U.S. patent application number 16/229904 was filed with the patent office on 2019-07-11 for diarylhydantoin compounds.
This patent application is currently assigned to The Regents of the University of California. The applicant listed for this patent is The Regents of the University of California. Invention is credited to Charlie D. CHEN, Michael JUNG, Samedy OUK, Charles L. SAWYERS, Chris TRAN, Derek WELSBIE, John WONGVIPAT, Dongwon YOO.
Application Number | 20190210974 16/229904 |
Document ID | / |
Family ID | 37590460 |
Filed Date | 2019-07-11 |
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United States Patent
Application |
20190210974 |
Kind Code |
A1 |
SAWYERS; Charles L. ; et
al. |
July 11, 2019 |
DIARYLHYDANTOIN COMPOUNDS
Abstract
The present invention relates to diarylhydantoin compounds,
including diarylthiohydantoins, and methods for synthesizing them
and using them in the treatment of hormone refractory prostate
cancer.
Inventors: |
SAWYERS; Charles L.; (New
York, NY) ; JUNG; Michael; (Los Angeles, CA) ;
CHEN; Charlie D.; (Shanghai, CN) ; OUK; Samedy;
(San Diego, CA) ; WELSBIE; Derek;
(Lutherville-Timonium, MD) ; TRAN; Chris; (New
York, NY) ; WONGVIPAT; John; (New York, NY) ;
YOO; Dongwon; (Los Angeles, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
The Regents of the University of California |
Oakland |
CA |
US |
|
|
Assignee: |
The Regents of the University of
California
Oakland
CA
|
Family ID: |
37590460 |
Appl. No.: |
16/229904 |
Filed: |
December 21, 2018 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15655458 |
Jul 20, 2017 |
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16229904 |
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14496973 |
Sep 25, 2014 |
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15655458 |
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14137991 |
Dec 20, 2013 |
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14496973 |
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13448964 |
Apr 17, 2012 |
9126941 |
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14137991 |
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12708523 |
Feb 18, 2010 |
8183274 |
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13448964 |
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11433829 |
May 15, 2006 |
7709517 |
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12708523 |
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60786837 |
Mar 29, 2006 |
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60756552 |
Jan 6, 2006 |
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60750351 |
Dec 15, 2005 |
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60680835 |
May 13, 2005 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 13/08 20180101;
A61P 35/00 20180101; C07D 233/86 20130101; A61P 17/10 20180101;
C07D 233/74 20130101; C07D 233/70 20130101; C07D 235/02 20130101;
C07D 471/10 20130101; A61P 17/14 20180101 |
International
Class: |
C07D 235/02 20060101
C07D235/02; C07D 233/86 20060101 C07D233/86; C07D 233/70 20060101
C07D233/70; C07D 233/74 20060101 C07D233/74; C07D 471/10 20060101
C07D471/10 |
Goverment Interests
[0002] This invention was made with Government support under Grant
No. W81XWH-04-1-0129 awarded by the United States Army, Medical
Research and Materiel Command and under Grant No. CA092131 awarded
by the National Institutes of Health. The Government has certain
rights in the invention.
Claims
1.-51. (canceled)
52. The compound having the formula ##STR00238##
Description
[0001] This application is a continuation of U.S. application Ser.
No. 15/655,458, filed Jul. 20, 2017, which is a continuation of
U.S. application Ser. No. 14/496,973, filed Sep. 25, 2014, which is
a continuation of U.S. application Ser. No. 14/137,991, filed Dec.
20, 2013, which is a continuation of U.S. application Ser. No.
13/448,964, filed Apr. 17, 2012, which is a continuation of U.S.
application Ser. No. 12/708,523, filed Feb. 18, 2010, which is a
divisional of U.S. application Ser. No. 11/433,829, filed May 15,
2006, which claims the benefit of U.S. Provisional Application No.
60/786,837, filed Mar. 29, 2006, U.S. Provisional Application No.
60/756,552, filed Jan. 6, 2006, U.S. Provisional Application No.
60/750,351, filed Dec. 15, 2005, and U.S. Provisional Application
No. 60/680,835, filed May 13, 2005, the specifications of which are
hereby incorporated by reference in their entirety.
FIELD OF THE INVENTION
[0003] The present invention relates to diarylhydantoin compounds
including diarylthiohydantoins, and methods for synthesizing them
and using them in the treatment of hormone refractory prostate
cancer.
BACKGROUND OF THE INVENTION
[0004] Prostate cancer is the most common incidence of cancer and
the second leading cause of cancer death in Western men. When the
cancer is confined locally, the disease can be cured by surgery or
radiation. However, 30% of such cancer relapses with distant
metastatic disease and others have advanced disease at diagnoses.
Advanced disease is treated by castration and/or administration of
antiandrogens, the so-called androgen deprivation therapy.
Castration lowers the circulating levels of androgens and reduces
the activity of androgen receptor (AR). Administration of
antiandrogens blocks AR function by competing away androgen
binding, therefore, reducing the AR activity. Although initially
effective, these treatments quickly fail and the cancer becomes
hormone refractory.
[0005] Recently, overexpression of AR has been identified and
validated as a cause of hormone refractory prostate cancer. See
Chen, C. D., Welsbie, D. S., Tran, C., Baek, S. H., Chen, R.,
Vessella, R., Rosenfeld, M. G., and Sawyers, C. L., Molecular
determinants of resistance to antiandrogen therapy, Nat. Med., 10:
33-39, 2004, which is hereby incorporated by reference.
Overexpression of AR is sufficient to cause progression from
hormone sensitive to hormone refractory prostate cancer, suggesting
that better AR inhibitors than the current drugs can slow the
progression of prostate cancer. It was demonstrated that AR and its
ligand binding are necessary for growth of hormone refractory
prostate cancer, indicating that AR is still a target for this
disease. It was also demonstrated that overexpression of AR
converts anti-androgens from antagonists to agonists in hormone
refractory prostate cancer (an AR antagonist inhibits AR activity
and an AR agonist stimulates AR activity). Data from this work
explains why castration and anti-androgens fail to prevent prostate
cancer progression and reveals unrecognized properties of hormone
refractory prostate cancer.
[0006] Bicalutamide (brand name: Casodex) is the most commonly used
anti-androgen. While it has an inhibitory effect on AR in hormone
sensitive prostate cancer, it fails to suppress AR when cancer
becomes hormone refractory. Two weaknesses of current antiandrogens
are blamed for the failure to prevent prostate cancer progression
from the hormone sensitive stage to the hormone refractory disease
and to effectively treat hormone refractory prostate cancer. One is
their weak antagonistic activities and the other is their strong
agonistic activities when AR is overexpressed in hormone refractory
prostate cancer. Therefore, better AR inhibitors with more potent
antagonistic activities and minimal agonistic activities are needed
to delay disease progression and to treat the fatal hormone
refractory prostate cancer.
[0007] Nonsteroidal anti-androgens, such as bicalutamide, have been
preferred over steroidal compounds for prostate cancer because they
are more selective and have fewer side effects. This class of
compounds has been described in many patents such as U.S. Pat. Nos.
4,097,578, 5,411,981, 5,705,654, PCT International Applications WO
97/00071 and WO 00/17163, and U.S. Published Patent Application
Number 2004/0009969, all of which are hereby incorporated by
reference.
[0008] U.S. Pat. No. 5,434,176 includes broad claims which
encompass a very large number of compounds, but synthetic routes
are only presented for a small fraction of these compounds and
pharmacological data are only presented for two of them, and one
skilled in the art could not readily envision other specific
compounds.
[0009] Because the mechanism of hormone refractory prostate cancer
was not known, there was no biological system to test these
compounds described in these patents for their effect on hormone
refractory prostate cancer. Particularly, the ability of AR
overexpression in hormone refractory prostate cancer to switch
inhibitors from antagonists to agonists was not recognized. Some
new properties of hormone refractory prostate cancer are reported
in PCT applications US04/42221 and US05/05529, which are hereby
incorporated by reference. PCT International Application US05/05529
presented a methodology for identifying androgen receptor
antagonist and agonist characteristics of compounds. However, for
each compound produced, the time consuming process of determining
the antagonist and agonist characteristics of a compound must be
determined. That is, there is no method to accurately predict
characteristics relevant to treating prostate cancer from the
chemical structure of a compound alone.
[0010] There is a need for new thiohydantoin compounds having
desirable pharmacological properties, and synthetic pathways for
preparing them. Because activities are sensitive to small
structural changes, one compound may be effective in treating
prostate cancer, whereas a second compound may be ineffective, even
if it differs from the first compound only slightly, say by the
replacement of a single substituent.
[0011] Identification of compounds which have high potency to
antagonize the androgen activity, and which have minimal agonistic
activity should overcome hormone refractory prostate cancer (HRPC)
and avoid or slow down the progression of hormone sensitive
prostate cancer (HSPC). Therefore, there is a need in the art for
the identification of selective modulators of the androgen
receptor, such as modulators which are non-steroidal, non-toxic,
and tissue selective.
SUMMARY OF THE INVENTION
[0012] The invention provides a series of compounds having strong
antagonistic activities with minimal agonistic activities against
AR. These compounds inhibit the growth of hormone refractory
prostate cancer.
[0013] The invention includes a compound having the formula
##STR00001##
wherein X is selected from the group consisting of trifluoromethyl
and iodo, wherein W is selected from the group consisting of O and
NR5, wherein R5 is selected from the group consisting of H, methyl,
and
##STR00002##
wherein D is S or O and E is N or O and G is alkyl, aryl,
substituted alkyl or substituted aryl; or D is S or O and E-G
together are C1-C4 lower alkyl,
[0014] wherein R1 and R2 together comprise eight or fewer carbon
atoms and are selected from the group consisting of alkyl,
substituted alkyl including haloalkyl, and, together with the
carbon to which they are linked, a cycloalkyl or substituted
cycloalkyl group,
[0015] wherein R3 is selected from the group consisting of
hydrogen, halogen, methyl, C1-C4 alkoxy, formyl, haloacetoxy,
trifluoromethyl, cyano, nitro, hydroxyl, phenyl, amino,
methylcarbamoyl, methoxycarbonyl, acetamido, methanesulfonamino,
methanesulfonyl, 4-methanesulfonyl-1-piperazinyl, piperazinyl, and
C1-C6 alkyl or alkenyl optionally substituted with hydroxyl,
methoxycarbonyl, cyano, amino, amido, nitro, carbamoyl, or
substituted carbamoyl including methylcarbamoyl, dimethylcarbamoyl,
and hydroxyethylcarbamoyl,
[0016] wherein R4 is selected from the group consisting of
hydrogen, halogen, alkyl, and haloalkyl, and
[0017] wherein R3 is not methylaminomethyl or
dimethylaminomethyl.
[0018] R5 may be
##STR00003##
[0019] The compound may have the formula
##STR00004##
wherein R3 is selected from the group consisting of hydroxy,
methylcarbamoyl, methylcarbamoylpropyl, methylcarbamoylethyl,
methylcarbamoylmethyl, methylsulfonecarbamoylpropyl,
methylaminomethyl, dimethylaminomethyl, methylsulfonyloxymethyl,
carbamoylmethyl, carbamoylethyl, carboxymethyl,
methoxycarbonylmethyl, methanesulfonyl,
4-cyano-3-trifluoromethylphenylcarbamoylpropyl, carboxypropyl,
4-methanesulfonyl-1-piperazinyl, piperazinyl, methoxy carbonyl,
3-cyano-4-trifluoromethylphenylcarbamoyl,
hydroxyethylcarbamoylethyl, and hydroxyethoxycarbonylethyl, and
[0020] wherein R10 and R11 are both H or, respectively, F and H, or
H and F. In certain embodiments, R10 and R11 may both be H or,
respectively, F and H. R3 may be methylcarbamoyl.
[0021] In some embodiments, R1 and R2 are independently methyl or,
together with the carbon to which they are linked, a cycloalkyl
group of 4 to 5 carbon atoms, and R3 is selected from the group
consisting of carbamoyl, alkylcarbamoyl, carbamoylalkyl, and
alkylcarbamoylalkyl, and R4 is H or F or R4 is 3-fluoro.
[0022] In other embodiments, R1 and R2 are independently methyl or,
together with the carbon to which they are linked, a cycloalkyl
group of 4 to 5 carbon atoms, R3 is selected from the group
consisting of cyano, hydroxy, methylcarbamoyl,
methylcarbamoyl-substituted alkyl,
methylsulfonecarbamoyl-substituted alkyl, methylaminomethyl,
dimethylaminomethyl, methylsulfonyloxymethyl, methoxycarbonyl,
acetamido, methanesulfonamido, carbamoyl-substituted alkyl,
carboxymethyl, methoxycarbonylmethyl, methanesulfonyl,
4-cyano-3-trifluoromethylphenylcarbamoyl-substituted alkyl,
carboxy-substituted alkyl,
4-(1,1-dimethylethoxy)carbonyl)-1-piperazinyl,
4-methanesulfonyl-1-piperazinyl, piperazinyl,
hydroxyethylcarbamoyl-substituted alkyl,
hydroxyethoxycarbonyl-substituted alkyl, and
3-cyano-4-trifluoromethylphenylcarbamoyl, and R4 is F.
[0023] Compounds of the invention may have the formula
##STR00005##
wherein R3 is selected from the group consisting of methylcarbonyl,
methoxycarbonyl, acetamido, and methanesulfonamido, and R4 is
selected from the group consisting of F and H.
[0024] Compounds of the invention may have the formula
##STR00006##
wherein R4 is selected from the group consisting of F and H.
[0025] In embodiments of the invention, wherein R1 and R2 together
with the carbon to which they are linked are
##STR00007##
[0026] Compounds of the invention may be those listed in Tier 1,
Tier 2, Tier 3, and/or Tier 4, below. Particular compounds of the
invention include
##STR00008##
[0027] The invention also provides a pharmaceutical composition
comprising a therapeutically effective amount of a compound
according to any of the preceding compounds or a pharmaceutically
acceptable salt thereof, and a pharmaceutically acceptable carrier
or diluent.
[0028] The invention encompasses a method for treating a
hyperproliferative disorder comprising administering such a
pharmaceutical composition to a subject in need of such treatment,
thereby treating the hyperproliferative disorder. The
hyperproliferative disorder may be hormone refractory prostate
cancer. The dosage may be in the range of from about 0.001 mg per
kg body weight per day to about 100 mg per kg body weight per day,
about 0.01 mg per kg body weight per day to about 100 mg per kg
body weight per day, about 0.1 mg per kg body weight per day to
about 10 mg per kg body weight per day, or about 1 mg per kg body
weight per day.
[0029] The compound may be administered by intravenous injection,
by injection into tissue, intraperitoneally, orally, or nasally.
The composition may have a form selected from the group consisting
of a solution, dispersion, suspension, powder, capsule, tablet,
pill, time release capsule, time release tablet, and time release
pill.
[0030] The administered compound may be selected from the group
consisting of RD162', RD162'', RD 169, or RD170, or a
pharmaceutically acceptable salt thereof. The administered compound
may be RD162 or a pharmaceutically acceptable salt thereof.
[0031] The invention provides a method of synthesizing a diaryl
compound of formula:
##STR00009##
comprising mixing Compound I
##STR00010##
with Compound II
##STR00011##
in a first polar solvent to form a mixture, heating the mixture,
adding a second polar solvent, the same as or different from the
first polar solvent, and an aqueous acid to the mixture, refluxing
the mixture, cooling the mixture and combining with water, and
separating the diaryl compound from the mixture, wherein R51
comprises an alkyl chain of from 1 to 4 carbon atoms, R52 is
selected from the group consisting of cyano, hydroxy,
methylcarbamoyl, methylcarbamoyl-substituted alkyl,
methylsulfonecarbamoyl-substituted alkyl, methylaminomethyl,
dimethylaminomethyl, methylsulfonyloxymethyl, methoxycarbonyl,
3-cyano-4-trifluoromethylphenylcarbamoyl, carbamoyl-substituted
alkyl, carboxymethyl, methoxycarbonylmethyl, methanesulfonyl,
4-cyano-3-trifluoromethylphenylcarbamoyl-substituted alkyl,
carboxy-substituted alkyl, 4-methanesulfonyl-1-piperazinyl,
piperazinyl, hydroxyethylcarbamoyl-substituted alkyl, and
hydroxyethoxycarbonyl-substituted alkyl, and R53 is selected from
the group consisting of F and H.
[0032] R51 may comprise an alkyl chain of from 1 to 2 carbon atoms,
R52 may be selected from the group consisting of carbamoyl and
methylcarbamoyl, and R53 may be F.
