under
vacuum. The resulting oil was dried under high vacuum to afford
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylme-
thyl)-phenyl]-acetonitrile (55) as an orange oil (assume 0.180
mmol) which was used without further purification.
Step 6: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-phen-
yl]-acetic acid (56)
A mixture of
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-phen-
yl]-acetonitrile (55) (assume 0.180 mmol) in concentrated
hydrochloric acid (4 mL) was heated to reflux for 24 h. The
reaction mixture was cooled to room temperature, diluted with ethyl
acetate (75 mL) and washed with a saturated aqueous sodium chloride
solution (20 mL) and water (10 mL). The water layers were combined
and extracted with ethyl acetate (50 mL). The organic layers were
combined, dried over magnesium sulfate, filtered, and concentrated
under vacuum. The resulting oil was purified by HPLC
(acetonitrile/water with 0.1% trifluoroacetic acid). The desired
fractions were collected, concentrated and freeze dried. The
resulting solid was purified by flash chromatography using silica
gel eluted with 10% methanol in methylene chloride. The desired
fractions were collected and concentrated under vacuum to afford a
solid which was dried under high vacuum to afford
3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetic acid (56) (8.6 mg, 13% for 2 steps) as a white solid;
EI(+)-HRMS m/z calcd for C.sub.16H.sub.16Cl.sub.2N.sub.2O.sub.3
(M+H).sup.+ 355.0611, found 355.0611. MW=355.2235, Exact
Mass=354.0538
##STR00099## ##STR00100##
Example 15
Synthesis of
[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetic acid (64)
##STR00101##
Step 1: Preparation of 3,5-Dibromo-4-bromomethyl-benzoic acid
methyl ester (57)
A solution of methyl 3,5-dibromo-4-methylbenzoate (5 g, 16.24 mmol)
in carbon tetrachloride (50 mL) was treated with N-bromosuccinamide
(3.6 g, 20.23 mmol) and 2,2'-azobisisobutyronitrile (0.56 g, 3.410
mmol). The reaction mixture was heated to reflux for 24 h. The
reaction was cooled to room temperature and concentrated under
vacuum. The resulting mixture was absorbed onto silica and was
purified by flash chromatography using silica gel eluted with 3:1
hexanes: ethyl acetate. The desired fractions were collected and
concentrated under vacuum to afford a solid which was dried under
high vacuum to afford 3,5-dibromo-4-bromomethyl-benzoic acid methyl
ester (57) (6.22 g, 99%) as a yellow solid; LRMS for
C.sub.9H.sub.7Br.sub.3O.sub.2 (M+Na) m/z=407. MW=386.8669, Exact
Mass=383.7996
Step 2: Preparation of (3,5-Dibromo-4-bromomethyl-phenyl)-methanol
(58)
A solution of 3,5-dibromo-4-bromomethyl-benzoic acid methyl ester
(57) (5.2 g, 13.44 mmol) in tetrahydrofuran (50 mL) at 0.degree. C.
was treated with a 1M solution of diisobutylaluminum hydride in
tetrahydrofuran (30 mL, 30 mmol). The reaction mixture was stirred
for 2 h at 0.degree. C. TLC revealed starting material was still
present. Additional 1M solution of diisobutylaluminum hydride in
tetrahydrofuran (16 mL, 16 mmol) was added at 0.degree. C. and the
reaction mixture was stirred for 30 min at 0.degree. C. The
reaction was quenched cautiously by pouring it onto a mixture of
ice and concentrated hydrochloric acid (100 mL) and the mixture was
extracted with ethyl acetate (3.times.150 mL). The organic layers
were combined, dried with magnesium sulfate, filtered, and
concentrated under vacuum to afford a solid which was dried under
high vacuum to afford (3,5-dibromo-4-bromomethyl-phenyl)-methanol
(58) (4.56 g, 94%) as an off-white solid; LRMS-EI(+) for
C.sub.8H.sub.7Br.sub.3O (M.sup.+) m/z=356. MW=358.8564, Exact
Mass=355.8047
Step 3: Preparation of
(2,6-Dibromo-4-hydroxymethyl-phenyl)-acetonitrile (59)
This compound was prepared by a similar method to that described in
Example 14, Step 5 except that
(3,5-dibromo-4-bromomethyl-phenyl)-methanol (58) was used in place
6-(4-bromomethyl-2,6-dichloro-benzyl)-4-isopropyl-pyridazin-3-one
(54). The product was purified by flash chromatography using silica
gel eluted with a gradient of 3:1 to 1:1 hexanes:ethyl acetate. The
desired fractions were collected and concentrated under vacuum to
afford a solid which was dried under high vacuum to afford
2,6-dibromo-4-hydroxymethyl-phenyl)-acetonitrile (59) (78%) as a
yellow solid; EI(+)-HRMS m/z calcd for C.sub.9H.sub.7Br.sub.2NO (M
302.8894, found 302.8881. MW=304.9702, Exact Mass=302.8894
Step 4: Preparation of
[2,6-Dibromo-4-(tetrahydro-pyran-2-yloxymethyl)-phenyl]-acetonitrile
(60)
A solution of 2,6-dibromo-4-hydroxymethyl-phenyl)-acetonitrile (59)
(1.4 g, 4.59 mmol) in methylene chloride (30 mL) at room
temperature was treated with 3,4-dihydro-2H-pyran (0.46 mL, 5.04
mmol) and p-toluenesulfonic acid monohydrate (16.10 mg, 0.085
mmol). The reaction mixture was stirred for 45 min and was then
treated with a saturated aqueous sodium bicarbonate solution (3
mL), a saturated aqueous sodium chloride solution (10 mL) and water
(10 mL). This mixture was extracted with methylene chloride
(3.times.50 mL). The organic layers were combined and washed with a
saturated aqueous sodium chloride solution (30 mL), dried with
magnesium sulfate, filtered and concentrated under vacuum. The
resulting oil was purified by flash chromatography (Isco 120 g
column) using silica gel eluted with 15% ethyl acetate in hexanes.
The desired fractions were collected and concentrated under vacuum
to afford an oil which was dried under high vacuum to afford of
[2,6-dibromo-4-(tetrahydro-pyran-2-yloxymethyl)-phenyl]-acetonitrile
(60) as a pale yellow oil (1.52 g, 85.4%). Used without further
purification. Molecular Weight=389.0892; Exact Mass=386.9470
Step 5: Preparation of
[2,6-Dibromo-4-(tetrahydro-pyran-2-yloxymethyl)-phenyl]-acetonitrile
(61)
A solution of
[2,6-dibromo-4-(tetrahydro-pyran-2-yloxymethyl)-phenyl]-acetonitrile
(60) (1.52 g, 3.91 mmol) in N,N-dimethylformamide (9 mL) at
25.degree. C. was treated with sodium hydride (192.8 mg, 4.82 mmol,
60% dispersion in mineral oil). The reaction mixture was stirred at
25.degree. C. for 10 min. At this time, the reaction was treated
with a solution of 3,6-dichloro-4-isopropyl-pyridazine (7) (810 mg,
4.24 mmol) in N,N-dimethylformamide (2 mL). The reaction was then
heated to 90.degree. C. for 1 h. At this time, the reaction was
cooled to 25.degree. C., diluted with water (50 mL), a saturated
aqueous sodium chloride solution (50 mL) and a saturated aqueous
sodium bicarbonate solution (20 mL). This mixture was extracted
with ethyl acetate (200 mL). The organics were washed with a
saturated aqueous sodium chloride solution (100 mL), dried with
magnesium sulfate, filtered, and concentrated under vacuum. Isco
chromatography (120 g, Silica, 2:1 hexanes/ethyl acetate) afforded
(6-chloro-5-isopropyl-pyridazin-3-yl)-[2,6-dibromo-4-(tetrahydro-pyran-2--
yloxymethyl)-phenyl]-acetonitrile (61) (964 mg, 65%) as an orange
viscous oil; LRMS for C.sub.21H.sub.22Br.sub.2ClN.sub.3O.sub.2
(M+H) at m/z=542. Molecular Weight=543.6894; Exact
Mass=540.9767
Step 6: Preparation of
6-(2,6-Dibromo-4-chloromethyl-benzyl)-4-isopropyl-pyridazin-3-one
(62)
A mixture of
(6-chloro-5-isopropyl-pyridazin-3-yl)-[2,6-dibromo-4-(tetrahydro-pyran-2--
yloxymethyl)-phenyl]-acetonitrile (61) (332.8 mg, 0.61 mmol) and
sodium acetate (103.8 mg, 1.26 mmol) in glacial acetic acid (2.7
mL) was heated to reflux for 2.5 h. The reaction was then treated
with concentrated hydrochloric acid (2.0 mL) and heated to reflux
for 18 h. At this time, the reaction was treated with more
concentrated hydrochloric acid (6.0 mL) and additional glacial
acetic acid (3.0 mL) and heated to reflux for an additional 24 h.