[0033] The invention provides methods of synthesizing a compound of
formula:
##STR00012##
comprising mixing 4-isothiocyanato-2-trifluoromethylbenzonitrile
and N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide in
dimethylformamide to form a first mixture, heating the first
mixture to form a second mixture, adding alcohol and acid to the
second mixture to form a third mixture, refluxing the third mixture
to form a fourth mixture, cooling the fourth mixture, combining the
fourth mixture with water and extracting an organic layer;
isolating the compound from the organic layer.
[0034] Likewise, the invention provides a method of synthesizing
RD162' comprising mixing
N-Methyl-2-fluoro-4-(1,1-dimethyl-cyanomethyl)-aminobenzamide and
4-Isothiocyanato-2-trifluoromethylbenzonitrile in DMF and heating
to form a first mixture, and processing as above.
[0035] The invention also provides a method of synthesizing
RD162'', comprising mixing
N-Methyl-2-fluoro-4-(1-cyanocyclopentyl)aminobenzamide,
4-isothiocyanato-2-trifluoromethyl benzonitrile, and DMF and
heating under reflux to form a first mixture, and processing as
above.
[0036] The invention further provides a method of synthesizing
RD169, comprising mixing N,N-Dimethyl
4-[4-(1-cyanocyclobutylamino)phenyl]butanamide,
4-isothiocyanato-2-trifluoromethyl benzonitrile, and DMF and
heating under reflux to form a first mixture; and processing as
above.
[0037] The invention provides a method of synthesizing RD170,
comprising mixing DMSO, dichloromethane, and oxalyl chloride to
form a first mixture, adding
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)butanamide to the first mixture to form a
second mixture; adding triethylamine to the second mixture to form
a third mixture; warming the third mixture and quenching with
aqueous NH.sub.4Cl to form a fourth mixture; extracting an organic
layer from the fourth mixture; and isolating the compound from the
organic layer.
[0038] Further compounds according to the invention have the
formula
##STR00013##
wherein R5 is CN or NO2 or SO2R11, wherein R6 is CF3, alkyl,
substituted alkyl, alkenyl, substituted alkenyl, alkynyl,
substituted alkynyl, halogenated alkyl, halogenated alkenyl,
halogenated alkynyl, halogen, wherein A is sulfur (S) or oxygen
(O), wherein B is O or S or NR8, wherein R8 is selected from the
group consisting of H, methyl, aryl, substituted aryl, alkyl,
substituted alkyl, alkenyl, substituted alkenyl, alkynyl,
substituted alkynyl, arylalkyl, arylalkenyl, arylalkynyl,
heterocyclic aromatic or non-aromatic, substituted heterocyclic
aromatic or non-aromatic, cycloalkyl, substituted cycloalkyl,
SO2R11, NR11R12, (CO)OR11, (CO)NR11R12, (CO)R11, (CS)R11,
(CS)NR11R12, (CS)OR11,
##STR00014##
wherein D is S or O and E is N or O and G is alkyl, aryl,
substituted alkyl or substituted aryl; or D is S or O and E-G
together are C1-C4 lower alkyl,
[0039] wherein R1 and R2 are independently alkyl, haloalkyl,
hydrogen, aryl, substituted alkyl, alkenyl, substituted alkenyl,
alkynyl, substituted alkynyl, halogenated alkenyl, halogenated
alkynyl, arylalkyl, arylalkenyl, arylalkynyl, heterocylic aromatic
or non-aromatic, substituted heterocyclic aromatic or non-aromatic,
cycloalkyl, substituted cycloalkyl, or R1 and R2 are connected to
form a cycle which can be heterocyclic, substituted heterocyclic,
cycloalkyl, substituted cycloalkyl,
##STR00015##
[0040] wherein X is carbon or nitrogen and can be at any position
in the ring, and
[0041] wherein R3, R4, and R7 are independently selected from the
group consisting of hydrogen, halogen, methyl, methoxy, formyl,
haloacetoxy, trifluoromethyl, cyano, nitro, hydroxyl, phenyl,
amino, methylcarbamoyl, methylcarbamoyl-substituted alkyl,
dimethylcarbamoyl-substituted alkyl, methoxycarbonyl, acetamido,
methanesulfonamino, carbamoyl-substituted alkyl, methanesulfonyl,
4-methanesulfonyl-1-piperazinyl, piperazinyl,
hydroxyethylcarbamoyl-substituted alkyl, hydroxyl-substituted
alkyl, hydroxyl-substituted alkenyl, carbamoyl-substituted alkenyl,
methoxycarbonyl-substituted alkyl, cyano-substituted alkyl,
##STR00016##
aryl, substituted aryl, alkyl, substituted alkyl, alkenyl,
substituted alkenyl, alkynyl, substituted alkynyl, halogenated
alkenyl, halogenated alkynyl, SO2R11, NR11R12, NR12(CO)OR11,
NH(CO)NR11R12, NR12 (CO)R11, O(CO)R11, O(CO)OR11, O(CS)R11, NR12
(CS)R11, NH(CS)NR11R12, NR12 (CS)OR11, arylalkyl, arylalkenyl,
arylalkynyl, heterocyclic aromatic or non-aromatic, substituted
heterocyclic aromatic or non-aromatic, cycloalkyl, substituted
cycloalkyl, haloalkyl, methylsulfonecarbamoyl-substituted alkyl,
methylaminomethyl, dimethylaminomethyl, methylsulfonyloxymethyl,
methoxycarbonyl, acetamido, methanesulfonamido,
carbamoyl-substituted alkyl, carboxymethyl, methoxycarbonylmethyl,
methanesulfonyl,
4-cyano-3-trifluoromethylphenylcarbamoyl-substituted alkyl,
carboxy-substituted alkyl,
4-(1,1-dimethylethoxy)carbonyl)-1-piperazinyl,
hydroxyethylcarbamoyl-substituted alkyl,
hydroxyethoxycarbonyl-substituted alkyl,
3-cyano-4-trifluoromethylphenylcarbamoyl,
[0042] wherein R11 and R12 are independently hydrogen, aryl,
aralkyl, substituted aralkyl, alkyl, substituted alkyl, alkenyl,
substituted alkenyl, alkynyl, substituted alkynyl, halogenated
alkyl, halogenated alkenyl, halogenated alkynyl, arylalkyl,
arylalkenyl, arylalkynyl, heterocyclic aromatic or non-aromatic,
substituted heterocyclic aromatic or non-aromatic, cycloalkyl, or
substituted cycloalkyl, or R11 and R12 can be connected to form a
cycle which can be heterocyclic aromatic or non-aromatic,
substituted heterocyclic aromatic, cycloalkyl, or substituted
cycloalkyl.
[0043] Such compounds have substantial androgen receptor antagonist
activity and no substantial agonist activity on hormone refractory
prostate cancer cells.
[0044] The invention encompasses a method comprising providing at
least one such compound, measuring inhibition of androgen receptor
activity for the compound and determining if the inhibition is
above a first predetermined level, measuring stimulation of
androgen receptor activity in hormone refractory cancer cells for
the compound and determining if the stimulation is below a second
predetermined level, and selecting the compound if the inhibition
is above the first predetermined level and the stimulation is below
the second predetermined level. The predetermined levels may be
those of bicalutamide. The step of measuring inhibition may
comprise measuring inhibitory concentration (IC50) in an AR
response reporter system or a prostate specific antigen secreting
system. The step of measuring stimulation may comprise measuring
fold induction by increasing concentrations in an AR response
reporter system or a prostate specific antigen secreting system.
The method of measuring inhibition and/or stimulation may comprise
measuring an effect of the compound on tumor growth in an
animal.
BRIEF DESCRIPTION OF THE DRAWINGS
[0045] The following Figures present the results of pharmacological
examination of certain compounds.
[0046] FIG. 1 is a graph depicting that bicalutamide displays an
agonistic effect on LNCaP-AR. Agonistic activities of bicalutamide
in AR-overexpressed hormone refractory prostate cancer. LNCaP cells
with overexpressed AR were treated with increasing concentrations
of DMSO as vehicle or bicalutamide in the absence of R1881.
Activities of AR response reporter were measured.
[0047] FIG. 2 is a graph depicting an antagonistic assay of
bicalutamide on LNCaP-AR. Agonistic activities of bicalutamide in
hormone sensitive prostate cancer. LNCaP cells were treated with
increasing concentrations of DMSO as vehicle or bicalutamide in the
absence of R1881. Activities of AR response reporter were
measured.
[0048] FIG. 3 is a graph depicting the effect of compounds on
LNCaP-AR.
[0049] FIG. 4 is a graph depicting the effect of compounds on
LNCaP-AR.
[0050] FIG. 5 is a graph depicting the inhibition effect on
LNCaP-AR.
[0051] In FIGS. 6-10, example 5-3b is RD7 and example 7-3b is
RD37.
[0052] FIG. 6. Inhibition on growth of AR-overexpressed LNCaP
cells. Androgen starved LNCaP cells with overexpressed AR were
treated with increasing concentrations of DMSO as vehicle or test
substances in the presence of 100 pM of R1881. After 4 days of
incubation, cell growth was measured by MTS assay.
[0053] FIGS. 7A and 7B depict the inhibitory effect on growth of
AR-overexpressed LNCaP xenograft model. Mice with established LN-AR
xenograft tumors were randomized and treated with indicated
compounds orally once daily. Tumor size was measured by caliber.
FIG. 7A represents tumor volume in mice treated with 1 mg per kg of
bicaluta mide, example 7-3b, or vehicle for 44 days. FIG. 7B
represents tumor volume in mice treated with vehicle, 0.1, 1, or 10
mg per kg of example 7-3b for 44 days.
[0054] FIG. 8. Inhibitory effect on PSA expression of
AR-overexpressed LNCaP xenograft model. Mice were treated with
vehicle, 0.1, 1, or 10 mg per kg of example 7-3b for 44 days orally
once daily. The tumors were taken out from the mice after 44 days
of treatment, tumor lysate was extracted, and PSA level in tissue
lysate was determined by ELISA.
[0055] FIGS. 9A and 9B depict inhibitory effect on growth and PSA
of hormone refractory LAPC4 xenograft model. Mice with established
tumors were randomized and treated with 1 mg per kg of
bicalutamide, example 7-3b, or vehicle for 17 days orally once
daily. FIG. 9A depicts tumor size measured by caliber. FIG. 9B
depicts PSA level after the tumors were taken out from the mice
after 17 days of treatment, tumor lysate was extracted, and PSA
level in tissue lysate was determined by ELISA.
[0056] FIG. 10. Inhibitory effect on growth of hormone sensitive
prostate cancer cells. Androgen starved LNCaP cells were treated
with increasing concentrations of DMSO as vehicle or test
substances in the presence of 1 pM of R1881. After 4 days of
incubation, cell growth was measured by MTS assay.
[0057] FIG. 11 is a graph of tumor size. AR overexpressing LNCaP
cells were injected in the flanks of castrated SCID mice,
subcutaneously. When tumors reached about 100 cubic mm, they were
randomized into five groups. Each group had nine animals. After
they reached this tumor volume, they were given orally with either
vehicle, bicalutamide or RD162 at 10 or 50 mg/kg everyday. The
tumors were measured three-dimensionally, width, length and depth,
using a caliper.
[0058] FIGS. 12A through 12D depict experimental results of tumor
size. FIGS. 12A and 12B depict images of the animals at day 18
obtained via an optical CCD camera, 3 hours after last dose of
treatment. A ROI was drawn over the tumor for luciferase activity
measurement in photon/second. FIGS. 12C and 12D depict the ROIs
measurements.
[0059] FIG. 13 is a graph depicting the pharmacokinetic curves of
RD162 from intravenous (upper curve) and oral administration (lower
curve).
[0060] FIG. 14 is a graph depicting PSA absorbance measured for
LN-AR cells after treatment with various doses of several
compounds.
[0061] FIG. 15A presents a table providing several characteristics
of compounds. FIG. 15B presents a graph providing the
pharmacokinetic characteristics of several compounds in terms of
compound serum concentration as a function of time.
[0062] FIG. 16 is a chart depicting prostate weight after treatment
with various compounds. 10, 25, or 50 mg of compound per kilogram
body weight were administered per day, as indicated by the label of
a bar. The compounds were administered to healthy FVB mice. After
treatment with compound for 14 days, the urogenital tract weight
was determined by removing and weighing the semi-vesicles,
prostate, and bladder. Three mice were administered a given
compound to obtain the data presented by a bar in the chart. A set
of mice was not treated with a compound: data are presented in the
bar labeled "untreated". Another set of mice was treated only with
vehicle solution: data are presented in the bar labeled
"vehicle".
[0063] FIG. 17 is a graph presenting a PSA assay performed along
with the experimental protocol presented in FIG. 6.
[0064] FIG. 18 is a graph presenting the effect of various dose
regimens of RD162 on tumor volume.
[0065] FIG. 19A presents images showing the rate of photon emission
associated with luciferase activity at day 0 of treatment with
RD162 at doses of 0.1, 1, and 10 mg per kilogram body weight per
day and without treatment with RD162. FIG. 19B presents images
showing the rate of photon emission associated with luciferase
activity at day 17 after treatment with RD162 at doses of 0.1, 1,
and 10 mg per kilogram body weight per day and without treatment
with RD162. FIG. 19C is a graph that compares the rate of photon
emission associated with luciferase activity at day 0 and day 17
after treatment at doses of 0.1, 1, and 10 mg per kilogram body
weight per day and without treatment with RD162.
[0066] FIGS. 20A through 20C present the results of an experiment
in which SCID mice were injected with the LN-AR (HR) cell line to
induce tumor growth. One set of mice was treated with the compound
RD162 at a dose of 10 mg per kilogram body weight per day; the
other set of mice was treated only with vehicle solution. FIG. 20A
depicts relative tumor volume as a function of time shown for each
set of mice. FIG. 20B depicts images of each set of mice with
photon emission associated with luciferase activity at day 31 shown
as color contours. FIG. 20C depicts the rate of photon emission
associated with luciferase activity shown at several times for each
set of mice.
[0067] FIGS. 21A and 21B are graphs that depict PSA absorbance
associated with LN-AR cells treated with various concentrations of
RD162, RD162', RD162'', RD169, and RD170 and vehicle solution.
[0068] FIG. 22 is a graph presenting PSA absorbance associated with
LN-CaP cells treated with various concentrations of RD37, RD131,
RD162, bicalutamide, and DMSO.
[0069] FIGS. 23A and 23B present results of an experiment conducted
with wild type nontransgenic mice (WT), castrated luciferase
transgenic mice (Cast), and non-castrated luciferase transgenic
mice (Intact). Data are shown for castrated luciferase transgenic
mice treated with an implanted testosterone pellet yielding 12.5 mg
per kilogram body weight with a 90 day release period (T/Cast), and
data are shown for non-castrated luciferase transgenic mice treated
with an implanted testosterone pellet yielding 12.5 mg per kilogram
body weight with a 90 day release period (Intact+T). Data are shown
for castrated luciferase transgenic mice treated with the implanted
testosterone pellet and with bicalutamide (BIC+T/Cast) or with
RD162 (RD162+T/Cast) at 10 mg per kilogram body weight per day.
FIG. 23A presents urogenital tract weight at 14 days. FIG. 23B
presents photon emission rate at 14 days. In all cases, a hormone
refractory disease state was not induced.
[0070] FIG. 24 is a graph of luciferase activity of the LIAR cell
line dosed with various compounds administered at concentrations
ranging from 125 nmol to 1000 nmol.
[0071] FIG. 25 is a graph of luciferase activity for the LN/AR cell
line for various compounds administered at concentrations ranging
from 1.25 to 10 .mu.mol.
[0072] FIG. 26 is a graph of luciferase activity for the 4AR cell
line for various compounds administered at concentrations ranging
from 1.25 to 10 .mu.mol.
[0073] FIG. 27 is a graph of PSA levels for the 1AR cell line for
various compounds administered at concentrations ranging from 1.25
to 10 .mu.mol.
[0074] FIG. 28 is a graph of PSA levels for the LN/AR cell line for
various compounds administered at concentrations ranging from 125
nmol to 1000 nmol.
[0075] FIG. 29 is a graph of luciferase activity for various
compounds administered at concentrations ranging from 125 nmol to
1000 nmol.
[0076] FIG. 30 is an illustration of an X-ray structure of the
compound [RD93] (also designated as 35a).
DETAILED DESCRIPTION
[0077] Embodiments of the invention are discussed in detail below.