The reaction was then cooled to 25.degree. C. and diluted with
ethyl acetate (150 mL). This solution was washed with a saturated
aqueous sodium chloride solution (2.times.50 mL) and a saturated
aqueous sodium carbonate solution (1.times.100 mL), dried with
magnesium sulfate, filtered, and concentrated under vacuum. The
resulting brown oil was treated with glacial acetic acid (1.0 mL)
and concentrated hydrochloric acid (6.0 mL) and was heated to
reflux for 3 d. At this time, the reaction was cooled to 25.degree.
C. and diluted with ethyl acetate (150 mL). This solution was
washed with a saturated aqueous sodium chloride solution
(2.times.50 mL) and a saturated aqueous sodium carbonate solution
(1.times.100 mL), dried with magnesium sulfate, filtered, and
concentrated under vacuum. Flash chromatography (Merck Silica gel
60, 230-400 mesh, 2/1 hexanes/ethyl acetate) afforded
6-(2,6-dibromo-4-chloromethyl-benzyl)-4-isopropyl-pyridazin-3-one
(62) (96.3 mg, 36%) as an off-white solid; LRMS for
C.sub.15H.sub.15Br.sub.2ClN.sub.2O (M+H) at m/z=433. Molecular
Weight=434.5606; Exact Mass=431.9240
Step 7: Preparation of
[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetonitrile (63)
A suspension of sodium cyanide (279 mg, 5.69 mmol) in dimethyl
sulfoxide (2.0 mL) was treated with concentrated sulfuric acid
(0.10 mL),
6-(2,6-dibromo-4-chloromethyl-benzyl)-4-isopropyl-pyridazin-3-one
(62) (346 mg, 0.79 mmol) and an addition rinse of dimethyl
sulfoxide (3.0 mL). The reaction was stirred at 25.degree. C. for 5
min and at 40.degree. C. for 1 h. At this time, the reaction was
heated to 60.degree. C. for 1 h and then was cooled to 25.degree.
C. The mixture was poured onto a mixture of ice and a saturated
aqueous sodium bicarbonate solution (50 mL). This mixture was
extracted into ethyl acetate (3.times.50 mL). The combined organics
were washed with a saturated aqueous sodium chloride solution
(1.times.50 mL), dried over magnesium sulfate, filtered and
concentrated under vacuum. Flash chromatography (Merck Silica gel
60, 230-400 mesh, 1:2 hexanes/ethyl acetate) afforded
[3,5-dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetonitrile (63) (227 mg, 67%) as a light orange solid; LRMS
for C.sub.16H.sub.15Br.sub.2N.sub.3O (M+H) at m/z=424. Molecular
Weight=425.1255; Exact Mass=422.9582
Step 8: Preparation of
[3,5-Dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetic acid (64)
A mixture of
[3,5-dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetonitrile (63) (224.2 mg, 0.52 mmol) in concentrated
hydrochloric acid (4.0 mL) was heated to 135.degree. C. for 16 h.
At this time, the reaction was cooled to 25.degree. C. and diluted
with a saturated aqueous sodium chloride solution (30 mL). This
solution was extracted with ethyl acetate (175 mL). The organics
were washed with a saturated aqueous sodium chloride solution (30
mL), dried with magnesium sulfate, filtered and concentrated under
vacuum. ISCO chromatography (40 g column, 9:1 methylene
chloride/methanol) gave a tan solid. This solid was triturated with
acetonitrile (10 mL) and methylene chloride (2.0 mL), filtered, and
dried at 110.degree. C. overnight under high vacuum to afford
[3,5-dibromo-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-pheny-
l]-acetic acid (64) (114.4 mg, 48.8%) as an off-white solid;
EI(+)-HRMS m/z calcd for C.sub.16H.sub.16Br.sub.2N.sub.2O.sub.3
(M.sup.+) 240.9964, found 240.9959. Molecular Weight=444.1255;
Exact Mass=441.9528
##STR00102## ##STR00103##
Example 16
Synthesis of
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro
[1,2,4]triazine-6-carbonitrile (69)
##STR00104##
Step 1: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]-isoindole-1,3-dione (65)
A mixture of 4-amino-2,6-dichloro-phenol (50 g, 280.8 mmol) and
potassium tert-butoxide (33.16 g, 280.8 mmol) in
N,N-dimethylacetamide (200 mL) was heated to 90.degree. C. The
resulting solution was then treated with
3,6-dichloro-4-isopropyl-pyridazine (55.31 g, 280.8 mmol). The
reaction was heated at 90.degree. C. for 17 h. At this time, the
reaction was diluted with methyl tert-butyl ether (700 mL) and a
saturated aqueous sodium chloride solution (800 mL). The organic
layer was separated, washed with water (2.times.400 mL) and
concentrated to a volume of .about.200 mL. This solution was
diluted with toluene (800 mL) and then distilled to remove
.about.300 mL of solvent. The remaining solution was cooled to
80.degree. C. and was treated with phthalic anhydride (42.01 g,
280.8 mmol). This mixture was heated to reflux for 4 h while water
was distilled azeotropically. At this time, the reaction was
concentrated under vacuum to .about.200 mL, diluted with glacial
acetic acid (800 mL) and then concentrated to remove .about.300 mL
of solvent. The reaction was treated with sodium acetate (46.06 g,
561.6 mmol) and heated to reflux for 6 h. At this time, the
reaction was cooled to room temperature and diluted with water (500
mL). This mixture was warmed to 60.degree. C., stirred for 30 min
and then cooled to room temperature. The resulting solid was
collected by filtration, washed with a 1:1 mixture of glacial
acetic acid:water (300 mL) followed by water (150 mL), dried under
house vacuum and then dried at 55.degree. C. in a vacuum oven
overnight to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy-
)-phenyl]-isoindole-1,3-dione (84.8 g, 68%) as an off-white solid;
ES(+)-LRMS for C.sub.21H.sub.15Cl.sub.2N.sub.3O.sub.4 (M+H).sup.+
at m/z=444. Exact Mass=443.0440; Molecular Weight=444.28.
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-isoindole-1,3-dione (66)
A suspension of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yloxy)-pheny-
l]isoindole-1,3-dione (65) (84.5 g, 190.2 mmol) in
N,N-dimethylformamide dimethyl acetal (250 mL) was heated to reflux
for 5 h. At this time, the reaction was cooled to room temperature.