In describing embodiments, specific terminology is employed for the
sake of clarity. However, the invention is not intended to be
limited to the specific terminology so selected. A person skilled
in the relevant art will recognize that other equivalent parts can
be employed and other methods developed without parting from the
spirit and scope of the invention. All references cited herein are
incorporated by reference as if each had been individually
incorporated.
Synthesis of Diarylhydantoin Compounds
[0078] The invention provides for synthesis of diarylthiohydantoin
compound having the formula
##STR00017##
with R71 including an alkyl chain of from 1 to 4 carbon atoms. For
example, R72 can be carbamoyl, e.g., --(CO)NH.sub.2, or
methylcarbamoyl, e.g., --(CO)NHCH.sub.3. An amide group bonded at
the carbon atom of the carbonyl to another structure is termed a
carbamoyl substituent. For example, R73 can be a fluorine or a
hydrogen atom. That is, a fluorine atom can be attached to any one
of the carbons of the right-hand aryl ring which are not bonded to
the R72 substituent or the nitrogen atom. Alternatively, no
fluorine atom can be attached to the carbons of the right-hand aryl
ring which are not bonded to the R72 substituent or the nitrogen
atom. For example, a hydrogen atom can be attached to each of the
carbons of the right-hand aryl ring which are not bonded to the R72
substituent or the nitrogen atom.
[0079] For example, as further presented below (see, for example,
FIGS. 3, 5, 11-13), the compound having the formula
##STR00018##
[RD162]
[0080] exhibited surprisingly potent antagonistic activities with
minimal agonistic activities for overexpressed AR in hormone
refractory prostate cancer.
[0081] A list of several compounds according to this invention is
presented in Tables 5-11. The compounds are grouped into tiers,
with Tier 1 to Tier 3 compounds being expected to be superior to
bicalutamide for the treatment of prostate cancer, Tier 4 compounds
being comparable to bicalutamide in effectiveness, and Tier 5 and
Tier 6 compounds being worse than bicalutamide for the treatment of
prostate cancer. A more detailed description of the protocol used
to rank the compounds into tiers is presented below.
Definitions
[0082] As used herein, the term "alkyl" denotes branched or
unbranched hydrocarbon chains, preferably having about 1 to about 8
carbons, such as, methyl, ethyl, n-propyl, iso-propyl, n-butyl,
sec-butyl, iso-butyl, tert-butyl, 2-methylpentyl pentyl, hexyl,
isohexyl, heptyl, 4,4-dimethyl pentyl, octyl, 2,2,4-trimethylpentyl
and the like. "Substituted alkyl" includes an alkyl group
optionally substituted with one or more functional groups which may
be attached to such chains, such as, hydroxyl, bromo, fluoro,
chloro, iodo, mercapto or thio, cyano, alkylthio, heterocyclyl,
aryl, heteroaryl, carboxyl, carbalkoyl, alkyl, alkenyl, nitro,
amino, alkoxyl, amido, and the like to form alkyl groups such as
trifluoro methyl, 3-hydroxyhexyl, 2-carboxypropyl, 2-fluoroethyl,
carboxymethyl, cyanobutyl and the like.
[0083] Unless otherwise indicated, the term "cycloalkyl" as
employed herein alone or as part of another group includes
saturated or partially unsaturated (containing 1 or more double
bonds) cyclic hydrocarbon groups containing 1 to 3 rings, including
monocyclicalkyl, bicyclicalkyl and tricyclicalkyl, containing a
total of 3 to 20 carbons forming the rings, preferably 3 to 10
carbons, forming the ring and which may be fused to 1 or 2 aromatic
rings as described for aryl, which include cyclopropyl, cyclobutyl,
cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl and
cyclododecyl, cyclohexenyl. "Substituted cycloalkyl" includes a
cycloalkyl group optionally substituted with 1 or more substituents
such as halogen, alkyl, alkoxy, hydroxy, aryl, aryloxy, arylalkyl,
cycloalkyl, alkylamido, alkanoylamino, oxo, acyl,
arylcarbonylamino, amino, nitro, cyano, thiol and/or alkylthio
and/or any of the substituents included in the definition of
"substituted alkyl." For example,
##STR00019##
and the like.
[0084] Unless otherwise indicated, the term "alkenyl" as used
herein by itself or as part of another group refers to straight or
branched chain radicals of 2 to 20 carbons, preferably 2 to 12
carbons, and more preferably 2 to 8 carbons in the normal chain,
which include one or more double bonds in the normal chain, such as
vinyl, 2-propenyl, 3-butenyl, 2-butenyl, 4-pentenyl, 3-pentenyl,
2-hexenyl, 3-hexenyl, 2-heptenyl, 3-heptenyl, 4-heptenyl,
3-octenyl, 3-nonenyl, 4-decenyl, 3-undecenyl, 4-dodecenyl,
4,8,12-tetradecatrienyl, and the like. "Substituted alkenyl"
includes an alkenyl group optionally substituted with one or more
substituents, such as the substituents included above in the
definition of "substituted alkyl" and "substituted cycloalkyl."
[0085] Unless otherwise indicated, the term "alkynyl" as used
herein by itself or as part of another group refers to straight or
branched chain radicals of 2 to 20 carbons, preferably 2 to 12
carbons and more preferably 2 to 8 carbons in the normal chain,
which include one or more triple bonds in the normal chain, such as
2-propynyl, 3-butynyl, 2-butynyl, 4-pentynyl, 3-pentynyl,
2-hexynyl, 3-hexynyl, 2-heptynyl, 3-heptynyl, 4-heptynyl,
3-octynyl, 3-nonynyl, 4-decynyl, 3-undecynyl, 4-dodecynyl and the
like. "Substituted alkynyl" includes an alkynyl group optionally
substituted with one or more substituents, such as the substituents
included above in the definition of "substituted alkyl" and
"substituted cycloalkyl."
[0086] The terms "arylalkyl", "arylalkenyl" and "arylalkynyl" as
used alone or as part of another group refer to alkyl, alkenyl and
alkynyl groups as described above having an aryl substituent.
Representative examples of arylalkyl include, but are not limited
to, benzyl, 2-phenylethyl, 3-phenylpropyl, phenethyl, benzhydryl
and naphthylmethyl and the like. "Substituted arylalkyl" includes
arylalkyl groups wherein the aryl portion is optionally substituted
with one or more substituents, such as the substituents included
above in the definition of "substituted alkyl" and "substituted
cycloalkyl."
[0087] The terms "arylalkyl", "arylalkenyl" and "arylalkynyl" as
used alone or as part of another group refer to alkyl, alkenyl and
alkynyl groups as described above having an aryl substituent.
Representative examples of arylalkyl include, but are not limited
to, benzyl, 2-phenylethyl, 3-phenylpropyl, phenethyl, benzhydryl
and naphthylmethyl and the like. "Substituted arylalkyl" includes
arylalkyl groups wherein the aryl portion is optionally substituted
with one or more substituents, such as the substituents included
above in the definition of "substituted alkyl" and "substituted
cycloalkyl."
[0088] The term "halogen" or "halo" as used herein alone or as part
of another group refers to chlorine, bromine, fluorine, and
iodine.
[0089] The terms "halogenated alkyl", "halogenated alkenyl" and
"alkynyl" as used herein alone or as part of another group refers
to "alkyl", "alkenyl" and "alkynyl" which are substituted by one or
more atoms selected from fluorine, chlorine, bromine, fluorine, and
iodine.
[0090] Unless otherwise indicated, the term "aryl" or "Ar" as
employed herein alone or as part of another group refers to
monocyclic and polycyclic aromatic groups containing 6 to 10
carbons in the ring portion (such as phenyl or naphthyl including
1-naphthyl and 2-naphthyl) and may optionally include one to three
additional rings fused to a carbocyclic ring or a heterocyclic ring
(such as aryl, cycloalkyl, heteroaryl or cycloheteroalkyl
rings).
[0091] "Substituted aryl" includes an aryl group optionally
substituted with one or more functional groups, such as halo,
haloalkyl, alkyl, haloalkyl, alkoxy, haloalkoxy, alkenyl,
trifluoromethyl, trifluoromethoxy, alkynyl, cycloalkyl-alkyl,
cycloheteroalkyl, cycloheteroalkylalkyl, aryl, heteroaryl,
arylalkyl, aryloxy, aryloxyalkyl, arylalkoxy, alkoxycarbonyl,
arylcarbonyl, arylalkenyl, aminocarbonylaryl, arylthio,
arylsulfinyl, arylazo, heteroarylalkyl, heteroarylalkenyl,
heteroarylheteroaryl, heteroaryloxy, hydroxy, nitro, cyano, amino,
substituted amino wherein the amino includes 1 or 2 substituents
(which are alkyl, aryl or any of the other aryl compounds mentioned
in the definitions), thiol, alkylthio, arylthio, heteroarylthio,
arylthioalkyl, alkoxyarylthio, alkylcarbonyl, arylcarbonyl,
alkylaminocarbonyl, arylaminocarbonyl, alkoxycarbonyl,
aminocarbonyl, alkylcarbonyloxy, arylcarbonyloxy,
alkylcarbonylamino, arylcarbonylamino, arylsulfinyl,
arylsulfinylalkyl, arylsulfonylamino or arylsulfonaminocarbonyl
and/or any of the alkyl substituents set out herein.
[0092] Unless otherwise indicated, the term "heterocyclic" or
"heterocycle", as used herein, represents an unsubstituted or
substituted stable 5- to 10-membered monocyclic ring system which
may be saturated or unsaturated, and which consists of carbon atoms
and from one to four heteroatoms selected from N, O or S, and
wherein the nitrogen and sulfur heteroatoms may optionally be
oxidized, and the nitrogen heteroatom may optionally be
quaternized. The heterocyclic ring may be attached at any
heteroatom or carbon atom which results in the creation of a stable
structure. Examples of such heterocyclic groups include, but is not
limited to, piperidinyl, piperazinyl, oxopiperazinyl,
oxopiperidinyl, oxopyrrolidinyl, oxoazepinyl, azepinyl, pyrrolyl,
pyrrolidinyl, furanyl, thienyl, pyrazolyl, pyrazolidinyl,
imidazolyl, imidazolinyl, imidazolidinyl, pyridyl, pyrazinyl,
pyrimidinyl, pyridazinyl, oxazolyl, oxazolidinyl, isooxazolyl,
isoxazolidinyl, morpholinyl, thiazolyl, thiazolidinyl,
isothiazolyl, thiadiazolyl, tetrahydropyranyl, thiamorpholinyl,
thiamorpholinyl sulfoxide, thiamorpholinyl sulfone, and
oxadiazolyl. The term "heterocyclic aromatic" as used here in alone
or as part of another group refers to a 5- or 7-membered aromatic
ring which includes 1, 2, 3 or 4 hetero atoms such as nitrogen,
oxygen or sulfur and such rings fused to an aryl, cycloalkyl,
heteroaryl or heterocycloalkyl ring (e.g. benzothiophenyl,
indolyl), and includes possible N-oxides. "Substituted heteroaryl"
includes a heteroaryl group optionally substituted with 1 to 4
substituents, such as the substituents included above in the
definition of "substituted alkyl" and "substituted cycloalkyl."
Examples of heteroaryl groups include the following:
##STR00020##
and the like.
Example 1
4-isothiocyanato-2-trifluoromethylbenzonitrile, (1a)
[0093] 4-Amino-2-trifluoromethylbenzonitrile, (2.23 g, 12 mmol) was
added portionwise over 15 minutes into the well-stirred
heterogeneous mixture of thiophosgene (1 ml, 13 mmol) in water (22
ml) at room temperature. Stirring was continued for an additional 1
h. The reaction medium was extracted with chloroform (3.times.15
ml). The combined organic phase was dried over MgSO.sub.4 and
evaporated to dryness under reduced pressure to yield desired
product, 4-isothiocyanato-2-trifluoromethylbenzonitrile, (1a), as
brownish solid and was used as such for the next step (2.72 g, 11.9
mmol, 99%).
Example 2
2-1). (4-aminophenyl)carbamic acid tert-butyl ester, (2a)
[0094] An aqueous solution of potassium carbonate (1.52 g, 11 mmol
in 5 ml of water) was added to a solution of 1,4-diaminobenzene
(3.24 g, 30 mmol) in THF (30 ml) and DMF (10 ml). To this mixture
was added di-tert-butyl pyrocarbonate, Boc.sub.2O (2.18 g, 10
mmol), dropwise over 0.5 h. The reaction mixture was stirred for an
additional 4 h at room temperature. The mixture was then poured
into cold water (40 ml) and extracted with chloroform (3.times.50
ml). The combined organic phase was dried over MgSO.sub.4 and
concentrated to yield a brown residue which was subjected to flash
chromatography (dichloromethane/acetone, 4:1) to afford
(4-aminophenyl)carbamic acid tert-butyl ester, (2a) as a yellow
solid (1.98 g, 9.5 mmol, 95%) (yield based on Boc.sub.2O).
2-2). {4-[(1-cyano-1-methylethyl)amino]phenyl}carbamic acid
tert-butyl ester, 2b
[0095] The mixture of 2a (0.83 g, 4 mmol), acetone cyanohydrin (4
ml) and MgSO.sub.4 (2 g) was heated to 80.degree. C. and stirred
over 2.5 h. After cooling down to room temperature, compound 2b was
crystallized into water (30 ml). The solid was filtered and dried
to yield {4-[(1-cyano-1-methylethyl)amino]phenyl}carbamic acid
tert-butyl ester, 2b (1.08 g, 3.9 mmol, 98%).
2-3).
{4-[3-(4-cyano-3-trifluoromethylphenyl)-4-imino-5,5-dimethyl-2-thiox-
o-imidazolidin-1-yl]phenyl}carbamic acid tert-butyl ester, (2c)
[0096] Triethylamine (0.202 g, 2 mmol) was added to a solution of
1a (0.456 g, 2 mmol) and 2b (0.57 g, 2 mmol) in dry THF (5 ml). The
reaction mixture was stirred at room temperature for 15 h and then
concentrated to yield a dark residue which was subjected to flash
chromatography (ethyl ether/acetone, 97:3) to afford
{4-[3-(4-cyano-3-trifluoromethylphenyl)-4-imino-5,5-dimethyl-2-thioxo-imi-
dazolidin-1-yl]phenyl}carbamic acid tert-butyl ester, (2c) (0.15 g,
0.3 mmol, 15%).
2-4).
4-[3-(4-aminophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-
-trifluoromethylbenzonitrile, 2d, [RD9]
[0097] The mixture of 2c (0.15 g, 0.3 mmol) in HCl aq, 3N. (1 ml)
and methanol (4 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (5 ml) and extracted with dichloromethane (8 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane/acetone, 9:1) to yield
4-[3-(4-aminophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trif-
luoromethylbenzonitrile, 2d, [RD9] (0.118 g, 0.29 mmol, 97%) as a
yellow solid.
##STR00021##
[0098] .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 1.54 (s, 6H),
6.73-6.75 (m, 2H), 7.00-7.03 (m, 2H), 8.02 (dd, J.sub.1=8.2 Hz,
J.sub.2=1.8 Hz, 1H), 8.16 (d, J=1.8 Hz, 1H), 8.20 (d, J=8.2 Hz,
1H); .sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 22.7, 66.2, 109.1,
114.3, 114.9, 120.4, 122.0 (q, J=272.5 Hz), 127.0 (q, J=4.9 Hz),
130.4, 132.5 (q, J=33.0 Hz), 133.4, 135.6, 138.5, 149.2, 175.3,
180.4.
2-5).
4-[3-(4-azidophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-
-trifluoromethylbenzonitrile, 2e, [RD10]
[0099] An aqueous solution of sulfuric acid (25% wt, 1 ml) was
added to a solution of 2d (0.10 g, 0.25 mmol) in acetone (1 ml) at
-5.degree. C. An aqueous solution of NaNO.sub.2 (0.024 g, 0.35
mmol, in 0.5 ml of water) was added slowly the above mixture over
0.1 h. The reaction mixture was allowed to stir at -5.degree. C.
for an additional 1 h and then an aqueous solution of NaN.sub.3
(0.02 g, 0.3 mmol in 0.3 ml of water) was added dropwise. Upon
completion of the addition, the reaction medium was warmed to room
temperature and stirred for an additional 3 h. The product was
extracted with dichloromethane (3.times.5 ml). The combined organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield
4-[3-(4-azidophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trif-
luoromethylbenzonitrile, 2e, [RD10] (0.08 g, 0.18 mmol, 72%) as a
yellowish solid.
##STR00022##
[0100] .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 1.54 (s, 6H),
7.17-7.20 (m, 2H), 7.27-7.30 (m, 2H), 7.84 (dd, J.sub.1=8.3 Hz,
=1.8 Hz, 1H), 7.96 (d, J=1.8 Hz, 1H), 7.97 (d, J=8.3 Hz, 1H);
.sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 23.7, 66.4, 110.1,
114.8, 120.4, 122.1 (q, J=272.5 Hz), 127.0 (q, J=4.7 Hz), 131.1,
131.5, 132.3, 133.3 (q, J=33.0 Hz), 135.3, 137.1, 141.7, 174.8,
180.1. MS for C.sub.19H.sub.13F.sub.3N.sub.6OS, calculated 430.4,
found 430.1.
Example 3
3-1). 2-(4-hydroxyphenylamino)-2-methylpropanenitrile, 3a
[0101] A mixture of 4-aminophenol (1.09 g, 10 mmol), acetone
cyanohydrin (10 ml) and MgSO4 (2 g) was heated to 80.degree. C. and
stirred for 4 h. After concentration of the medium under vacuum,
compound 3a was crystallized from water (20 ml). The solid was
filtered and dried to yield
2-(4-hydroxyphenylamino)-2-methylpropanenitrile, 3a (1.69 g, 9.6
mmol, 96%).
3-2).
4-[3-(4-hydroxyphenyl)-5-imino-4,4-dimethyl-2-thioxoimidazolidin-1-y-
l]-2-trifluoromethylbenzonitrile, 3b
[0102] Triethylamine (0.101 g, 1 mmol) was added to a solution of
1a (0.456 g, 2 mmol) and 3a (0.352 g, 2 mmol) in dry THF (5 ml).
The reaction mixture was stirred at 0.degree. C. for 48 h and then
concentrated to yield a dark residue which was subjected to flash
chromatography (dichloromethane/acetone, 85:15) to afford
4-[3-(4-hydroxyphenyl)-5-imino-4,4-dimethyl-2-thioxoimidazolidin-1-yl]-2--
trifluoromethylbenzonitrile, 3b (0.274 g, 0.68 mmol, 34%).
3-3).
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-
-2-trifluoromethylbenzonitrile, 3c, [RD8]
[0103] A mixture of 3b (0.202 g, 0.5 mmol) in HCl aq., 2N (2 ml)
and methanol (5 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (10 ml) and extracted with ethyl acetate (10 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane/acetone, 9:1) to yield
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-tr-
ifluoromethylbenzonitrile, 3c, [RD8] (0.198 g, 0.49 mmol, 98%) as a
white powder.
##STR00023##
[0104] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57 (s, 6H), 6.26
(s, OH), 6.90-6.93 (m, 2H), 7.11-7.14 (m, 2H), 7.84 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.95-7.98 (m, 2H); .sup.13C
NMR (CDCl.sub.3, 100 MHz) .delta. 23.6, 66.5, 109.9, 114.9, 115.7,
116.8, 121.9 (q, J=272.7 Hz), 127.2 (q, J=4.7 Hz), 130.6, 132.3,
133.5 (q, J=33.2 Hz), 135.3, 137.2, 157.0, 175.3, 180.2.
Example 4
Chloroacetic acid
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxoimidazo-
lidin-1-yl]phenyl ester, 4a, [RD13]
[0105] Chloroacetyl chloride (0.045 g, 0.4 mmol) was added to a
mixture of 3c (0.101 g, 0.25 mmol) and triethylamine (0.041 g, 0.41
mmol) in dry THF (1.5 ml). The mixture was stirred at room
temperature for 4 h. Triethylamine hydrochloride was filtered off.
The filtrate was concentrated and chromatographed
(dichloromethane/acetone, 95:5) to yield 84% of Chloroacetic acid
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxoimidazo-
lidin-1-yl]phenyl ester, 4a, [RD13] (0.101 g, 0.21 mmol) as white
powder.
##STR00024##
[0106] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.58 (s, 6H), 4.32
(s, 2H), 7.33 (s, 4H), 7.83 (dd, J.sub.1=8.3 Hz, J.sub.2=1.9 Hz,
1H), 7.95-7.97 (m, 2H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta.
23.7, 40.8, 66.5, 110.1, 114.8, 121.9 (q, J=272.5 Hz), 122.7, 127.1
(q, J=4.7 Hz), 130.9, 132.3, 132.9, 133.5 (q, J=33.2 Hz), 135.3,
137.1, 150.9, 165.5, 174.8, 180.0.
Example 5
5-1a). 2-methyl-2-(4-methylphenyl)aminopropanenitrile, 5a
[0107] A mixture of p-toluidine (1.07 g, 10 mmol) and acetone
cyanohydrin (10 ml) was heated to 80.degree. C. and stirred for 4
h. The medium was concentrated and dried under vacuum to yield
2-methyl-2-(4-methylphenyl)aminopropanenitrile, 5a (1.72 g, 9.9
mmol, 99%) as brown solid.
5-1b). 2-methyl-2-(4-methylphenyl)aminopropanenitrile, 5a
[0108] Sodium cyanide (0.735 g, 15 mmol) was added to a mixture of
p-toluidine (1.07 g, 10 mmol) and acetone (1.16 g, 20 mmol) in 90%
acetic acid (10 ml). The reaction mixture was stirred at room
temperature for 12 h and then ethyl acetate (50 ml) was added. The
organic layer was washed with water (4.times.30 ml), dried over
magnesium sulfate and concentrated under vacuum to dryness to yield
2-methyl-2-(4-methylphenyl)aminopropanenitrile, 5a (1.65 g, 9.5
mmol, 95%) as a brown solid.
5-2).
4-[3-(4-methylphenyl)-5-imino-4,4-dimethyl-2-thioxoimidazolidin-1-yl-
]-2-trifluoromethylbenzonitrile, 5b
[0109] Triethylamine (0.101 g, 1 mmol) was added to a solution of
1a (0.456 g, 2 mmol) and 5a (0.348 g, 2 mmol) in dry THF (3 ml).
The reaction mixture was stirred at 0.degree. C. for 2 days and
then concentrated to yield a dark residue which was subjected to
flash chromatography (dichloromethane/acetone, 95:5) to afford
4-[3-(4-methylphenyl)-5-imino-4,4-dimethyl-2-thioxoimidazolidin-1-yl]-2-t-
rifluoromethylbenzonitrile, 5b (0.136 g, 0.34 mmol, 17%).
5-3a).
4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-
-2-trifluoromethylbenzonitrile, 5c
[0110] A mixture of 5b (0.121 g, 0.3 mmol) in HCl aq., 2N (2 ml)
and methanol (5 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (10 ml) and extracted with ethyl acetate (10 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield
4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-tri-
fluoromethylbenzonitrile, 5c (0.118 g, 0.294 mmol, 98%) as a white
powder.
5-3b).
4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-
-2-trifluoromethyl-benzonitrile, 5c, [RD7]
[0111] A mixture of 1a (0.547 g, 2.4 mmol) and 5a (0.348 g, 2 mmol)
in dry DMF (0.6 ml) was stirred for 36 h. To this mixture were
added methanol (20 ml) and 2N HCl (5 ml). The second mixture was
refluxed for 6 h. After being cooled to room temperature, the
reaction mixture was poured into cold water (30 ml) and extracted
with ethyl acetate (40 ml). The organic layer was dried over
MgSO.sub.4, concentrated and chromatographed (dichloromethane) to
yield
4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-tri-
fluoromethyl-benzonitrile, 5c, [RD7] (0.596 g, 1.48 mmol, 74%) as a
white powder.
##STR00025##
[0112] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.61 (s, 6H), 2.44
(s, 3H), 7.17-7.20 (m, 2H), 7.33-7.36 (m, 2H), 7.86 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.96-7.98 (m, 2H); .sup.13C
NMR (CDCl.sub.3, 100 MHz) .delta. 21.3, 23.6, 66.4, 110.0, 114.9,
121.9 (q, J=272.6 Hz), 127.1 (q, J=4.7 Hz), 129.2, 130.6, 132.2,
132.3, 133.4 (q, J=33.2 Hz), 135.2, 137.2, 140.1, 175.1, 179.9.
Example 6
6-1). 2-methyl-2-phenylaminopropanenitrile, 6a
[0113] A mixture of aminobenzene (0.931 g, 10 mmol) and acetone
cyanohydrin (2 ml) was heated to reflux and stirred for 20 h. After
being cold to room temperature, the reaction mixture was poured
into ethyl acetate (40 ml) and washed with cold water (2.times.30
ml). The organic layer was dried over MgSO.sub.4, concentrated
under vacuum to dryness to yield
2-methyl-2-phenylaminopropanenitrile, 6a (1.51 g, 9.4 mmol, 94%) as
slurry brown liquid.
6-2).
4-[3-phenyl-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trifluor-
omethylbenzonitrile, 6b, [RD10]
[0114] A mixture of 1a (0.274 g, 1.2 mmol) and 6a (0.160 g, 1 mmol)
in dry DMF (0.2 ml) was stirred for 48 h. To this mixture were
added methanol (10 ml) and 2N HCl (3 ml). The second mixture was
refluxed for 6 h. After being cooled to room temperature, the
reaction mixture was poured into cold water (20 ml) and extracted
with ethyl acetate (20 ml). The organic layer was dried over
MgSO.sub.4, concentrated and chromatographed (dichloromethane) to
yield
4-[3-phenyl-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trifluorometh-
ylbenzonitrile, 6b, [RD10] (0.276 g, 0.71 mmol, 71%) as a white
powder.
##STR00026##
[0115] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60 (s, 6H),
7.28-7.31 (m, 2H), 7.50-7.58 (m, 3H), 7.85 (dd, J.sub.1=8.3 Hz,
=1.8 Hz, 1H), 7.96-7.99 (m, 2H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 23.7, 66.4, 110.2, 114.8, 121.9 (q, J=272.6 Hz), 127.1 (q,
J=4.7 Hz), 129.5, 129.8, 129.9, 132.2, 133.4 (q, J=33.2 Hz), 135.1,
135.2, 137.2, 175.0, 179.9.
Example 7
7-1a). 1-(4-methylphenyl)aminocyclobutanenitrile, 7a
[0116] Sodium cyanide (0.147 g, 3 mmol) was added to a mixture of
p-toluidine (0.214 g, 2 mmol) and cyclobutanone (0.21 g, 3 mmol) in
90% acetic acid (3 ml). The reaction mixture was stirred at room
temperature for 12 h and then 20 ml of ethyl acetate was added. The
organic layer was washed with water (3.times.10 ml), dried over
magnesium sulfate and concentrated under vacuum to dryness to yield
1-(4-methylphenyl)aminocyclobutanenitrile, 7a (0.343 g, 1.84 mmol,
92%) as a brown solid.
7-1b). 1-(4-methylphenyl)aminocyclobutanenitrile, 7a
[0117] Trimethylsilyl cyanide (0.93 ml, 7 mmol) was added dropwise
to a mixture of p-toluidine (0.535 g, 5 mmol) and cyclobutanone
(0.42 g, 6 mmol). The reaction mixture was stirred at room
temperature for 6 h and then concentrated under vacuum to obtain a
brown liquid which was subjected to chromatography
(dichloromethane) to yield
1-(4-methylphenyl)aminocyclobutanenitrile, 7a (0.912 g, 4.9 mmol,
98%) as a yellowish solid.
7-2).
4-(8-imino-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)--
2-trifluoromethylbenzonitrile, 7b
[0118] To a solution of 1a (2.28 g, 10 mmol) in dry DMF (3 ml) was
added progressively, over 20 hours, a solution of 7a (1.764 g, 9
mmol) in dry DMF (3 ml) at room temperature. The medium was stirred
for an additional 4 h. After DMF being evaporated, the residue was
chromatographed (dichloromethane/acetone, 95:5) to afford
4-(8-imino-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-tri-
fluoromethylbenzonitrile, 7b (1.937 g, 4.68 mmol, 52%).
7-3a).
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-
-trifluoromethylbenzonitrile, 7c [RD37]
[0119] A mixture of 7b (0.041 g, 0.1 mmol) in HCl aq., 2N (3 ml)
and methanol (1 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (5 ml) and extracted with ethyl acetate (6 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trifl-
uoromethylbenzonitrile,
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trifl-
uoromethylbenzonitrile, 7c (0.04 g, 0.096 mmol, 96%) as a white
powder.
7-3b).
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-
-trifluoromethylbenzonitrile, 7c, [RD37]
[0120] A mixture of 1a (0.912 g, 4 mmol) and 7a (0.558 g, 3 mmol)
in dry DMF (0.5 ml) was stirred at room temperature for 24 h. To
this mixture were added methanol (30 ml) and HCl aq. 2N (6 ml). The
second mixture was refluxed for 6 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (50
ml) and extracted with ethyl acetate (60 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trifl-
uoromethylbenzonitrile, 7c (0.959 g, 2.31 mmol, 77%) as a white
powder.
##STR00027##
[0121] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.62-1.69 (m, 1H),
2.16-2.22 (m, 1H), 2.46 (s, 3H), 2.55-2.66 (m, 4H), 7.19-7.26 (m,
2H), 7.36-7.42 (m, 2H), 7.86 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz,
1H), 7.96 (d, J=8.3 Hz, 1H), 7.99 (d, J=1.8 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 13.7, 21.3, 31.4, 67.4, 109.9, 114.9,
121.9 (q, J=272.6 Hz), 127.1 (q, J=4.7 Hz), 129.5, 130.8, 132.2,
132.4, 133.3 (q, J=33.2 Hz), 135.2, 137.3, 140.1, 175.0, 180.0.
Example 8
8-1). 1-(4-methylphenyl)aminocyclopentanenitrile, 8a
[0122] Trimethylsilyl cyanide (0.865 ml, 7 mmol) was added dropwise
to a mixture of p-toluidine (0.535 g, 5 mmol) and cyclopentanone
(0.589 g, 7 mmol). The reaction mixture was stirred at room
temperature for 6 h and then concentrated under vacuum to obtain a
brown liquid which was subjected to chromatography
(dichloromethane) to yield
1-(4-methylphenyl)aminocyclopentanenitrile, 8a (0.981 g, 4.9 mmol,
98%) as a yellowish solid.
8-2).
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.4]non-3-yl)-2--
trifluoromethylbenzonitrile, 8b, [RD35]
[0123] A mixture of 1a (0.296 g, 1.3 mmol) and 8a (0.2 g, 1 mmol)
in dry DMF (0.2 ml) was stirred for 48 h. To this mixture were
added methanol (10 ml) and HCl aq. 2N (3 ml). The second mixture
was refluxed for 6 h. After being cooled to room temperature, the
reaction mixture was poured into cold water (20 ml) and extracted
with ethyl acetate (30 ml). The organic layer was dried over
MgSO.sub.4, concentrated and chromatographed (dichloromethane) to
yield
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.4]non-3-yl)-2-trifl-
uoromethylbenzonitrile, 8b, [RD35] (0.3 g, 0.7 mmol, 70%) as a
white powder.
##STR00028##
[0124] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.47-1.57 (m, 2H),
1.81-1.92 (m, 2H), 2.20-2.24 (m, 2H), 2.27-2.34 (m, 2H), 2.43 (s,
3H), 7.18-7.22 (m, 2H), 7.33-7.36 (m, 2H), 7.86 (dd, J.sub.1=8.2
Hz, J.sub.2=1.8 Hz, 1H), 7.96 (d, J=8.2 Hz, 1H), 7.98 (d, J=1.8 Hz,
1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 21.3, 25.2, 36.3,
75.1, 110.0, 114.9, 121.9 (q, J=272.5 Hz), 127.1 (q, J=4.7 Hz),
129.5, 130.7, 123.2, 133.0, 133.4 (q, J=33.2 Hz), 135.1, 137.4,
140.0, 176.3, 180.2.
Example 9
9-1). 1-(4-methylphenyl)aminocyclohexanenitrile, 9a
[0125] Sodium cyanide (0.147 g, 3 mmol) was added to a mixture of
p-toluidine (0.214 g, 2 mmol) and cyclohexanone (0.294 g, 3 mmol)
in acetic acid 90% (3 ml). The reaction mixture was stirred at room
temperature for 12 h and then 20 ml of ethyl acetate was added. The
organic layer was washed with water (3.times.10 ml), dried over
magnesium sulfate and concentrated under vacuum to dryness to yield
1-(4-methylphenyl)aminocyclohexanenitrile, 9a (0.398 g, 1.86 mmol,
93%) as a brown solid.
9-2).
4-(4-imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-3-yl)--
2-trifluoromethylbenzonitrile, 9b
[0126] Triethylamine (0.05 g, 0.5 mmol) was added to a solution of
1a (0.228 g, 1 mmol) and 9a (0.214 g, 1 mmol) in dry THF (2 ml).