The solids that resulted were collected by filtration, washed with
a 1:1 mixture of methyl tert-butyl ether:heptane (70 mL) followed
by heptane (70 mL) and dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-yloxy)-phenyl]-isoindole-1,3-dione (66) (63.57 g, 73%) as an
off-white solid; ES(+)-LRMS for
C.sub.22H.sub.17Cl.sub.2N.sub.3O.sub.4 (M+H).sup.+ at m/z=458.
Exact Mass=457.0596; Molecular Weight=458.30.
Step 3: Preparation of
6-(4-Amino-2,6-dichlorophenoxy)-4-isopropyl-2-methyl-2H-pyridazin-3-one
(67)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-isoindole-1,3-dione (66) (64 g, 139.6 mmol) in methanol
(500 mL) was treated with butylamine (34.67 mL, 349 mmol). The
mixture was heated to reflux for 1.5 h. At this time, the reaction
was cooled to room temperature and treated dropwise with water (384
mL). The resulting solids were collected by filtration, washed with
a 1:1 solution of methanol/water (180 mL) followed by water (250
mL) and dried under vacuum to afford
6-(4-amino-2,6-dichloro-phenoxy)-4-isopropyl-2-methyl-2H-pyrida-
zin-3-one (67) (34.12 g, 74.4%) as an off-white solid; ES(+)-LRMS
for C.sub.14H.sub.15Cl.sub.2N.sub.3O.sub.2 (M+H).sup.+ at m/z=328.
Exact Mass=327.0541; Molecular Weight=328.20.
Step 4: Preparation of
(2-Cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-yloxy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(68)
A mixture of
6-(4-amino-2,6-dichloro-phenoxy)-4-isopropyl-2-methyl-2H-pyridazin-3-one
(67) (10 g, 30.47 mmol) in glacial acetic acid (60 mL) and
concentrated hydrochloric acid (9.06 mL) cooled to 5-10.degree. C.
was treated dropwise with a solution of sodium nitrite (2.3 g, 32.3
mmol) in water (6 mL). The reaction was stirred at 5-10.degree. C.
for 30 min. At this time, the reaction was treated with
N-cyanoacetylurethane (5.34 g, 33.52 mmol) followed by a solution
of sodium acetate (7.5 g, 91.41 mmol) in water (22.5 mL). The
reaction was stirred at 5-10.degree. C. for 30 min and then was
diluted with water (50 mL). The resulting solids were collected by
filtration, washed with a 1:1 mixture of glacial acetic acid:water
(40 mL) followed by water (2.times.60 mL), dried under house
vacuum, and then dried under vacuum at 50.degree. C. to afford
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-yloxy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(68) (15.04 g, 95%) as an orange solid; ES(+)-LRMS for
C.sub.20H.sub.20Cl.sub.2N.sub.6O.sub.5 (M+H).sup.+ at m/z=495.
Exact Mass=494.0872; Molecular Weight=495.33.
Step 5: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylo-
xy)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(69)
A solution of
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-yloxy)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(68) (6.5 g, 13.12 mmol) and potassium acetate (1.42 g, 14.43 mmol)
in N,N-dimethylacetamide was heated to 120.degree. C. for 2 h. At
this time, the reaction was cooled to room temperature and was
treated with glacial acetic acid (1.5 mL) and water (65 mL). The
mixture was stirred at room temperature for 15 min. The solids
which resulted were collected by filtration, washed with water (65
mL) and dried under vacuum to give an orange solid. The solid was
suspended in acetonitrile (36 mL), heated to reflux for 3 h and
then cooled to room temperature. The resulting solids were then
collected by filtration, washed with acetonitrile (12 mL) and
methyl tert-butyl ether (24 mL) and dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo
-1,6-dihydro-pyridazin-3-yloxy)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro[1,2,-
4]triazine-6-carbonitrile (69) (5.06 g, 85.8%) as an orange solid;
ES(+)-HRMS m/e calcd for C.sub.18H.sub.14Cl.sub.2N.sub.6O.sub.4
(M+H).sup.+ 449.0527, found 449.0527. Exact Mass=448.0454;
Molecular Weight=449.26.
##STR00105## ##STR00106##
Example 17
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)-ph-
enyl]-acetic acid (76)
##STR00107##
Step 1: Preparation of
4-[2-(tert-Butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenol
(70)
A solution of 2,6-dichloro-4-(2-hydroxy-ethyl)-phenol (16) (2.0 g,
9.66 mmol) in methylene chloride (20 mL) at 25.degree. C. was
treated with triethylamine (1.35 mL, 9.66 mmol) and tert-butyl
diphenylsilylchloride (2.48 mL, 9.66 mmol). The reaction was
stirred at 25.degree. C. for 18 h. At this time, the reaction was
diluted with methylene chloride. The organics were washed with a 1N
aqueous hydrochloric acid solution, water, and a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, filtered
and concentrated under vacuum. Flash chromatography (Merck Silica
gel 60, 230-400 mesh, 95:5 petroleum ether/ethyl acetate) afforded
4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenol
(70) (2.60 g, 60%) as a pale yellow oil; HRMS m/e calcd for
C.sub.24H.sub.26Cl.sub.2O.sub.2Si (M+Na).sup.+ 467.0971, found
467.0977. Exact Mass=444.1079; Molecular Weight=445.47.
Step 2: Preparation of Dimethyl-thiocarbamic acid
0-{4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenyl}ester
(71)
A solution of
4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenol
(70) (289 mg, 0.78 mmol) in N,N-dimethylformamide (4 mL) at
25.degree. C. was treated with 1,4-diazabicyclo[2.2.2]octane (172
.mu.L, 1.56 mol) and dimethylthiocarbamoyl chloride (154 mg, 1.25
mmol). The reaction was stirred at 25.degree. C. for 18 h. At this
time, the reaction was diluted with ethyl acetate and was then
washed with a 1N aqueous hydrochloric acid solution, water, and a
saturated aqueous sodium chloride solution, dried with magnesium
sulfate, filtered and concentrated under vacuum. Flash
chromatography (Merck Silica gel 60, 230-400 mesh, 95:5 petroleum
ether/ethyl acetate) afforded dimethyl-thiocarbamic acid
O-{4-[2-(tert-butyl-diphenyl-silanyloxy)ethyl]-2,6-dichloro-phenyl}ester
(71) (376 mg, 90%) as a white solid; EI(+)-HRMS m/e calcd for
C.sub.27H.sub.31Cl.sub.2N.sub.2O.sub.2SSi (M+H).sup.+ 532.1295,
found 532.1292. Exact Mass=531.1222; Molecular weight=532.61
Step 3: Preparation of Dimethyl-thiocarbamic acid
S-{4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenyl}ester
(72)
Dimethyl-thiocarbamic acid
0-{4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenyl}ester
(71) (2.98 g, 5.60 mmol) was heated to 190-200.degree. C. for 24 h.