The reaction mixture was stirred at room temperature for 2 days and
then concentrated to yield a dark residue which was subjected to
flash chromatography (dichloromethane/acetone, 95:5) to afford
4-(4-imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-3-yl)-2-tri-
fluoromethylbenzonitrile, 9b (0.035 g, 0.08 mmol, 8%).
9-3).
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-3-yl)-2--
trifluoromethylbenzonitrile, 9c, [RD48]
[0127] A mixture of 9b (0.035 g, 0.08 mmol) in HCl aq., 2N (1 ml)
and methanol (3 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (5 ml) and extracted with ethyl acetate (6 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-3-yl)-2-trifl-
uoromethylbenzonitrile, 9c, [RD48] (0.034 g, 0.076 mmol, 95%) as a
white powder.
##STR00029##
[0128] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.02-1.05 (m, 1H),
1.64-1.76 (m, 4H), 2.03-2.12 (m, 5H), 2.44 (s, 3H), 7.12-7.15 (m,
2H), 7.33-7.36 (m, 2H), 7.85 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz,
1H), 7.96 (d, J=8.3 Hz, 1H), 7.97 (d, J=1.8 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 20.7, 21.3, 24.0, 32.6, 67.4, 109.9,
114.9, 122.0 (q, J=272.5 Hz), 127.3 (q, J=4.6 Hz), 130.0, 130.5,
132.0, 132.5, 133.3 (q, J=33.2 Hz), 135.2, 137.3, 140.1, 174.1,
180.1.
Example 10
10-1). 1-(4-methylphenyl)aminocyclohexanenitrile, 10a
[0129] Sodium cyanide (0.147 g, 3 mmol) was added to a mixture of
p-toluidine (0.214 g, 2 mmol) and cycloheptanone (0.337 g, 3 mmol)
in acetic acid 90% (3 ml). The reaction mixture was stirred at room
temperature for 12 h and then 20 ml of ethyl acetate was added. The
organic layer was washed with water (3.times.10 ml), dried over
magnesium sulfate and concentrated under vacuum to dryness to yield
1-(4-methylphenyl)aminocyclohexanenitrile, 10a (0.438 g, 1.92 mmol,
96%) as a brown solid.
10-2).
4-(4-imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]undec-3-y-
l)-2-trifluoromethylbenzonitrile, 10b
[0130] Triethylamine (0.05 g, 0.5 mmol) was added to a solution of
1a (0.228 g, 1 mmol) and 9a (0.228 g, 1 mmol) in dry THF (2 ml).
The reaction mixture was stirred at room temperature for 2 days and
then concentrated to yield a dark residue which was subjected to
flash chromatography (dichloromethane/acetone, 95:5) to afford
4-(4-imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]undec-3-yl)-2-t-
rifluoromethylbenzonitrile, 10b (0.036 g, 0.08 mmol, 8%).
10-3).
4-(4-oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]undec-3-yl)-
-2-trifluoromethylbenzonitrile, 10c, [RD49]
[0131] A mixture of 9b (0.036 g, 0.08 mmol) in HCl aq., 2N (1 ml)
and methanol (3 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (5 ml) and extracted with ethyl acetate (6 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 10c (0.034 g, 0.075
mmol, 94%) as a white powder.
##STR00030##
[0132] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.24-134 (m, 2H),
1.37-1.43 (m, 2H), 1.53-1.60 (m, 2H), 1.74-1.82 (m, 2H), 2.19-2.25
(m, 4H), 2.44 (s, 3H), 7.16-7.19 (m, 2H), 7.32-7.35 (m, 2H), 7.83
(dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.95-7.97 (m, 2H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 21.4, 22.2, 30.9, 36.3,
71.1, 110.0, 114.9, 121.9 (q, J=272.5 Hz), 127.2 (q, J=4.6 Hz),
129.6, 130.5, 132.3, 133.0, 133.2 (q, J=33.2 Hz), 135.1, 137.4,
140.0, 175.9, 179.7.
Example 11
11-1). 1-(4-hydroxyphenyl)aminocyclobutanenitrile, 11a
[0133] Trimethylsilyl cyanide (0.93 ml, 7 mmol) was added dropwise
to a mixture of 4-hydroxyaniline (0.545 g, 5 mmol) and
cyclobutanone (0.42 g, 6 mmol). The reaction mixture was stirred at
room temperature for 6 h and then concentrated under vacuum to
obtain a brown liquid which was subjected to chromatography
(dichloromethane:acetone, 98:2) to yield 11a (0.903 g, 4.8 mmol,
96%) as a yellowish solid.
11-2).
4-(8-oxo-6-thioxo-5-(4-hydroxyphenyl)-5,7-diazaspiro[3.4]oct-7-yl)--
2-trifluoromethylbenzonitrile, 11b, [RD58]
[0134] A mixture of 1a (0.57 g, 2.5 mmol) and 7a (0.376 g, 2 mmol)
in dry DMF (0.5 ml) was stirred at room temperature for 40 h. To
this mixture were added methanol (30 ml) and HCl aq. (5 ml). The
second mixture was refluxed for 6 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (40
ml) and extracted with ethyl acetate (50 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 98:2) to yield 11b (0.659 g, 1.58 mmol,
79%) as a white powder.
##STR00031##
[0135] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.55-1.63 (m, 1H),
2.01-2.09 (m, 1H), 2.50-2.65 (m, 4H), 6.97-7.01 (m, 2H), 7.20-7.24
(m, 2H), 8.02 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 8.14 (d,
J=1.8 Hz, 1H), 8.21 (d, J=8.3 Hz, 1H); .sup.13C NMR
(Acetone-d.sub.6, 100 MHz) .delta. 13.4, 31.3, 67.5, 108.9, 114.8,
116.1, 123.5 (q, J=271.5 Hz), 127.4 (q, J=4.9 Hz), 131.3, 131.8 (q,
J=32.7 Hz), 133.3, 135.5, 136.2, 138.5, 158.1, 175.1, 180.7.
Example 12
12-1). 1-(4-biphenylamino)cyclobutanecarbonitrile, 12a
[0136] Trimethylsilyl cyanide (0.2 ml, 1.5 mmol) was added dropwise
to a mixture of 4-biphenylamine (0.169 g, 1 mmol) and cyclobutanone
(0.098 g, 1.4 mmol). The reaction mixture was stirred at room
temperature for 6 h and then concentrated under vacuum to obtain a
brown liquid which was subjected to chromatography
(dichloromethane) to yield 12a (0.24 g, 0.97 mmol, 97%) as a white
solid.
12-2).
4-(8-oxo-6-thioxo-5-(4-biphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-tri-
fluoromethylbenzonitrile, 12b [RD57]
[0137] A mixture of 1a (0.137 g, 0.6 mmol) and 12a (0.124 g, 0.5
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 3
days. To this mixture were added methanol (5 ml) and HCl aq. 2N (1
ml). The second mixture was refluxed for 6 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(10 ml) and extracted with ethyl acetate (15 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 12b (0.162 g, 0.34 mmol, 68%) as a white
powder.
##STR00032##
[0138] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.67-1.76 (m, 1H),
2.19-2.31 (m, 1H), 2.59-2.74 (m, 4H), 7.40-7.44 (m, 3H), 7.47-7.53
(m, 2H), 7.64-7.67 (m, 2H), 7.79-7.82 (m, 2H), 7.88 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.97 (d, J=8.2 Hz, 1H), 8.02
(d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
31.5, 67.5, 110.0, 114.9, 122.0 (q, J=272.6 Hz), 127.1 (q, J=4.7
Hz), 127.3, 128.1, 128.7, 129.0, 130.2, 132.3, 133.5 (q, J=33.2
Hz), 134.2, 135.2, 137.2, 139.6, 142.8, 174.9, 179.9.
Example 13
13-1). 1-(2-naphthylamino)cyclobutanecarbonitrile, 13a
[0139] Trimethylsilyl cyanide (0.27 ml, 2 mmol) was added dropwise
to a mixture of 2-aminonaphthalene (0.143 g, 1 mmol) and
cyclobutanone (0.098 g, 1.4 mmol). The reaction mixture was stirred
at room temperature for 12 h and then concentrated under vacuum to
obtain a brown liquid which was subjected to chromatography
(dichloromethane) to yield 13a (0.209 g, 0.94 mmol, 94%) as a
yellow solid.
13-2).
4-(8-oxo-6-thioxo-5-(4-biphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-tri-
fluoromethylbenzonitrile, 12b, [RD85]
[0140] A mixture of 1a (0.137 g, 0.6 mmol) and 13a (0.111 g, 0.5
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 3
days. To this mixture were added methanol (5 ml) and HCl aq. (1
ml). The second mixture was refluxed for 6 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(10 ml) and extracted with ethyl acetate (15 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 12b (0.146 g, 0.325 mmol, 65%) as a
white powder.
##STR00033##
[0141] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 158-1.68 (m, 1H),
2.17-2.29 (m, 1H), 2.61-2.75 (m, 4H), 7.40 (dd, J.sub.1=8.6 Hz,
J.sub.2=2.0 Hz, 1H), 7.58-7.65 (m, 2H), 7.86-8.00 (m, 5H), 8.04
(J=1.8 Hz, 1H), 8.06 (d, J=8.6 Hz, 1H); .sup.13C NMR (CDCl.sub.3,
100 MHz) .delta. 13.7, 31.6, 67.7, 110.0, 114.9, 122.0 (q, J=272.6
Hz), 126.8, 127.1 (q, J=4.8 Hz), 127.2, 127.7, 128.0, 128.3, 129.1,
130.2, 132.2, 132.5, 133.4, 133.5 (q, J=33.1 Hz), 133.6, 135.2,
137.2, 175.0, 180.1.
Example 14
14-1). 2-(4-methyl-2-pyridinamino)-2-methylpropanenitrile, 14a
[0142] Trimethylsilyl cyanide (0.27 ml, 2 mmol) was added dropwise
to a mixture of 2-amino-4-methylpyridine (0.108 g, 1 mmol) and
acetone (0.58 g, 10 mmol). The reaction mixture was stirred at room
temperature for 6 days and then concentrated under vacuum to obtain
a brown liquid which was subjected to chromatography
(dichloromethane:acetone, 60:40) to yield 14a (0.133 g, 0.76 mmol,
76%) as a white solid.
14-2).
4-[4,4-dimethyl-3-(4-methylpyridin-2-yl)-5-oxo-2-thioxoimidazolidin-
-1-yl]-2-trifluoromethylbenzonitrile, 14b, [RD83]
[0143] A mixture of 1a (0.91 g, 0.4 mmol) and 14a (0.053 g, 0.3
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 6
days. To this mixture were added methanol (5 ml) and HCl aq. (1
ml). The second mixture was refluxed for 5 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(10 ml) and extracted with ethyl acetate (15 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 14b (0.07 g, 0.174 mmol, 58%) as a white
powder.
##STR00034##
[0144] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.70 (s, 6H), 2.44
(s, 3H), 7.19 (d, J=4.4 Hz, 1H), 7.45 (t, J=0.6 Hz, 1H), 7.82 (dd,
J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (d, J=1.8 Hz, 1H), 7.97
(d, J=8.2 Hz, 1H), 8.47 (d, J=5.0 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 21.1, 24.1, 67.1, 110.2, 114.8, 121.9
(q, J=272.6 Hz), 124.4, 125.1, 127.3 (q, J=4.8 Hz), 132.4, 133.5
(q, J=33.2 Hz), 135.3, 137.1, 149.2, 149.5, 150.0, 175.2,
179.0.
Example 15
15-1). 2-(2-pyridinamino)-2-methylpropanenitrile, 15a
[0145] Trimethylsilyl cyanide (0.27 ml, 2 mmol) was added dropwise
to a mixture of 2-aminopyridine (0.094 g, 1 mmol) and acetone (0.58
g, 10 mmol). The reaction mixture was stirred at room temperature
for 6 days and then concentrated under vacuum to obtain a brown
liquid which was subjected to chromatography
(dichloromethane:acetone, 60:40) to yield 15a (0.131 g, 0.81 mmol,
81%) as a white solid.
15-2).
4-[4,4-dimethyl-3-(4-pyridin-2-yl)-5-oxo-2-thioxoimidazolidin-1-yl]-
-2-trifluoromethylbenzonitrile, 15b, [RD82]
[0146] A mixture of 1a (0.91 g, 0.4 mmol) and 15a (0.048 g, 0.3
mmol) in dry DMF (0.3 ml) was stirred at room temperature for 10
days. To this mixture were added methanol (5 ml) and of HCl aq. (1
ml). The second mixture was refluxed for 5 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(10 ml) and extracted with ethyl acetate (15 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 15b (0.059 g, 0.153 mmol, 51%) as a
white powder.
##STR00035##
[0147] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.73 (s, 6H), 7.38
(dd, J.sub.1=7.3 Hz, J.sub.2=5.4 Hz, 1H), 7.71 (d, J=8.0 Hz, 1H),
7.87 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (td,
J.sub.1=7.8 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (d, J=1.3 Hz, 1H), 7.98
(d, J=8.2 Hz, 1H), 8.62 (dd, J.sub.1=4.7 Hz, J.sub.2=1.3 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 24.2, 67.1, 110.3,
114.8, 121.9 (q, J=272.6 Hz), 123.7, 123.8, 127.3 (q, J=4.8 Hz),
132.4, 133.6 (q, J=33.2 Hz), 135.3, 137.1, 138.2, 149.5, 149.6,
175.1, 179.0.
Example 16
16-1). 1-(5-methyl-2H-pyrazol-3-ylamino)-cyclobutanecarbonitrile,
16a
[0148] Trimethylsilyl cyanide (0.532 ml, 4.0 mmol) was added
dropwise to the mixture of 3-amino-5-methylpyrazole (0.194 g, 2.0
mmol) and cyclobutanone (0.154 g, 2.2 mmol). The reaction mixture
was stirred at room temperature for 40 h and then concentrated
under vacuum to obtain a dark liquid which was subjected to
chromatography (dichloromethane) to yield 16a (0.267 g, 1.52 mmol,
76%) as a off-white powder.
16-2).
4-[5-(5-methyl-2H-pyrazol-3-yl)-8-oxo-6-thioxo-5,7-diaza-spiro[3.4]-
oct-7-yl]-2-trifluoromethyl-benzonitrile, 16b, [RD84]
[0149] A mixture of 1a (0.0684 g, 0.3 mmol) and 16a (0.053 g, 0.3
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 4
days. To this mixture were added methanol (10 ml) and HCl aq. 2N (2
ml). The second mixture was refluxed for 5 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(30 ml) and extracted with ethyl acetate (30 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 97:3) to yield 16b (0.0826 g, 0.2 mmol,
67%) as a white powder.
##STR00036##
[0150] .sup.1H NMR (acetone d.sub.6, 400 MHz) .delta. .delta.
1.66-1.76 (m, 1H), 2.00-2.07 (m, 1H), 3.35 (s, 3H), 2.56-2.63 (m,
2H), 2.85-2.93 (m, 2H), 8.04 (dd, J.sub.1=8.2 Hz, J.sub.2=1.6 Hz,
1H), 8.18 (d, J=1.6 Hz, 1H), 8.22 (d, J=8.2 Hz, 1H), 11.99 (s, 1H);
.sup.13C NMR (acetone d.sub.6, 100 MHz) .delta. 10.2, 13.1, 31.1,
67.4, 102.5, 109.1, 114.8, 122.5 (q, J=271.4 Hz), 127.8 (q, J=4.8
Hz), 131.9 (q, J=33.6 Hz), 133.6, 135.6, 138.4, 139.9, 145.0,
175.0, 179.6.
Example 17
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-2,5-dithioxoimidazolidin-1-yl]-2-trifl-
uoromethylbenzonitrile, 17a, [RD59]
[0151] A mixture of 3c (0.081 g, 0.2 mmol) and Lawesson reagent
(0.097 g, 0.24 mmol) in toluene (3 ml) was heated to reflux for 15
h. After being cooled to room temperature, the reaction mixture was
poured into cold water (10 ml) and extracted with ethyl acetate (10
ml). The organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:pentane, 9:1) to yield 17a (0.0185
g, 0.044 mmol, 22%) as a white powder.
##STR00037##
[0152] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.65 (s, 6H),
6.95-6.97 (m, 2H), 7.15-7.18 (m, 2H), 7.75 (d, J=8.2 Hz, 1H), 7.86
(d, J=1.8 Hz, 1H), 7.98 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 27.9, 77.8, 110.9,
114.7, 116.7, 121.9 (q, J=272.6 Hz), 128.1 (q, J=4.8 Hz), 129.1,
130.7, 133.3, 133.5 (q, J=33.2 Hz), 135.5, 140.3, 156.8, 179.9,
207.9.
Example 18
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-2,5-dioxoimidazolidin-1-yl]-2-trifluor-
omethylbenzonitrile, 18a, [RD60]
[0153] Hydrogen peroxide, 30% (3 ml, 26 mmol) was added dropwise to
a solution of 3c (0.121 g, 0.4 mmol) in glacial acetic acid (3 ml).
The mixture was stirred at room temperature for 12 h and then 20 ml
of ethyl acetate was added. The organic layer was washed with water
(3.times.15 ml), dried over magnesium sulfate, concentrated and
chromatographed (dichloromethane) to yield 18a (0.102 g, 0.261
mmol, 87%) as a white powder.
##STR00038##
[0154] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.52 (s, 6H),
6.70-6.73 (m, 2H), 7.01-7.04 (m, 2H), 7.92 (d, J=8.4 Hz, 1H), 8.00
(dd, J.sub.1=8.4 Hz, J.sub.2=1.8 Hz, 1H), 8.15 (d, J=1.8 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 23.7, 63.7, 108.4,
115.0, 116.7, 121.9 (q, J=272.6 Hz), 123.5 (q, J=4.8 Hz), 124.0,
128.5, 130.5, 133.6 (q, J=33.2 Hz), 135.5, 136.2, 153.4, 157.2,
174.5.
Example 19
19-1). 3-fluoro-2-methyl-2-(4-methylphenyl)aminopropionitrile,
19a
[0155] Trimethylsilyl cyanide (0.146 ml, 1.1 mmol) was added
dropwise to the mixture of p-toluidine (0.107 g, 1 mmol) and
fluoroacetone (0.082 g, 1.1 mmol). The reaction mixture was stirred
at room temperature for 12 h and then concentrated under vacuum to
obtain a brown liquid which was subjected to chromatography
(dichloromethane) to yield 19a (0.179 g, 0.93 mmol, 93%) as a
yellowish solid.
19-2).
4-(4-fluoromethyl-4-methyl-5-oxo-2-thioxo-3-(4-methylphenyl)imidazo-
lidin-1-yl)-2-trifluoromethylbenzonitrile, 19b, [RD68]
[0156] A mixture of 1a (0.16 g, 0.7 mmol) and 19a (0.096 g, 0.5
mmol) in dry DMF (0.3 ml) was stirred at room temperature for 48 h.
To this mixture were added methanol (10 ml) and HCl aq. 2N (2 ml).
The second mixture was refluxed for 6 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (30
ml) and extracted with ethyl acetate (30 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 19b (0.168 g, 0.4 mmol, 80%) as a white
powder.
##STR00039##
[0157] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.49 (s, 3H), 2.44
(s, 3H), 4.35 (dd, J.sub.1=47.2 Hz, J.sub.2=10.0 Hz, 1H), 4.71 (dd,
J.sub.1=45.2 Hz, J.sub.2=10 Hz, 1H), 7.22-7.26 (m, 2H), 7.35-7.39
(m, 2H), 7.82 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.93 (d,
J=1.8 Hz, 1H), 7.98 (d, J=8.2 Hz, 1H); .sup.13C NMR (CDCl.sub.3,
100 MHz) .delta. 17.0 (d, J=4.6 Hz), 21.3, 69.3 (d, J=18.3 Hz),
81.9 (d, J=179.5 Hz), 109.9, 114.8, 121.8 (q, J=272.6 Hz), 127.2
(q, J=4.7 Hz), 129.3, 130.9, 131.6, 132.3, 133.3 (q, J=33.2 Hz),
135.3, 137.0, 140.5, 174.1, 181.4; .sup.19F NMR (CDCl.sub.3, 376
MHz) .delta. -62.5, 110.9.
Example 20
20-1). 2-methyl-2-(4-trifluoromethylphenyl)aminopropanenitrile,
20a
[0158] A mixture of 4-trifluoromethylaniline (1.61 g, 10 mmol),
acetone cyanohydrin (5 ml) and magnesium sulfate (2 g) was heated
to 80.degree. C. and stirred for 12 h. To the medium was added
ethyl acetate (50 ml) and then washed with water (3.times.30 ml).
The organic layer was dried over MgSO.sub.4 and concentrated under
vacuum to dryness to yield 20a (2.166 g, 9.5 mmol, 95%) as brown
solid.
20-2).
4-(4,4-dimethyl-5-oxo-2-thioxo-3-(4-trifluoromethylphenyl)imidazoli-
din-1-yl)-2-trifluoromethylbenzonitrile, 20b, [RD66]
[0159] A mixture of 1a (0.114 g, 0.5 mmol) and 20a (0.092 g, 0.4
mmol) in dry DMF (0.3 ml) was stirred at room temperature for 48 h.
To this mixture were added methanol (10 ml) and HCl aq. (3 ml). The
second mixture was refluxed for 6 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (20
ml) and extracted with ethyl acetate (20 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 20b (0.117 g, 0.256 mmol, 64%) as a
white powder.
##STR00040##
[0160] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.61 (s, 6H),
7.45-7.49 (m, 2H), 7.80-7.83 (m, 2H), 7.85 (dd, J.sub.1=8.3 Hz,
=1.8 Hz, 1H), 7.97 (d, J=1.8 Hz, 1H), 7.99 (d, J=8.2 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 23.8, 66.6, 110.3,
114.8, 121.8 (q, J=272.6 Hz), 123.5 (q, J=271.1 Hz), 127.0 (q,
J=4.6 Hz), 127.1 (q, J=4.7 Hz), 130.3, 131.9 (q, J=32.9 Hz), 132.2,
133.5 (q, J=33.3 Hz), 135.3, 136.9, 138.4, 174.6, 179.9.
Example 21
21-1).
3-chloro-2-chloromethyl-2-(4-methylphenyl)aminopropanenitrile,
21a
[0161] Trimethylsilyl cyanide (0.27 ml, 2 mmol) was added dropwise
to a mixture of p-toluidine (0.107 g, 1 mmol) and
1,3-dichloroacetone (0.254 g, 2 mmol). The reaction mixture was
heat to 80.degree. C. and stirred for 6 h. To the mixture was added
20 ml of ethyl acetate and then wash with water (2.times.20 ml).
The organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 21a (0.192 g, 0.79 mmol,
79%) as a brown powder.
21-2).
4-(4,4-bischloromethyl-5-oxo-2-thioxo-3-(4-methylphenyl)imidazolidi-
n-1-yl)-2-trifluoromethylbenzonitrile, 21b, [RD67]
[0162] A mixture of 1a (0.16 g, 0.7 mmol) and 21a (0.122 g, 0.5
mmol) in dry DMF (0.5 ml) was stirred at room temperature for 10
days. To this mixture were added methanol (10 ml) and of HCl aq. 2N
(2 ml). The second mixture was refluxed for 6 h. After being cooled
to room temperature, the reaction mixture was poured into cold
water (20 ml) and extracted with ethyl acetate (30 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 21b (0.09 g, 0.19 mmol, 38%) as a white
powder.
##STR00041##
[0163] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 2.44 (s, 3H), 3.54
(d, J=11.8 Hz, 2H), 3.93 (d, J=11.8 Hz, 2H), 7.37-7.40 (m, 2H),
7.48-7.51 (m, 2H), 7.79 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H),
7.88 (d, J=1.8 Hz, 1H), 7.98 (d, J=8.2 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 21.4, 42.8, 74.3, 110.7, 114.7, 121.7
(q, J=272.6 Hz), 127.2 (q, J=4.7 Hz), 128.8, 131.0, 131.1, 132.4,
133.8 (q, J=33.2 Hz), 135.5, 136.9, 140.9, 169.5, 182.5.
Example 22
22-1). 1-(4-methylphenyl)aminocyclohexanenitrile, 22a
[0164] Sodium cyanide (0.245 g, 5 mmol) was added to a mixture of
anthranilic acid (0.411 g, 3 mmol) and acetone (1 ml, 13.6 mmol) in
acetic acid 90% (3 ml). The reaction mixture was stirred at room
temperature for 12 h and then 50 ml of ethyl acetate was added. The
organic layer was washed with brine (3.times.30 ml). The organic
layer was dried over magnesium sulfate, concentrated and
chromatographed (dichloromethane:acetone, 90:10) to yield 22a
(0.551 g, 2.7 mmol, 90%) as a brown solid.
22-2).
2-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxoi-
midazolidin-1-yl]benzoic acid, 22b, [RD65]
[0165] A mixture of 1a (0.114 g, 0. mmol) and 22a (0.103 g, 0.5
mmol) in dry DMF (0.5 ml) was stirred at room temperature for 3
days. To this mixture were added methanol (10 ml) and HCl aq. 2N,
(3 ml). The second mixture was refluxed for 6 h. After being cooled
to room temperature, the reaction mixture was poured into cold
water (20 ml) and extracted with ethyl acetate (30 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(ethyl acetate:pentane, 2:1) to yield 22b (0.143 g, 0.33 mmol, 66%)
as a white powder.
##STR00042##
[0166] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.47 (s, 3H), 1.78
(s, 3H), 7.39 (d, J=7.7 Hz, 1H), 7.63 (t, J=7.7 Hz, 1H) 7.76-7.82
(m, 2H), 7.90-7.98 (m, 2H), 8.22 (d, J=6.8 Hz, 1H), 8.96 (bs, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 20.6, 26.2, 67.6, 110.1,
114.8, 121.9 (q, J=272.6 Hz), 127.2 (q, J=4.7 Hz), 128.9, 131.0,
130.2, 132.5, 133.2 (q, J=33.3 Hz), 133.7, 134.7, 135.4, 135.8,
137.3, 169.8, 175.3, 180.7.
Example 23
23-1). 1-(2-methylphenyl)aminocyclobutanenitrile, 23a
[0167] Trimethylsilyl cyanide (0.66 ml, 5 mmol) was added dropwise
to the mixture of p-toluidine (0.321 g, 3 mmol) and cyclobutanone
(0.28 g, 4 mmol). The reaction mixture was stirred at room
temperature for 6 h and then concentrated under vacuum to obtain a
brown liquid which was subjected to chromatography
(dichloromethane) to yield 23a (0.541 g, 2.91 mmol, 97%) as a
yellowish solid.
23-2).
4-(8-oxo-6-thioxo-5-(2-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-
-trifluoromethylbenzonitrile, 23b, [RD71]
[0168] A mixture of 1a (0.114 g, 0.5 mmol) and 23a (0.093 g, 0.5
mmol) in dry DMF (0.3 ml) was stirred at room temperature for 3
days. To this mixture were added methanol (10 ml) and HCl aq. 2N,
(3 ml). The second mixture was refluxed for 6 h. After being cooled
to room temperature, the reaction mixture was poured into cold
water (20 ml) and extracted with ethyl acetate (30 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 23b (0.116 g, 0.28 mmol, 56%) as a white
powder.
##STR00043##
[0169] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.69 (m, 1H),
2.26 (s, 3H), 2.28-2.41 (m, 2H), 2.58-2.76 (m, 3H), 7.21 (d, J=7.6
Hz, 1H), 7.39-7.49 (m, 3H), 7.89 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8
Hz, 1H), 7.97 (d, J=8.2 Hz, 1H), 8.00 (d, J=1.8 Hz, 1H); .sup.13C
NMR (CDCl.sub.3, 100 MHz) .delta. 14.2, 18.0, 30.7, 32.2, 67.6,
109.9, 114.9, 121.9 (q, J=272.6 Hz), 127.0 (q, J=4.7 Hz), 127.5,
129.8, 130.2, 131.9, 132.3, 133.4, 133.5 (q, J=34.3 Hz), 135.2,
135.8, 137.1, 138.0, 175.3, 178.7.
Example 24
24-1). 1-aminocyclopentanecarbonitrile, 24a
[0170] Ammonia anhydrous was bubble into a mixture of
cyclopentanone (0.452 g) and trimethylsilyl cyanide (0.66 ml, 5
mmol). The excess of ammonia was refluxed by a dry ice-acetone
condenser. After 1 h of reflux, the ammonia was allowed to degas
form the medium and then the remaining mixture was concentrated
under vacuum to yield 24a (0.522 g, 4.75 mmol, 95%) as a colorless
liquid.
24-2).
4-(4-imino-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2-trifluoromethylb-
enzonitrile, 24b
[0171] Triethylamine (0.101 g, 0.1 mmol) was added to a solution of
1a (0.684 g, 3 mmol) and 24a (0.33 g, 3 mmol) in dry THF (5 ml).
The reaction mixture was stirred at room temperature for 5 h and
then concentrated to yield a brown residue which was subjected to
flash chromatography (dichloromethane/acetone, 93:7) to afford 24b
(0.741 g, 2.19 mmol, 73%).
24-3).
4-(4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2-trifluoromethylben-
zonitrile, 24c, [RD77]
[0172] A mixture of 24b (0.741 g, 2.19 mmol) in HCl aq., 2N (4 ml)
and methanol (20 ml) was heated to reflux for 1 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (20 ml) and extracted with ethyl acetate (40 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 24c (0.72 g, 2.12 mmol,
97%) as a white powder.
##STR00044##
[0173] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.86-190 (m, 2H),
1.96-2.05 (m, 4H), 2.26-2.30 (m, 2H), 7.80 (dd, J.sub.1=8.2 Hz,
J.sub.2=1.8 Hz, 1H), 7.92 (d, J=1.8 Hz, 1H), 7.97 (d, J=8.2 Hz, 1H)
8.20 (bs, NH); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 25.3,
38.1, 71.0, 110.1, 114.8, 121.8 (q, J=272.7 Hz), 126.8 (q, J=4.7
Hz), 131.9, 133.6 (q, J=34.3 Hz), 135.3, 136.7, 176.1, 179.8.
Example 25
25).
4-[1-(4-nitrophenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl]-2-tr-
ifluoromethylbenzonitrile, 25a, [RD55]
[0174] A mixture of 25c (0.0678 g, 0.2 mmol),
1,8-Diazabicyclo[5.4.0]undec-7-ene (0.05 g, 0.33 mmol) and
4-fluoronitrobenzene (0.056 g, 0.4 mmol) in dimethylformamide (0.5
ml) was placed under argon in a sealed-tube and heated to
130.degree. C. for 40 h. The reaction mixture was poured into ethyl
acetate (5 ml) and washed with water (2.times.10 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 25a (0.038 g, 0.084 mmol, 42%) as a
white powder.
##STR00045##
[0175] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.53-1.56 (m, 2H),
1.90-1.93 (m, 2H), 2.14-2.18 (m, 2H), 2.37-2.40 (m, 2H), 7.54-7.57
(m, 2H), 7.85 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.97 (d,
J=1.8 Hz, 1H), 7.98 (d, J=8.2 Hz, 1H), 8.39-8.43 (m, 2H); .sup.13C
NMR (CDCl.sub.3, 100 MHz) .delta. 25.2, 36.5, 75.3, 110.3, 114.8,
121.8 (q, J=272.6 Hz), 125.2, 127.0 (q, J=4.7 Hz), 131.4, 132.1,
133.6 (q, J=34.3 Hz), 135.3, 136.9, 141.7, 148.1, 175.6, 180.2.
Example 26
26).
4-[1-(4-cyanophenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl]-2-tr-
ifluoromethylbenzonitrile, 26a, [RD54]
[0176] A mixture of 24c (0.0678 g, 0.2 mmol),
1,8-diazabicyclo[5.4.0]undec-7-ene (0.061 g, 0.4 mmol) and
4-fluorocyanobenzene (0.048 g, 0.4 mmol) in dimethylformamide (0.5
ml) was placed under argon in a sealed-tube and heated to
140.degree. C. for 5 days. The reaction mixture was poured into
ethyl acetate (5 ml) and washed with water (2.times.10 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 26a (0.023 g, 0.052
mmol, 26%) as a white powder.
##STR00046##
[0177] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.51-1.55 (m, 2H),
1.90-1.93 (m, 2H), 2.12-2.16 (m, 2H), 2.33-2.38 (m, 2H), 7.47-7.50
(m, 2H), 7.81-7.87 (m, 3H), 7.95-7.99 (m, 2H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 25.2, 36.5, 75.3, 110.3, 113.9,
114.7, 117.5, 121.8 (q, J=272.6 Hz), 127.0 (q, J=4.8 Hz), 131.2,
132.1, 133.6 (q, J=34.3 Hz), 133.8, 135.3, 136.9, 140.0, 175.6,
180.1.