At this time, the residue was cooled to 25.degree. C. and was
dissolved in methylene chloride. Flash chromatography (Merck Silica
gel 60, 230-400 mesh, 95/5 petroleum ether/ethyl acetate) afforded
dimethyl-thiocarbamic acid
S-{4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenyl}-
ester (72) (2.08 g, 70%) as a pale oil; EI(+)-HRMS m/e calcd for
C.sub.27H.sub.31Cl.sub.2NO.sub.2SSi (M+H).sup.+ 532.1295, found
532.1301. Exact Mass=531.1222; Molecular weight=532.61
Step 4: Preparation of 2-(3,5-Dichloro-4-mercapto-phenyl)-ethanol
(73)
A solution of dimethyl-thiocarbamic acid
S-{4-[2-(tert-butyl-diphenyl-silanyloxy)-ethyl]-2,6-dichloro-phenyl}ester
(72) (2.0 g, 3.76 mmol) in ethanol (6.0 mL) was treated with a 3N
aqueous potassium hydroxide solution (4.4 mL, 13.2 mmol). The
reaction was then heated to 95.degree. C. for 18 h. At this time,
the reaction was poured onto water (200 mL), acidified to pH=2 with
a 3N aqueous hydrochloric acid solution, and then extracted into
methylene chloride (3.times.100 mL). The organics were washed with
water (1.times.100 mL) and a saturated aqueous sodium chloride
solution (1.times.100 mL), dried with magnesium sulfate, filtered,
and concentrated under vacuum. Flash chromatography (Merck Silica
gel 60, 230-400 mesh, 80:20 petroleum ether/ethyl acetate) afforded
2-(3,5-dichloro-4-mercapto-phenyl)-ethanol (73) (639 mg, 76%) as a
white solid; EI(+)-HRMS m/e calcd for C.sub.8H.sub.8Cl.sub.2OS
(M.sup.+) 221.9673, found 221.9672. Exact Mass=221.9673; Molecular
Weight=223.12
Step 5: Preparation of
2-[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-e-
thanol (74)
A solution of 2-(3,5-dichloro-4-mercapto-phenyl)-ethanol (73) (620
mg, 2.78 mmol) in dimethyl sulfoxide (25 mL) was treated with
3,6-dichloro-4-isopropyl-pyridazine (7) (530 mg, 2.78 mmol) and
potassium carbonate (1.54 g, 11.12). The resulting mixture was
heated to 90.degree. C. for 18 h. At this time, the reaction was
cooled to 25.degree. C., poured into a solution of water (200 mL)
and a 1N aqueous hydrochloric acid solution (25 mL). The mixture
was extracted with ethyl acetate (2.times.200 mL). The organics
were then washed with water (1.times.100 mL) and a saturated
aqueous sodium chloride solution (1.times.100 mL), dried with
magnesium sulfate, filtered, and concentrated under vacuum. Flash
chromatography (Merck Silica gel 60, 230-400 mesh, 70:30 petroleum
ether/ethyl acetate) afforded
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-e-
thanol (74) (550 mg, 52%) as a pale yellow oil; EI(+)-HRMS m/e
calcd for C.sub.15H.sub.15Cl.sub.3N.sub.2OS (M+H).sup.+ 377.0044,
found 377.0043. Exact Mass=375.9971; Molecular weight=377.72
Step 6: Preparation of
[3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-ace-
tic acid (75)
A solution of
2-[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-e-
thanol (74) (100 mg, 0.26 mmol) in acetone (2.0 mL) cooled to
0.degree. C. was treated with a 2.7M solution of Jones reagent
(0.21 mL, 0.57 mmol, prepared via standard method). The reaction
was stirred at 0.degree. C. for 1.25 h. At this time, the reaction
was treated with 2-propanol until the red solution turned green.
This mixture was then diluted with ethyl acetate (50 mL) and water
(25 mL). The organics were separated, dried with magnesium sulfate,
filtered, and concentrated under vacuum to afford
[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-ace-
tic acid (75) (92.5 mg, 89%) as an off-white foam. This material
was used without further purification; LRMS-APCI for
C.sub.15H.sub.13Cl.sub.3N.sub.2O.sub.2S (M+H).sup.+ at m/z=390.
Exact Mass=389.9763; Molecular weight=391.71
Step 7: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)-ph-
enyl]-acetic acid (76)
A mixture of
[3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenyl]-ace-
tic acid (75) (90 mg, 0.23 mmol) and sodium acetate (66 mg, 0.80
mmol) in glacial acetic acid (2.5 mL) was heated to 100.degree. C.
for 7 h. At this time, the reaction was concentrated in vacuo. The
residue was slurried three times in a 1:1 solution of methylene
chloride:hexanes followed by concentration under vacuum. The
resulting solid was diluted with water (100 mL) and was then
treated with a 1N aqueous sodium hydroxide solution to adjust the
pH between 10-11. This solution was extracted with ethyl acetate.
The organics were discarded. The aqueous layer was then acidified
to pH=2-3 with a 1N aqueous hydrochloric acid solution. This
solution was then extracted with ethyl acetate (2.times.100 mL).
The organics were washed with a saturated aqueous sodium chloride
solution (1.times.100 mL), dried with magnesium sulfate, filtered,
and concentrated under vacuum. Flash chromatography (Merck Silica
gel 60, 230-400 mesh, 70:30 hexanes/ethyl acetate with 2% glacial
acetic acid) afforded
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)-ph-
enyl]acetic acid (76) (41 mg, 17.5%) as a white solid; EI(+)-HRMS
m/e calcd for C.sub.15H.sub.14Cl.sub.2N.sub.2O.sub.3S (M+H).sup.+
373.0175, found 373.0175. Exact Mass=372.0102; Molecular
Weight=373.26
##STR00108##
Example 18
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfinyl)-phe-
nyl]-acetic acid (78) and
Example 19
Synthesis of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-phe-
nyl]-acetic acid (79)
##STR00109##
Step 1: Preparation of
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfinyl)-phe-
nyl]-acetic acid (77) and
[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-phe-
nyl]-acetic acid (78)
A mixture of
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)-ph-
enyl]acetic acid (76) (75 mg, 0.2 mmol) in formic acid (1.0 mL) at
0.degree. C. was treated with a 30% aqueous hydrogen peroxide
solution (61 .mu.L, 0.6 mmol). The resulting suspension was stirred
at 0.degree. C. for 30 min and then was stirred at 25.degree. C.
for 7 d during which time more 30% aqueous hydrogen peroxide
solution (322 .mu.L total), formic acid (1.0 mL) and ethanol (1.0
mL) were added to the reaction. At this time, the reaction was
poured onto ethyl acetate/water and was extracted with ethyl
acetate (2.times.50 mL). The organics were dried with magnesium
sulfate, filtered and concentrated under vacuum. HPLC (20-70
acetonitrile/water with 0.1% trifluoroacetic acid over 30 min)
afforded
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulf-
inyl)-phenyl]-acetic acid (77) (32 mg, 41%) as a white solid
ES-HRMS m/e calcd for C.sub.15H.sub.14Cl.sub.2N.sub.2O.sub.4S
(M+H).sup.+ 389.0124, found 389.0124: Exact Mass=388.0051;
Molecular weight=389.26; and
[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-phe-
nyl]-acetic acid (78) (22 mg, 27%) as a white solid; EI(+)-HRMS m/e
calcd for C.sub.15H.sub.14Cl.sub.2N.sub.2O.sub.5S (M+H).sup.+
377.0044, found 377.0043: Exact Mass=404.0000; Molecular
weight=405.26
##STR00110## ##STR00111## ##STR00112##
Example 20
Synthesis of Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(88)
##STR00113##
Step 1: Preparation of Dimethyl-thiocarbamic acid
O-(2,6-dichloro-4-nitro-phenyl)ester (79)
A solution of 2,6-dichloro-4-nitro-phenol (3.0 g, 14.4 mmol) in
N,N-dimethylformamide (70 mL) at 25.degree. C. was treated with
1,4-diazabicyclo[2.2.2]octane (3.16 mL, 28.8 mmol) and
dimethylthiocarbamoyl chloride (2.85 g, 23.04 mmol). The reaction
was stirred at 25.degree. C. for 18 h. At this time, the reaction
was diluted with ethyl acetate (250 mL) and was then washed with a
1N aqueous hydrochloric acid solution (1.times.125 mL), water
(1.times.125 mL), and a saturated aqueous sodium chloride solution
(1.times.125 mL), dried with magnesium sulfate, filtered and
concentrated under vacuum. Flash chromatography (Merck Silica gel
60, 230-400 mesh, 80:20 petroleum ether/ethyl acetate) afforded
dimethyl-thiocarbamic acid O-(2,6-dichloro-4-nitro-phenyl)ester
(79) (3.2 g, 75%) as a white solid; EL-LRMS for
C.sub.9H.sub.8Cl.sub.2N.sub.2O.sub.3S (M-Cl).sup.+ at m/z=259.