Example 27
27-1). 1-methyl-4-(4-methylphenylamino)piperidine-4-carbonitrile,
27a
[0178] Sodium cyanide (0.318 g, 6.5 mmol) was added to a mixture of
p-toluidine (0.535 g, 5 mmol) and 1-methyl-4-piperidinone (0.678 g,
6 mmol) in acetic acid 90% (5 ml). The reaction mixture was stirred
at room temperature for 6 h and then 100 ml of dichloromethane was
added. The organic layer was washed with a solution NaOH, 2N
(2.times.50 ml), dried over magnesium sulfate, concentrated and
chromatographed (DCM and then acetone) to obtained 27a (0.722 g,
3.15 mmol, 63%).
27.2).
4-(4-imino-8-methyl-2-thioxo-1-(4-methylphenyl)-1,3,8-triazaspiro[4-
.5]dec-3-yl)-2-trilluoromethylbenzonitrile, 27b
[0179] Triethylamine (0.02, 0.2 mmol) was added to a solution of 1a
(0.228 g, 1 mmol) and 27a (0.114 g, 0.5 mmol) in dry THF (2 ml).
The reaction mixture was stirred at room temperature for 20 h and
then concentrated to yield a dark residue which was subjected to
flash chromatography (dichloromethane/acetone, 90:10, and then
acetone) to afford 27b (0.059 g, 0.13 mmol, 26%).
27-3).
4-(8-methyl-4-oxo-2-thioxo-1-(4-methylphenyl)-1,3,8-triazaspiro[4.5-
]dec-3-yl)-2-trifluoromethylbenzonitrile, 27c, [RD53]
[0180] A mixture of 27b (0.059 g, 0.13 mmol) in HCl aq., 2N (1 ml)
and methanol (3 ml) was heated to reflux for 2 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (5 ml) and extracted with ethyl acetate (10 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 60:40) to yield 27c
(0.055 g, 0.012 mmol, 92%) as a white powder.
##STR00047##
[0181] .sup.1H NMR (Acetone-d.sub.6, 400 MHz) .delta. 1.93-1.99 (m,
1H), 2.00-2.04 (m, 1H), 2.18 (s, 3H), 2.24-2.28 (m, 2H), 2.38 (s,
3H), 2.61-2.72 (m, 4H), 7.18-7.20 (m, 2H), 7.32-7.35 (m, 2H), 8.03
(dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 8.16 (d, J=1.8 Hz, 1H),
8.22 (d, J=8.2 Hz, 1H); .sup.13C NMR (Acetone-d.sub.6, 100 MHz)
.delta. 20.3, 31.4, 45.1, 49.8, 65.1, 109.1, 114.8, 122.4 (q,
J=275.1 Hz), 127.7 (q, J=4.8 Hz), 130.0, 130.5, 131.9 (q, J=32.6
Hz), 132.6, 133.5, 135.6, 138.3, 139.4, 174.0, 180.6.
Example 28
4-(8-methyl-4-oxo-2-thioxo-1,3,8-triazaspiro[4.5]dec-3-yl)-2-trifluorometh-
ylbenzonitrile, 28a, [RD52]
[0182] Compound 28a was synthesized according to the procedure
described in U.S. Pat. No. 5,958,936.
##STR00048##
[0183] .sup.1H NMR (Acetone-d.sub.6, 400 MHz) .delta. 1.93-2.00 (m,
2H), 2.09-2.16 (m, 2H), 2.25 (s, 3H), 2.42-2.49 (m, 2H), 2.75-2.80
(m, 2H), 7.97 (dd, J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 8.11 (d,
J=1.8 Hz, 1H), 8.20 (d, J=8.2 Hz, 1H), 9.80 (bs, NH); .sup.13C NMR
(Acetone-d.sub.6, 100 MHz) .delta. 32.9, 45.4, 50.1, 62.3, 109.1,
114.8, 122.4 (q, J=271.6 Hz), 127.5 (q, J=4.8 Hz), 131.8 (q, J=32.7
Hz), 133.2, 135.6, 135.6, 138.0, 175.2, 180.4.
Example 29
4-[3-(4-hydroxybutyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trif-
luoromethylbenzonitrile, RU 59063
[0184] Compound RU 59063 was synthesized according to the procedure
described by Teutsch et al [J. Steroid. Biochem. Molec. Biol. 1994,
48(1), 111-119].
##STR00049##
[0185] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.55 (s, 6H),
1.58-1.62 (m, 2H), 1.86-1.89 (m, 2H), 2.25 (bs, OH), 3.65-3.71 (m,
4H), 7.74 (dd, J.sub.1=8.0 Hz, J.sub.2=1.8 Hz, 1H), 7.92 (d, J=1.8
Hz, 1H), 7.98 (d, J=8.0 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 23.1, 24.7, 29.6, 43.9, 61.7, 65.2, 109.7, 114.9, 121.9 (q,
J=272.6 Hz), 127.1 (q, J=4.8 Hz), 132.2, 133.7 (q, J=34.3 Hz),
135.2, 137.2, 175.3, 178.2.
Example 30
30-1). 1-methylaminocyclobutanecarbonitrile, 30a
[0186] Methylamine was bubbled into a refrigerated mixture of
cyclobutanone (0.21 g, 3 mmol) and trimethylsilyl cyanide (0.396 g,
4 mmol) until the volume doubled. The mixture was stirred 3 h and
then concentrated to dryness to obtain 30a (0.33 g,
quantitative).
30-2).
4-(5-methyl-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-yl)-2-trifluoro-
methylenzonitrile, 30b, [RD73]
[0187] A mixture of 1a (0.114 g, 0.5 mmol) and 30a (0.055 g, 0.5
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 0.5
h. To this mixture were added 10 ml of methanol and 2 ml of 2N HCl.
The second mixture was refluxed for 2 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (20
ml) and extracted with ethyl acetate (30 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 30b (0.148 g, 0.435 mmol, 87%) as a
white powder.
##STR00050##
[0188] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.95-2.06 (m, 1H),
2.21-2.32 (m, 1H), 2.58-2.71 (m, 4H), 3.44 (s, 3H), 7.77 (dd,
J.sub.1=8.2 Hz, J.sub.2=2.0 Hz, 1H), 7.89 (d, J=2.0 Hz, 1H), 7.93
(d, J=8.2 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
30.3, 30.4, 66.1, 109.7, 114.9, 121.9 (q, J=272.6 Hz), 126.9 (q,
J=4.8 Hz), 132.1, 133.2 (q, J=34.3 Hz), 135.2, 137.3, 175.1,
178.7.
30-3).
4-(5-methyl-6,8-dioxo-5,7-diazaspiro[3.4]oct-7-yl)-2-trifluoromethy-
lbenzonitrile, 30c, [RD74]
[0189] Hydrogen peroxide (2 ml, 30%) was added to the mixture of
30b (0.068 g, 0.2 mmol) in glacial acetic acid (3 ml). After being
stirred at room temperature for 10 h, the reaction mixture was
poured into ethyl acetate (20 ml) and then washed with water
(2.times.20 ml). The organic layer was dried over MgSO.sub.4,
concentrated and chromatographed (dichloromethane:acetone) to yield
30c (0.057 g, 0.176 mmol, 88%) as a white powder.
##STR00051##
[0190] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.91-2.35 (m, 1H),
2.21-2.31 (m, 1H), 2.50-2.61 (m, 4H), 3.12 (s, 3H), 7.89 (d, J=8.2
Hz, 1H), 7.97 (dd, J.sub.1=8.2 Hz, J.sub.2=2.0 Hz, 1H), 8.12 (d,
J=2.0 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.9,
25.4, 29.3, 63.4, 108.1, 115.1, 121.6 (q, J=272.6 Hz), 122.9 (q,
J=4.8 Hz), 127.9, 133.5 (q, J=34.3 Hz), 135.3, 136.5, 152.7,
174.4.
Example 31
31-1). 1-methylaminocyclopentanecarbonitrile, 31a
[0191] Methylamine was bubbled into a refrigerated mixture of
cyclopentanone (0.252 g, 3 mmol) and trimethylsilyl cyanide (0.396
g, 4 mmol) until the volume doubled. The mixture was stirred 3 h
and then concentrated to dryness to obtain 31a (0.372 g,
quantitative).
31-2).
4-(1-methyl-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2-trifluoro-
methylbenzonitrile, 31b, [RD75]
[0192] A mixture of 1a (0.114 g, 0.5 mmol) and 31a (0.062 g, 0.5
mmol) in dry DMF (0.2 ml) was stirred at room temperature for 0.5
h. To this mixture were added 10 ml of methanol and 2 ml of 2N HCl.
The second mixture was refluxed for 2 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (20
ml) and extracted with ethyl acetate (30 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane) to yield 31b (0.159 g, 0.45 mmol, 90%) as a white
powder.
##STR00052##
[0193] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.91-2.05 (m, 6H),
2.16-2.21 (m, 2H), 3.27 (s, 3H), 7.77 (dd, J.sub.1=8.2 Hz, =1.8 Hz,
1H), 7.89 (d, J=1.8 Hz, 1H), 7.91 (d, J=8.2 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 26.4, 30.3, 35.4, 73.2, 109.5, 114.9,
121.9 (q, J=272.6 Hz), 126.9 (q, J=4.8 Hz), 132.2, 133.2 (q, J=34.3
Hz), 135.2, 137.5, 176.8, 178.5.
31-3).
4-(1-methyl-2,4-dioxo-1,3-diaza-spiro[4.4]non-3-yl)-2-trifluorometh-
ylbenzonitrile, 31c, [RD76]
[0194] Hydrogen peroxide (2 ml, 30%) was added to the mixture of
31b (0.07 g, 0.2 mmol) in glacial acetic acid (3 ml). After being
stirred at room temperature for 10 h, the reaction mixture was
poured into ethyl acetate (20 ml) and then washed with water
(2.times.20 ml). The organic layer was dried over MgSO.sub.4,
concentrated and chromatographed (dichloromethane:acetone) to yield
31c (0.057 g, 0.168 mmol, 84%) as a white powder.
##STR00053##
[0195] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.88-1.99 (m, 6H),
2.12-2.17 (m, 2H), 2.98 (s, 3H), 7.88 (d, J=8.2 Hz, 1H), 7.97 (dd,
J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 8.12 (d, J=1.8 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 25.2, 26.5, 34.8, 70.1,
108.0, 115.1, 122.0 (q, J=272.5 Hz), 122.9 (q, J=4.9 Hz), 127.9,
133.5 (q, J=32.9 Hz), 135.3, 136.6, 152.7, 176.1.
Example 32
4-(8-methylimino-6-thioxo-5-p-tolyl-5,7-diaza-spiro[3.4]oct-7-yl)-2-triflu-
oromethyl-benzonitrile, 32a, [RD90]
[0196] A mixture of 7b (0.042 g, 0.1 mmol), DBU (0.023 g, 0.15
mmol) and iodomethane (0.073 g, 0.5 mmol) in DMF (0.3 ml) was
stirred for 15 h at room temperature. After DMF being evaporated,
the medium was chromatographed (dichloromethane) to yield 32a
(0.011 g, 0.026 mmol, 26%) as white powder.
##STR00054##
[0197] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.58-1.65 (m, 1H),
2.04-2.13 (m, 1H), 2.45 (s, 3H), 2.70-2.77 (m, 2H), 3.06-3.10 (m,
2H), 3.58 (s, CH.sub.3--N, major isomer) [2.70 (s, CH.sub.3--N,
minor isomer)], 7.20-7.34 (m, 4H), 7.75-7.91 (m, 3H); (CDCl.sub.3,
100 MHz) .delta. 12.6, 21.4, 30.2, 33.7 (35.3 for the other
isomer), 66.9, 109.1, 115.2, 122.1 (q, J=272.5 Hz), 128.5 (q, J=4.9
Hz), 129.8, 130.4, 130.6, 132.8, 133.2 (q, J=32.9 Hz), 133.5,
134.9, 139.8, 157.0, 180.2.
Example 33
1-[3-(4-cyano-3-trifluoromethyl-phenyl)-5,5-dimethyl-2-thioxo-1-p-tolyl-im-
idazolidin-4-ylidene]-3-ethyl-thiourea, 33a, [RD91]
[0198] A mixture of 5b (0.06 g, 0.149 mmol), ethylthioisocyanate
(0.087 g, 1 mmol) and CuI (0.01 g, 0.05 mmol) in DMF (0.1 ml) was
heated under microwave for 45 minutes. Then the medium was washed
with brine and extracted with ethyl acetate. The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed (HPLC,
alumina column) to yield 33a (0.054 g, 0.108 mmol, 72%) as white
powder.
##STR00055##
[0199] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.15 (t, J=7.23
Hz, 3H), 1.70 [1.75 minor isomer] (s, 6H), 2.42 (s, 3H), 3.28-3.39
(m, 2H) [3.15-3.22 (m, 2H), minor isomer], 6.50 (bs, 1H) [6.93 (bs,
1H), minor isomer], 7.14-7.18 (m, 2H), 7.32-7.35 (m, 2H), 7.77-7.94
(m, 3H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.31 (13.83
minor), 21.3, 25.22 (24.89 minor), 40.31 (40.67 minor), 68.1,
109.9, 114.9, 122.3 (q, J=272.5 Hz), 127.6 (q, J=4.9 Hz), 129.1,
129.59 (129.55 minor), 130.52 (130.57 minor), 132.27 (132.15
minor), 132.9 (q, J=32.9 Hz), 134.27 (134.15 minor), 134.9, 135.2,
156.33 (156.06 minor), 180.28 (180.06 minor), 187.24 (186.63
minor).
Example 34
1-[7-(4-cyano-3-trifluoromethyl-phenyl)-6-thioxo-5-p-tolyl-5,7-diaza-spiro-
[3.4]oct-8-ylidene]-3-phenyl-thiourea, 34a, [RD92]
[0200] A mixture of 7b (0.021 g, 0.05 mmol) and phenylthioisocyante
(0.027 g, 0.2 mmol) in DMF (0.3 ml) was stirred for 2 days at
60.degree. C. After DMF being evaporated, the medium was
chromatographed (dichloromethane) to yield 34a (0.015 g, 0.028
mmol, 57%) as white powder.
##STR00056##
[0201] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.59-1.67 (m, 1H),
2.12-2.22 (m, 1H), 2.45 (s, 3H), 2.61-2.71 (m, 2H), 2.81-2.87 (m,
2H), 7.18-7.27 (m, 6H), 7.33-7.41 (m, 5H), 7.60-7.62 (m, 1H), 8.40
(bs, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.6, 21.4,
32.3, 69.6, 110.7, 114.8, 121.6, 122.0 (q, J=272.5 Hz), 126.3,
128.0 (q, J=4.9 Hz), 128.9, 129.4, 130.7, 132.5, 133.2 (q, J=32.9
Hz), 134.1, 134.9, 137.7, 139.2, 140.2, 154.8, 180.3, 185.5.
Example 35
1-(4-Cyano-3-trifluoromethyl-phenyl)-3-[7-(4-cyano-3-trifluoromethyl-pheny-
l)-6-thioxo-5-p-tolyl-5,7-diaza-spiro[3.4]oct-8-ylidene]-thiourea,
35a, [RD93]
[0202] A mixture of 1a (0.5.02 g, 2.2 mmol) and 7a (0.186 g, 1
mmol) in DMF (1 ml) was stirred at room temperature. After 20 hours
of stirring, the mixture was concentrated under reduced pressure to
yield an orange viscous liquid, which was chromatographed
(dichloromethane:acetone, 99:1) to yield 35a (0.269 g, 0.42 mmol,
42%) as a yellow powder.
##STR00057##
[0203] The X-ray structure of 35a is illustrated in FIG. 30.
Example 36
36-1). 1-(4-hydroxymethylphenylamino)-cyclobutanecarbonitrile,
36a
[0204] Trimethylsilyl cyanide (0.66 ml, 5 mmol) was added dropwise
to a mixture of 4-aminobenzoic acid (0.492 g, 4 mmol) and
cyclobutanone (0.35 g, 5 mmol) in dichloromethane (10 ml). The
reaction mixture was stirred at room temperature for 6 h and then
concentrated under vacuum to obtain a brown liquid which was
subjected to chromatography (dichloromethane) to yield 36a (0.677
g, 3.36 mmol, 84%) as a brown solid.
36-2).