Exact Mass=293.9633; Molecular weight=295.15
Step 2: Preparation of Dimethyl-thiocarbamic acid
S-(2,6-dichloro-4-nitro-phenyl)ester (80)
Dimethyl-thiocarbamic acid O-(2,6-dichloro-4-nitro-phenyl)ester
(79) (3.2 g, 10.8 mmol) was heated to 180.degree. C. for 20 min to
afford dimethyl-thiocarbamic acid
S-(2,6-dichloro-4-nitro-phenyl)ester (80) (3.23 g) as a tan solid.
The material was used without further purification; EL-LRMS for
C.sub.9H.sub.8Cl.sub.2N.sub.2O.sub.3S (M+H).sup.+ at m/z=295. Exact
Mass=293.9633; Molecular weight=295.15
Step 3: Preparation of Dimethyl-thiocarbamic acid
S-(4-amino-2,6-dichloro-phenyl)ester (81)
A mixture of dimethyl-thiocarbamic acid
S-(2,6-dichloro-4-nitro-phenyl)ester (80) (1.6 g, 5.4 mmol) in
glacial acetic acid (24 mL), 2-propanol (48 mL), and water (24 mL)
heated to 50.degree. C. was treated with iron powder (2.1 g, 37.8
mmol). The resulting mixture was heated to 95.degree. C. for 2 h.
At this time, the reaction was filtered hot through a pad of celite
and was washed with water and ethyl acetate. The filtrates were
concentrated to remove the majority of organics. The remaining
solution was diluted with water (500 mL) and was then brought to
pH=8 with a concentrated ammonium hydroxide solution. The solution
was extracted with ethyl acetate (3.times.200 mL). The combined
organics were dried with magnesium sulfate, filtered and
concentrated under vacuum. The resulting residue was slurried with
diethyl ether and was cooled in the freezer for 30 min. The solid
that formed was collected by filtration and was washed with cold
diethyl ether to afford dimethyl-thiocarbamic acid
S-(4-amino-2,6-dichloro-phenyl)ester (81) (1.33 g, 93%) as a white
solid; EI(+)-HRMS m/e calcd for C.sub.9H.sub.10Cl.sub.2N.sub.2OS
(M+H).sup.+ 264.9964, found 264.9964. Exact Mass=263.9891;
Molecular weight=265.16
Step 4: Preparation of 4-Amino-2,6-dichloro-benzenethiol (82)
A solution of dimethyl-thiocarbamic acid
S-(4-amino-2,6-dichloro-phenyl)ester (81) (2.0 g, 7.5 mmol) in
ethanol was treated with a 3N aqueous potassium hydroxide solution.
The reaction was heated to 95.degree. C. for 2 d. At this time, the
reaction was cooled to 25.degree. C. and was then acidified to pH=2
with a 3N aqueous hydrochloric acid solution. This solution was
diluted with water (500 mL) and then was extracted with ethyl
acetate (2.times.250 mL). The organics were dried with magnesium
sulfate, filtered, and concentrated under vacuum to an orange
solid. The solid was slurried in chloroform, collected by
filtration and washed with chloroform. The filtrate was
concentrated under vacuum and purified by flash chromatography
(Merck Silica gel 60, 230-400 mesh, 85:15 petroleum ether/ethyl
acetate) to afford 4-amino-2,6-dichloro-benzenethiol (82) (1.0 g,
69%) as a white solid; EI(+)-HRMS m/e calcd for
C.sub.6H.sub.5Cl.sub.2NS (M.sup.+) 192.9520, found 192.9519. Exact
Mass=192.9520; Molecular weight=194.08
Step 5: Preparation of
3,5-Dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenylamine
(83)
A solution of 4-amino-2,6-dichloro-benzenethiol (82) (1.0 g, 5.2
mmol) in N,N-dimethylformamide at 25.degree. C. was treated with
3,6-dichloro-4-isopropyl-pyridazine (7) (990 mg, 5.2 mmol) and
potassium carbonate (2.16 g, 15.6 mmol). The resulting mixture was
heated to 90.degree. C. for 18 h. At this time, the reaction was
cooled to 25.degree. C., poured onto a mixture of ice water and a
1N aqueous hydrochloric acid solution (10 mL). The resulting
solution was brought to pH=7 with additional 1N aqueous
hydrochloric acid solution and then was extracted with ethyl
acetate (2.times.100 mL). The organics were washed with a saturated
aqueous sodium chloride solution (1.times.50 mL), dried with
magnesium sulfate, filtered and concentrated under vacuum. Flash
chromatography (Merck Silica gel 60, 230-400 mesh, 80/20 petroleum
ether/ethyl acetate) afforded
3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenylamine
(83) (1.29 g, 72%) as an off-white foam; ES(+)-HRMS m/e calcd for
C.sub.13H.sub.12Cl.sub.3N.sub.3S (M+H).sup.+ 347.9891, found
347.9889. Exact Mass=346.9817; Molecular weight=348.68
Step 6: Preparation of
N-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-acetamide (84)
A solution of
3,5-dichloro-4-(6-chloro-5-isopropyl-pyridazin-3-ylsulfanyl)-phenylamine
(83) (1.23 g, 3.50 mmol) in glacial acetic acid (35 mL) was treated
with sodium acetate (1.0 g, 12.25 mmol). This mixture was heated to
95.degree. C. for 18 h. At this time, the reaction mixture was
cooled to 25.degree. C., poured onto water (200 mL) and neutralized
with a 3N aqueous sodium hydroxide solution. This solution was then
extracted with ethyl acetate (2.times.200 mL). The combined
organics were washed with a saturated aqueous sodium chloride
solution (1.times.100 mL), dried with magnesium sulfate, filtered
and concentrated under vacuum to afford
N-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yl-sulfanyl)-
-phenyl]-acetamide (84) (1.3 g, 99%) as an off-white solid. This
material was used without further purification; ES(+)-HRMS m/e
calcd for C.sub.15H.sub.15Cl.sub.2N.sub.3O.sub.2S (M+H).sup.+
372.0335, found 372.0335. Exact Mass=371.0262; Molecular
weight=372.28
Step 7: Preparation of
6-(4-Amino-2,6-dichloro-phenylsulfanyl)-4-isopropyl-2H-pyridazin-3-one
(85)
A mixture of
N-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-acetamide (84) (4.0 g, 10.7 mmol) in methanol (15 mL) and
water (15 mL) was treated with powdered sodium hydroxide (2.14 g,
53.5 mmol). The reaction was then heated to reflux for 18 h. At
this time, the reaction was cooled to 25.degree. C., diluted with
water (500 mL) and extracted with ethyl acetate (3.times.300 mL).
The organics were washed with a saturated aqueous sodium chloride
solution (1.times.200 mL), dried with magnesium sulfate, filtered
and concentrated under vacuum to afford
6-(4-amino-2,6-dichloro-phenyl
sulfanyl)-4-isopropyl-2H-pyridazin-3-one (85) (3.5 g, 99%) as a
light tan solid. The material was used without further
purification; ES(+)-HRMS m/e calcd for
C.sub.13H.sub.13Cl.sub.2N.sub.3OS (M+H).sup.+ 330.0229, found
330.0229. Exact Mass=329.0156; Molecular weight=330.24
Step 8: Preparation of
(2-Cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yl-
sulfanyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester
(86)
A mixture of
6-(4-amino-2,6-dichloro-phenylsulfanyl)-4-isopropyl-2H-pyridazin-3-one
(85) (3.40 g, 10.4 mmol) in water (135 mL) and concentrated
hydrochloric acid (68 mL) cooled to 0.degree. C. was treated with a
solution of sodium nitrite (853 mg, 12.36 mmol) in water (7.0 mL)
via Pasteur pipette under the surface of the reaction mixture. This
was followed by a water rinse (1.0 mL) of the Pasteur pipette. The
resulting yellow mixture was stirred at 0.degree. C. for 30 min.
More concentrated hydrochloric acid (7.0 mL) was added. The
reaction was stirred at 0.degree. C. for an additional 1.3 h. The
solids were removed by filtration through filter paper and were
rinsed with water. The clear, yellow diazonium salt solution of the
filtrate was quickly poured into a solution of cyanoacetylurethane
(1.77 g, 11.33 mmol), pyridine (75 mL) and water (204 mL) cooled to
0.degree. C. Upon mixing, orange-red solids immediately formed.
This mixture was stirred at 0.degree. C. for 1 h. At this time, the
solids were collected by filtration through filter paper. The
solids were washed with water, air-dried under house vacuum for 2
h, and then dried under vacuum to afford
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridaz-
in-3-ylsulfanyl)-phenyl]hydrazono}-acetyl)-carbamic acid ethyl
ester (86) (2.98 g, 58%) as an orange solid; ES(+)-HRMS m/e calcd
for C.sub.19H.sub.18Cl.sub.2N.sub.6O.sub.4S (M+H).sup.+ 497.0560,
found 497.0559. Exact Mass=496.0487; Molecular weight=497.36
Step 9: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(87)
A solution of
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-yl-
sulfanyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl ester (86)
(2.98 g, 5.99 mmol) in glacial acetic acid (60 mL) was treated with
sodium acetate (2.46 g, 29.95 mmol). The resulting mixture was
heated to 120.degree. C. for 3 h. At this time, LCMS indicated
complete consumption of the starting material and conversion to
product. The reaction was cooled to 25.degree. C., poured onto
water (200 mL) and was extracted with ethyl acetate (2.times.150
mL). The organics were washed with a saturated aqueous sodium
chloride solution (1.times.150 mL), dried with magnesium sulfate,
filtered and concentrated under vacuum to give an orange solid.
This solid was dissolved in a 1:1 methylene chloride:hexanes
mixture and concentrated in vacuo three times. It was then
dissolved in a 1:1 methylene chloride: methanol mixture and
concentrated under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(87) (2.79 g) as an orange solid. The material was used without
further purification; ES(+)-HRMS m/e calcd for
C.sub.17H.sub.12Cl.sub.2N.sub.6O.sub.3S (M+H).sup.+ 451.0142, found
451.0143. Exact Mass=450.0069; Molecular weight=451.29
Step 10: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(88)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(87) (45 mg, 0.09 mmol) in glacial acetic acid (2.5 mL) was treated
with a 30% aqueous hydrogen peroxide solution (0.28 mL, 0.98 mmol).
The reaction was heated to 80.degree. C. for 3 d. An additional
amount of the 30% aqueous hydrogen peroxide solution (0.28 mL, 0.98
mmol) was added each day. After 3 d, the reaction was diluted with
ethyl acetate and water. The organics were washed with water, dried
with magnesium sulfate, filtered and concentrated under vacuum.
HPLC (20-70 acetonitrile/water aith 0.1% trifluoroacetic acid over
30 min) afforded
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(88) (2.0 mg, 4%) as a white solid; ES(+)-HRMS m/e calcd for
C.sub.17H.sub.12Cl.sub.2N.sub.6O.sub.5S (M+H).sup.+ 483.9940, found
483.0041. Exact Mass=481.9967; Molecular weight=483.29
##STR00114## ##STR00115##
Example 21
Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-2H-[1,2,4]triazine-3,5-dione (91)
##STR00116##
Step 1: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (89)
A solution of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(87) (1.35 g, 2.99 mmol) in glacial acetic acid (30 mL) and
concentrated hydrochloric acid (6.7 mL) was heated to 120.degree.
C. for 3 d. At this time, the reaction was cooled to 0.degree. C.,
diluted with water (50 mL) and stirred at 0.degree. C. for 15 min.
The product that precipitated was collected by filtration, washed
with water and petroleum ether, air-dried under house vacuum for 15
min, and then dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (89) (1.17 g, 83%) as a light brown solid; ES(+)-HRMS m/e
calcd for C.sub.17H.sub.13Cl.sub.2N.sub.5O.sub.5S (M+H).sup.+
470.0087, found 470.0088. Exact Mass=469.0014; Molecular
weight=470.29
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-2H-[1,2,4]triazine-3,5-dione (90)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carboxylic
acid (89) (1.0 g, 2.13 mmol) and thioglycolic acid (21.3 mL, 2.13
mmol) were heated at 170.degree. C. for 3.5 h. At this time, the
reaction was cooled to 25.degree. C. and diluted with water (500
mL). The aqueous layer was brought to pH=4 by the addition of a
saturated aqueous sodium bicarbonate solution. The solids that
precipitated were collected by filtration, washed consecutively
with water, petroleum ether, a pH=5 buffer and water. The solids
were air-dried under house vacuum for 15 min and then dried under
vacuum to give a light brown solid. This solid was dissolved in a
1N aqueous sodium hydroxide solution (100 mL) and extracted with
ethyl acetate (1.times.120 mL). The aqueous layer was neutralized
by the addition of a 1N aqueous hydrochloric acid solution (15 mL).
The resulting solids were collected by filtration, washed with
water (200 mL) and petroleum ether (100 mL), air-dried under house
vacuum and then dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-2H-[1,2,4]triazine-3,5-dione (90) (338 mg, 37%) as tan
solid; ES(+)-HRMS m/e calcd for
C.sub.16H.sub.13Cl.sub.2N.sub.5O.sub.3S (M+H).sup.+ 426.0189, found
426.0189. Exact Mass=425.0116; Molecular weight=426.28
Step 3: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-2H-[1,2,4]triazine-3,5-dione (91)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylsulfanyl)--
phenyl]-2H-[1,2,4]triazine-3,5-dione (90) (165 mg, 0.38 mmol) in
glacial acetic acid (10 mL) was treated with a 30% aqueous hydrogen
peroxide solution (0.93 mL, 3.87 mmol). The reaction mixture was
then heated to 90.degree. C. for 3 d. At this time, the reaction
was cooled to 25.degree. C. and was poured onto water (100 mL).
This solution was extracted with ethyl acetate (1.times.125 mL).
The organics were washed with water, dried with magnesium sulfate,
filtered and concentrated under vacuum. HPLC (20-80-30
acetonitrile/water with 0.1% trifluoroacetic acid over 30 min
conditions) afforded
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazine-3-sulfonyl)-p-
henyl]-2H-[1,2,4]triazine-3,5-dione (91) (48.5 mg, 27%) as a white
solid; ES(+)-HRMS m/e calcd for
C.sub.16H.sub.13Cl.sub.2N.sub.5O.sub.5S (M+H).sup.+ 458.0087, found
458.0090. Exact Mass=457.0014; Molecular weight=458.28
##STR00117## ##STR00118##
Example 22
Synthesis of
2-[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(96)
##STR00119##
Step 1: Preparation of
[2,6-Dichloro-4-(1,3-dioxo-1,3-dihydro-isoindol-2-yl)-phenyl]-acetonitril-
e (92)
A solution of
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-2H-pyridazin-3-one (910
mg, 2.90 mmol) in glacial acetic acid (12 mL) was treated with
phthalic anhydride (430 mg, 2.9 mmol). The reaction was then heated
to 130.degree. C. for 3.5 h. At this time, the reaction was poured
onto water (200 mL) and extracted into ethyl acetate (300 mL). The
organics were then washed with water, dried with magnesium sulfate,
filtered and concentrated under vacuum. The resulting orange solid
was slurried in cold acetonitrile, collected by filtration, washed
with cold acetonitrile and dried under vacuum to afford
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-isoindole-1,3-dione (92) (780 mg, 61%) as a light, tan solid;
ES(+)-LRMS for C.sub.22H.sub.17Cl.sub.2N.sub.3O.sub.3 (M+H).sup.+
at m/z=442. Exact Mass=441.0647; Molecular Weight=442.3051
Step 2: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-isoindole-1,3-dione (93)
A mixture of
2-[3,5-dichloro-4-(5-isopropyl-6-oxo-1,6-dihydro-pyridazin-3-ylmethyl)-ph-
enyl]-isoindole-1,3-dione (92) (570 mg, 1.29 mmol) and
N,N-dimethylformamide dimethyl acetal (12 mL) was heated to
105.degree. C. for 4.5 h. At this time, the reaction was cooled to
25.degree. C. and diluted with methylene chloride. This solution
was washed with water (1.times.50 mL) and a saturated aqueous
sodium chloride solution (1.times.50 mL), dried with magnesium
sulfate, filtered and concentrated under vacuum. Flash
chromatography (Merck Silica gel 60, 230-400 mesh, 95:5 methylene
chloride/methanol) afforded
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-isoindole-1,3-dione (93) (310 mg, 52%) as an
off-white solid; ES_HRMS m/e calcd for
C.sub.23H.sub.19Cl.sub.2N.sub.3O.sub.3 (M+H).sup.+ 456.0876, found
456.0876. Exact Mass=455.0803; Molecular Weight=456.3322.
Step 3: Preparation of
6-(4-Amino-2,6-dichloro-benzyl)-4-isopropyl-2-methyl-2H-pyridazin-3-one
(94)
A solution of
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-isoindole-1,3-dione (93) (300 mg, 0.66 mmol) in
glacial acetic acid was heated to 110.degree. C. for 3 h. At this
time, the reaction was cooled to 25.degree. C. and was poured onto
water (100 mL). This mixture was extracted with ethyl acetate
(3.times.50 mL). The organics were washed with a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, filtered
and concentrated under vacuum to a brown oil. Recrystallization
from diethyl ether and hexanes followed by flash chromatography
(Merck Silica gel 60, 230-400 mesh, 30:70 ethyl acetate/petroleum
ether) afforded
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-2-methyl-2H-pyridazin-3-one
(94) (148 mg, 69%) as an off-white solid; ES_HRMS m/e calcd for
C.sub.15H.sub.17Cl.sub.2N.sub.3O (M+H).sup.+ 326.0822, found
326.0822. Exact Mass=325.0749; Molecular Weight=326.2282.
Step 4: Preparation of
(2-Cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-ylmethyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl
ester (95)
A mixture of
6-(4-amino-2,6-dichloro-benzyl)-4-isopropyl-2-methyl-2H-pyridazin-3-one
(94) (148 mg, 0.45 mmol) in water (6 mL) and concentrated
hydrochloric acid (3 mL) cooled to 0.degree. C. was treated with a
solution of sodium nitrite (37.3 mg, 0.54 mmol) in water (0.5 mL)
via Pasteur pipette under the surface of the reaction mixture. This
was followed by a water rinse (0.5 mL) of the Pasteur pipette. The
resulting pale yellow solution was stirred at 0.degree. C. for 45
min. At this time, the reaction was filtered through a pad of
cotton and was drained directly into a solution of
cyanoacetylurethane (77.3 mg, 0.49 mmol), pyridine (3 mL) and water
(9 mL) cooled to 0.degree. C. Upon mixing, orange-red solids
immediately formed. This mixture was stirred at 0.degree. C. for 30
min. At this time, the solids were collected by filtration through
filter paper. The solids were washed with water and petroleum
ether, air-dried under house vacuum, and then dried under vacuum to
afford
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-ylmethyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl
ester (95) (192 mg, 86%) as an orange solid; ES-HRMS m/e calcd for
C.sub.21H.sub.22Cl.sub.2N.sub.6O.sub.4 (M+H).sup.+ 493.1153, found
493.1155. Exact Mass=492.1080; Molecular weight=493.3533.
Step 5: Preparation of
2-[3,5-Dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(96)
A solution of
(2-cyano-2-{[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyrid-
azin-3-ylmethyl)-phenyl]-hydrazono}-acetyl)-carbamic acid ethyl
ester (95) (190 mg, 0.38 mmol) in glacial acetic acid (4 mL) was
treated with sodium acetate (158 mg, 1.15 mmol). The resulting
mixture was heated to 120.degree. C. for 3.5 h. The reaction was
cooled to 25.degree. C. and was then poured onto water (125 mL).
The resulting orange mixture was extracted with ethyl acetate (150
mL). The combined organics were washed with a saturated aqueous
sodium chloride solution, dried with magnesium sulfate, treated
with Norite neutral decolorizing carbon, filtered through celite
and washed with ethyl acetate. The filtrate was concentrated under
vacuum. Purification by HPLC (10-90 acetonitrile/water with 0.1%
trifluoroacetic acid over 30 min) afforded
2-[3,5-dichloro-4-(5-isopropyl-1-methyl-6-oxo-1,6-dihydro-pyridazin-3-ylm-
ethyl)-phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-[1,2,4]triazine-6-carbonitrile
(96) (87.2 mg, 44%) as an off-white solid; ES-HRMS m/e calcd for
C.sub.19H.sub.16Cl.sub.2N.sub.6O.sub.3 (M+H).sup.+ 447.0734, found
447.0735. Exact Mass=446.0661; Molecular Weight=447.2838.
Example 23
TR/RXR/GRIP Assay
In this Example, a TR/RXR/GRIP assay was used to test
representative compounds of formula (I). Abbreviations used:
H6-TR.beta., ligand binding domain of thyroid hormone receptor
.beta. with hexa His tag; H6-TR.alpha., ligand binding domain of
thyroid hormone receptor .alpha. with hexa His tag; EE-RxR.alpha.,
ligand binding domain of retinoid X receptor with EE-tag; APC,
allophycocyanin; BSA, bovine serum albumin; DMSO, dimethyl
sulfoxide.
Materials
The ligand binding domain (amino acids 148-410) of thyroid hormone
receptor .beta. (H6-TR.beta.) and the ligand binding domain (amino
acids 202-461) of thyroid hormone receptor .alpha. (H6-TR.alpha.)
were cloned into an E. coli expression vector pET28a (Novagen,
Milwaukee, Wis.) which contained a N-terminal hexaHis sequence. The
resulting recombinant hexaHis tagged proteins were produced in E.
coli BL21(DE3) cells. Cells were grown in Terrific Broth (in-house
preparation in-house prepared medium of Bacto tryptone (3.3%, w/v),
Difico yeast extract (2.0%, w/v) and NaCl (0.5%, w/v)) using shake
flasks with a 24 hour induction in 0.2 mM IPTG at 25.degree. C.,
harvested and lysed with five volumes of Buffer A (0.05M Tris, 0.3M
NaCL, 1% W/V Betaine, 0.01M imidazole, 0.02M b-mercapto ethanol, pH
8.0). Lysozyme (1.0 mg/ml, Sigma) and Complete Protease Inhibitor
Cocktail (Roche Diagnostics Gmbh) were added to slurry and solution
sonicated for one min five times at 4.degree. C. The suspension was
centrifuged in a Ti45 Beckmann rotor for two hours at 127, 300 RCF
and the supernatant was loaded onto NI_NTA Agarose (Quigen 30210)
column. After washing with Buffer A, H6-TR.beta. or H6-TR.alpha.
was eluted with Buffer A containing 0.25M Imidazole.
The ligand binding domain of human retinoid X receptor (amino acids
225-462) (RxR.alpha.) was engineered with N-terminal His6 and EE
(EFMPME) tags, a thrombin cleavage site between the His6 and EE
sequences, and cloned into pACYC vector. The resulting
His6-EE-tagged protein was produced in E. coli cells. Cells were
grown using shake flasks with an 18 hour induction in 0.1 mM IPTG
at 18.degree. C., harvested and suspended with five volumes of
Buffer B (0.025M Tris, 0.3M NaCl, 0.02 M imidazole, 0.01M
.beta.-mercaptoethanol, pH 8.0). Lysozyme (0.2 mg/ml, Sigma) and
Complete Protease Inhibitor Cocktail (Roche Diagnostics Gmbh) were
added and stirred for 30 min. at 4.degree. C. The suspension was
sonicated for 30 seconds, five times, at 4.degree. C. The
suspension was centrifuged for 20 min. at 12,000 RCF. The
supernatant was filtered by 0.45 .mu.m pore size membrane and 0.5%
NP-40 was added. The His6-tagged protein was bound to and eluted
from NiNTA metal-affinity resin (QIAGEN, Valencia, Calif.). The
protein was concentrated and dialyzed.
The His6 tag was removed from EE-RxR.alpha. by thrombin digestion,
using 10 units thrombin (Pharmacia, Piscataway, N.J.) per mg
protein and incubating for 2 hours at 25.degree. C. Removal of
thrombin was done batch-wise using Benzamidine-Sepharose 6B
(Pharmacia, Piscataway, N.J.). The protein was concentrated and
dialyzed. This protein was used in the coactivator peptide
recruitment assay.
Europium-conjugated anti-hexa H is antibody and APC-conjugated
streptavidin were purchased from PerkinElmer Life and Analytical
Sciences.
TR.beta./RXR/GRIP Coactivator Peptide Recruitment Assay
Thirty microliters of H6-TR.beta. (50 nM) in 50 mM Hepes, pH 7.0, 1
mM DTT, 0.05% NP40 and 0.2 mg/ml BSA (Binding Buffer) was mixed
with an equal volume of EE-RxR.alpha. (50 nM) in Binding Buffer.
Six microliters of T3 (0-14.8 uM) or test compound (0-1.2 mM) in
DMSO was then added and the solution incubated at 37.degree. C. for
30 min. Thirty microliters of biotin-GRIP peptide
(Biotin-Aca-HGTSLKEKHKILHRLLQDSSSPVDL-CONH.sub.2) (100 nM) in 30 ul
of Binding Buffer plus 5% DMSO was then added and the solution
incubated at 37.degree. C. for 30 min. Thirty microliters of
solution containing 12 nM europium-conjugated anti-hexa His
antibody and 160 nM APC-conjugated streptavidin in 50 mM Tris, pH
7.4, 100 mM NaCl and 0.2 mg/ml BSA was added and the solution
incubated at 4.degree. C. for over night. An aliquot (35 ul/sample)
was transferred to 384-well black microtiter plates. The HTRF
signal was read on the Victor 5 reader (PerkinElmer Life and
Analytical Sciences).
TR.alpha./RXR/GRIP Coactivator Peptide Recruitment Assay
The assay protocol is essentially the same as that of
TR.beta./RXR/GRIP coactivator peptide recruitment assay as
described above except that 125 nM of H6-TR.alpha., 125 nM of
EE-RxR.alpha. and 250 nM of biotin-GRIP were used.
As shown in the Table below, the tested compounds are thyroid
hormone receptor agonists, with EC.sub.50 values from the
THR-beta/RXR/GRIP recruitment assay:
TABLE-US-00001 THR-beta/RXR/GRIP Recruitment assay Example
EC.sub.50 (.mu.M) Systematic Name Example 1 7.745
[4-(5-Isopropyl-6-oxo-1,6- dihydro-pyridazin-3-yloxy)-3,5-
dimethyl-phenyl]-acetic acid Example 2 7.31
[3-Chloro-4-(5-isopropyl-6-oxo- 1,6-dihydro-pyridazin-3-yloxy)-5-
methyl-phenyl]-acetic acid (10b) Example 3 0.51875
[3,5-Dibromo-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenyl]-acetic acid Example 4 2.33425
[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenyl]-acetic acid Example 5 0.699
3-[3,5-Dibromo-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenyl]-proprionic acid Example 6 0.666
[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenylamino]-acetic acid Example 7 0.117
N-[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenyl]-oxalamic acid Example 8 0.1918
2-[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
yloxy)-phenyl]-3,5-dioxo-2,3,4,5- tetrahydro-[1,2,4]triazine-6-
carbonitrile Example 9 0.115 2-[3,5-Dichloro-4-(5-isopropyl-6-
oxo-1,6-dihydro-pyridazin-3- yloxy)-phenyl]-2H-
[1,2,4]triazine-3,5-dione Example 10 0.767
[3,5-Dichloro-4-(5-isopropyl-1- methyl-6-oxo-1,6-dihydro-
pyridazin-3-yloxy)-phenyl]-acetic acid Example 11 1.2 Synthesis of
[3,5-Dibromo-4-(1- isopropyl-6-oxo-1,6-dihydro-
pyridazin-3-yloxy)-phenyl]-acetic acid Example 12 0.133
2-[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
ylmethyl)-phenyl]-3,5-dioxo- 2,3,4,5-tetrahydro-1,2,4]triazine-
6-carbonitrile Example 13 0.092 2-[3,5-Dichloro-4-(5-isopropyl-6-
oxo-1,6-dihydro-pyridazin-3- ylmethyl)-phenyl]-[1,2,4]triazine-
3,5-dione Example 14 2.99 [3,5-Dichloro-4-(5-isopropyl-6-
oxo-1,6-dihydro-pyridazin-3- ylmethyl)-phenyl]-acetic acid Example
15 0.601 [3,5-Dibromo-4-(5-isopropyl-6-
oxo-1,6-dihydro-pyridazin-3- ylmethyl)-phenyl]acetic acid Example
16 0.066 2-[3,5-dichloro-4-(5-isopropyl-1-
methyl-6-oxo-1,6-dihydro- pyridazin-3-yloxy)-phenyl]-3,5-
dioxo-2,3,4,5-tetra- hydro[1,2,4]triazine-6-carbonitrile Example 17
2.04 [3,5-dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazin-3-
ylsulfanyl)-phenyl]-acetic acid Example 18 9.178
[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazine-3-
sulfinyl)-phenyl]-acetic acid Example 19 6.98
[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazine-3-
sulfonyl)-phenyl]-acetic acid Example 20 7.04
2-[3,5-Dichloro-4-(5-isopropyl-6- oxo-1,6-dihydro-pyridazine-3-
sulfonyl)-phenyl]-3,5-dioxo- 2,3,4,5-tetrahydro-[1,2,4]triazine-
6-carbonitrile Example 21 3.21 2-[3,5-Dichloro-4-(5-isopropyl-6-
oxo-1,6-dihydro-pyridazine-3- sulfonyl)-phenyl]-2H-
[1,2,4]triazine-3,5-dione Example 22 0.136
2-[3,5-Dichloro-4-(5-isopropyl-1- methyl-6-oxo-1,6-dihydro-
pyridazin-3-ylmethyl)-phenyl]- 3,5-dioxo-2,3,4,5-tetrahydro-
[1,2,4]triazine-6-carbonitrile
It is to be understood that the invention is not limited to the
particular embodiments of the invention described above, as
variations of the particular embodiments may be made and still fall
within the scope of the appended claims.
* * * * *