4-[8-(4-hydroxymethylphenyl)-5-oxo-7-thioxo-6-azaspiro[3.4]oct-6-yl-
]-2-trifluoromethyl-benzonitrile, 36b, [RD110]
[0205] A mixture of 1a (0.342 g, 1.5 mmol) and 36a (0.21 g, 1 mmol)
in dry DMF (0.5 ml) was stirred at room temperature for 24 h. To
this mixture were added methanol (20 ml) and HCl aq. 2N (5 ml). The
second mixture was refluxed for 6 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (40
ml) and extracted with ethyl acetate (60 ml). The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 90:10) to yield 36b (0.296 g, 0.69 mmol,
69%) as a white powder.
##STR00058##
[0206] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.68 (m, 1H),
2.17-2.26 (m, 1H), 2.52-2.68 (m, 4H), 4.75 (s, 2H), 7.30 (d, J=8.1
Hz, 2H), 7.58 (d, J=8.1 Hz, 2H), 7.88 (dd, J.sub.1=8.3 Hz,
J.sub.2=1.8 Hz, 1H), 7.95-7.98 (m, 2H); .sup.13C NMR (CDCl.sub.3,
100 MHz) .delta. 13.7, 31.5, 64.4, 67.5, 109.9, 114.9, 121.9 (q,
J=272.6 Hz), 127.1 (q, J=4.7 Hz), 128.3, 130.0, 132.2, 133.3, 133.4
(q, J=33.2 Hz), 134.2, 137.2, 142.9, 174.9, 179.9.
Example 37
4-[5-(4-formylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-yl]-2-triflu-
oromethyl-benzonitrile, 37a, [RD114]
[0207] To a mixture of 36b (0.303 g, 0.7 mmol) and Dess-Martin
periodinane (0.417 g, 1 mmol) in dichloromethane (5 ml) was added
pyridine (1.01 g, 1 mmol). The mixture was stirred for 2 hours at
room temperature and then ethyl ether (10 ml) was added to
precipitate the by-product of the reaction. After filtration and
concentration under reduced pressure, the mixture was
chromatographed (dichloromethane:acetone, 95:5) to yield 37a (0.24
g, 0.56 mmol, 80%) as white powder.
##STR00059##
[0208] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.62-1.73 (m, 1H),
2.24-2.30 (m, 1H), 2.50-2.58 (m, 2H), 2.69-2.75 (m, 2H), 7.53 (d,
J=8.1 Hz, 2H), 7.85 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H),
7.97-7.99 (m, 2H), 8.11 (d, J=8.1 Hz, 2H), 10.12 (s, 1H); .sup.13C
NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 31.7, 67.5, 110.2, 114.8,
121.9 (q, J=272.6 Hz), 127.0 (q, J=4.7 Hz), 129.1, 131.0, 131.2,
132.2, 133.3 (q, J=33.2 Hz), 135.3, 136.9, 140.5, 174.5, 179.8,
190.8.
Example 38
4-{5-[4-(1-hydroxyethyl)-phenyl]-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-y-
l}-2-trifluoromethyl-benzonitrile, 38a [RD116]
[0209] The mixture of 37a (0.043 g, 0.1 mmol) and dry THF (1 ml) in
a flamed-dried flash was placed under argon and cooled to
-78.degree. C. Then, methylmagnesium iodide (1.1 ml, 0.1 M) was
added. The mixture was stirred at -78.degree. C. for 30 minutes and
warmed slowly to room temperature. The medium was washed with water
(3 ml) and extracted with ethyl acetate (10 ml). The organic layer
was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 95:5) to yield 38a (0.037 g, 0.082 mmol,
82%) as a white powder.
##STR00060##
[0210] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57 (d, J=6.5 Hz,
3H), 1.61-1.71 (m, 1H), 2.09 (d, J=3.2 Hz, OH), 2.16-2.28 (m, 1H),
2.52-2.60 (m, 2H), 2.63-2.69 (m, 2H), 5.00 (dd, J.sub.1=6.5 Hz, q,
J.sub.2=3.1 Hz, 1H), 7.29 (d, J=8.3 Hz, 2H), 7.60 (d, J=8.2 Hz,
2H), 7.85 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.95-7.98 (m,
2H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 25.3, 31.5,
67.4, 69.8, 110.0, 114.9, 121.9 (q, J=272.6 Hz), 127.0 (q, J=4.7
Hz), 127.1, 129.9, 132.2, 133.4 (q, J=33.2 Hz), 134.1, 135.2,
137.1, 147.6, 174.9, 179.9.
Example 39
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.-
4]oct-5-yl]-phenyl}-acrylic acid ethyl ester, 39a [RD117]
[0211] A mixture of 37a (0.043 g, 0.1 mmol) and
(carbethoxyethylidene)triphenylphosphorane (0.039 g, 0.12 mmol) in
dichloromethane (2 ml) was stirred at room temperature for 10
hours. The medium was concentrated and chromatographed
(dichloromethane) to yield 39a (0.048 g, 0.096 mmol, 96%) as white
powder.
##STR00061##
[0212] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.35 (t, J=7.1 Hz,
3H), 1.66-1.70 (m, 1H), 2.19-2.65 (m, 1H), 2.51-2.69 (m, 2H),
2.66-2.72 (m, 2H), 4.28 (q, J=7.1 Hz, 2H), 6.51 (d, J=16.1 Hz, 1H),
7.35 (d, J=8.3 Hz, 2H), 7.72 (d, J=8.3 Hz, 2H), 7.73 (d, J=16.1 Hz,
1H), 7.85 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.96-7.98 (m,
2H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 14.3, 31.6,
60.8, 67.5, 110.0, 114.9, 120.5, 121.8 (q, J=272.6 Hz), 127.0 (q,
J=4.7 Hz), 129.5, 130.5, 132.2, 133.4 (q, J=33.2 Hz), 135.2, 136.0,
136.5, 137.0, 142.7, 166.5, 174.7, 179.8.
Example 40
4-{5-[4-(3-hydroxypropenyl)-phenyl]-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct--
7-yl}-2-trifluoromethylbenzonitrile, 40a [RD120]
[0213] To a mixture of 39a (0.05 g, 0.1 mmol) in dichloromethane (2
ml) at -78.degree. C. was added a solution of diisobutylaluminum
hydride in THF (0.11 ml, 1M, 0.11 mmol). The mixture was stirred at
-78.degree. C. for 3 hours. After being warmed to room temperature,
the mixture was washed with an aqueous solution of sodium
thiosulfate and extracted with ethyl acetate. The organic layer was
dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 95:5) to yield 40a (0.040 g, 0.089 mmol,
89%) as a white powder.
##STR00062##
[0214] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57-1.68 (m, 1H),
2.17-2.39 (m, 1H), 2.55-2.61 (m, 2H), 2.61-2.67 (m, 2H), 4.39 (d,
J=4.7 Hz, 2H), 6.47 (dt, J.sub.1=16.0 Hz, J.sub.2=5.3 Hz, 1H), 6.70
(d, J=16.0 Hz, 1H), 7.29 (d, J=8.3 Hz, 2H), 7.59 (d, J=8.3 Hz, 2H),
7.85 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.96-7.98 (m, 2H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 31.5, 63.4, 67.4,
110.0, 114.8, 120.5, 121.8 (q, J=272.6 Hz), 127.0 (q, J=4.7 Hz),
127.9, 129.2, 130.1, 131.1, 132.1, 133.4 (q, J=33.2 Hz), 135.2,
137.1, 138.4, 174.8, 179.9.
Example 41
41-1) 3-[4-(1-cyanocyclobutylamino)-phenyl]propionic acid, 41a
(41-1)
[0215] Trimethylsilyl cyanide (0.4 g, 4 mmol) was added dropwise to
a mixture of 3-(4-aminophenyl)-propionic acid (0.33 g, 2 mmol),
cyclobutanone (0.35 g, 5 mmol) and sodium sulfate (1 g) in
1,4-dioxane (5 ml). The mixture was stirred for 15 hours. After
filtration to eliminate sodium sulfate, the medium was concentrated
under vacuum to obtain a brown liquid which was subjected to
chromatography (dichloromethane:acetone, 50:50) to yield 41a (0.472
g, 1.93 mmol, 97%) as a yellowish solid.
41-2)
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazasp-
iro[3.4]oct-5-yl]-phenyl}-propionic acid methyl ester, 41b (41-2)
[RD128]
[0216] A mixture of 1a (0.661 g, 2.9 mmol) and 41a (0.472 g, 1.93
mmol) in dry DMF (2 ml) was stirred at room temperature for 15
hours. To this mixture were added methanol (10 ml) and HCl aq. (5
ml, 2M). The second mixture was refluxed for 3 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (10 ml) and extracted with ethyl acetate (3.times.30
ml). The organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 41b (0.582 g, 1.19 mmol,
62%) as a white powder.
##STR00063##
[0217] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60-1.70 (m, 1H),
2.14-2.26 (m, 1H), 2.51-2.56 (m, 2H), 2.58-2.67 (m, 2H), 2.71 (t,
J=7.8 Hz, 2H), 3.05 (t, J=7.8 Hz, 2H), 3.69 (s, 3H), 7.23 (d, J=8.2
Hz, 2H), 7.41 (d, J=8.2 Hz, 2H), 7.85 (dd, J.sub.1=8.3 Hz,
J.sub.2=1.8 Hz, 1H), 7.95 (d, J=8.3 Hz, 1H), 7.98 (d, J=1.8 Hz,
1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 30.5, 31.4,
35.1, 51.8, 67.5, 109.9, 114.9, 121.9 (q, J=272.7 Hz), 127.1 (q,
J=4.7 Hz), 129.9, 130.0, 133.2, 132.3, 133.3 (q, J=33.2 Hz), 135.7,
137.2, 142.5, 173.1, 174.9, 179.9.
41-3)
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-propionic acid, 41c (41-3) [RD132]
[0218] A mixture of 41b (0.487 g, 1 mmol) in methanol (10 ml) and
solution of sodium hydroxide (10 ml, 2M) was stirred at room
temperature for 5 hours. Methanol was evaporated. The residue was
adjusted to pH=5 by HCl aq. (2M) and then extracted with ethyl
acetate (3.times.50 ml). The organic layer was dried over
MgSO.sub.4 and concentrated to dryness to obtain 41c (0.472 g, 0.99
mmol, 99%).
41-4)
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-propionamide, 41d (41-4) [RD133]
[0219] To a suspension of 41c (0.094 g, 0.2 mmol) in THF (10 ml) at
-5.degree. C. was added thionyl chloride (0.019 ml, 0.26 mmol). The
medium was stirred at -5.degree. C. for one hour. Then ammonia was
bubbled into the mixture. The excess of ammonia was condensed by
reflux condenser at -78.degree. C. for 30 minutes and then was
allowed to evaporate. The medium was filtered. The filtrate was
concentrated and chromatographed (dichloromethane:acetone, 70:30)
to yield 41d (0.09 g, 0.19 mmol, 95%) as an off-white powder.
##STR00064##
[0220] .sup.1H NMR (acetone-d.sub.6, 400 MHz) .delta. 1.52-160 (m,
1H), 2.01-2.09 (m, 1H), 2.49-2.58 (m, 4H), 2.61-2.67 (m, 2H), 2.98
(t, J=7.5 Hz, 2H), 6.20 (bs, 1H), 6.78 (bs, 1H), 7.31 (d, J=8.2 Hz,
2H), 7.44 (d, J=8.2 Hz, 2H), 8.03 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8
Hz, 1H), 8.15 (d, J=1.8 Hz, 1H), 8.22 (d, J=8.3 Hz, 1H); .sup.13C
NMR (acetone-d.sub.6, 100 MHz) .delta. 13.4, 30.7, 31.2, 36.4,
67.5, 109.0, 114.8, 122.5 (q, J=271.5 Hz), 127.5 (q, J=4.7 Hz),
129.5, 130.0, 131.8 (q, J=32.5 Hz), 133.3, 133.8, 135.6, 138.4,
143.2, 171.6, 174.9, 178.0.
41-5)
3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-N-methyl-propionamide, 41e (41-5)
[RD134]
[0221] To a suspension of 41c (0.094 g, 0.2 mmol) in THF (10 ml) at
-5.degree. C. was added thionyl chloride (0.019 ml, 0.26 mmol). The
medium was stirred at -5.degree. C. for one hour. Then methylamine
was bubbled into the mixture at -5.degree. C. for 30 minutes. The
medium was filtered. The filtrate was concentrated and
chromatographed (dichloromethane:acetone, 75:25) to yield 41e
(0.092 g, 0.19 mmol, 95%) as an off-white powder.
##STR00065##
[0222] .sup.1H NMR (acetone-d.sub.6, 400 MHz) .delta. 1.51-1.60 (m,
1H), 2.01-2.11 (m, 1H), 2.48-2.58 (m, 4H), 2.61-2.67 (m, 2H), 2.77
(d, J=4.6 Hz, 3H), 2.98 (t, J=7.5 Hz, 2H), 7.03 (bs, NH), 7.33 (d,
J=8.2 Hz, 2H), 7.42 (d, J=8.2 Hz, 2H), 8.01 (dd, J.sub.1=8.3 Hz,
J.sub.2=1.8 Hz, 1H), 8.13 (d, J=1.8 Hz, 1H), 8.20 (d, J=8.3 Hz,
1H); .sup.13C NMR (acetone-d.sub.6, 100 MHz) .delta. 13.4, 25.3,
30.0, 31.2, 37.0, 67.6, 109.0, 114.8, 122.5 (q, J=271.5 Hz), 127.4
(q, J=4.7 Hz), 129.5, 130.0, 131.9 (q, J=32.5 Hz), 133.3, 133.8,
135.6, 138.4, 143.1, 171.7, 175.0, 178.0.
41-6)
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-N-(2-hydroxyethyl)-propionamide, 41f
(41-6) [RD135]
[0223] To a suspension of 41c (0.094 g, 0.2 mmol) in THF (10 ml) at
-5.degree. C. was added thionyl chloride (0.019 ml, 0.26 mmol). The
medium was stirred at -5.degree. C. for one hour. Then
2-aminoethanol (0.0183 g, 0.03 mmol) was added into the mixture at
-5.degree. C. After stirring of an additional 30 minutes, the
medium was filtered. The filtrate was concentrated and
chromatographed (dichloromethane:acetone, 50:50) to yield 41f
(0.093 g, 0.18 mmol, 90%) as an off-white powder.
##STR00066##
[0224] .sup.1H NMR (acetone-d.sub.6, 400 MHz) .delta. 1.51-161 (m,
1H), 2.01-2.11 (m, 1H), 2.49-2.66 (m, 6H), 2.99 (t, J=7.5 Hz, 2H),
3.27 (dd, J.sub.1=11.2 Hz, J.sub.2=5.6 Hz, 3H), 3.51 (dd,
J.sub.1=11.2 Hz, J.sub.2=5.6 Hz, 2H), 3.87 (bs, OH), 7.20 (bs, NH),
7.33 (d, J=8.2 Hz, 2H), 7.43 (d, J=8.2 Hz, 2H), 8.02 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 8.14 (d, J=1.8 Hz, 1H), 8.22
(d, J=8.3 Hz, 1H); .sup.13C NMR (acetone-d.sub.6, 100 MHz) .delta.
13.4, 31.0, 31.2, 37.1, 42.0, 61.2, 67.6, 109.0, 114.8, 122.5 (q,
J=271.5 Hz), 127.4 (q, J=4.7 Hz), 129.6, 130.0, 131.9 (q, J=32.5
Hz), 133.3, 133.8, 135.6, 138.4, 143.0, 171.9, 175.0, 178.1.
42-1) 4-[4-(1-Cyanocyclobutylamino)-phenyl]butyric acid, 42a
[0225] Trimethylsilyl cyanide (0.50 g, 5 mmol) was added dropwise
to a mixture of 4-(4-aminophenyl)-butyric acid (0.537 g, 3 mmol),
cyclobutanone (0.35 g, 5 mmol) and sodium sulfate (1 g) in
1,4-dioxane (10 ml). The mixture was stirred for 15 hours. After
filtration to eliminate sodium sulfate, the medium was concentrated
under vacuum to obtain a brown liquid which was subjected to
chromatography (dichloromethane:acetone, 50:50) to yield 42a (0.665
g, 2.58 mmol, 86%) as a yellowish solid.
42-2)
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazasp-
iro[3.4]oct-5-yl]-phenyl}-butyric acid methyl ester, 42b
[RD129]
[0226] A mixture of 1a (0.547 g, 2.4 mmol) and 42a (0.342 g, 1.5
mmol) in dry DMF (2 ml) was stirred at room temperature for 15
hours. To this mixture were added methanol (10 ml) and HCl aq. (5
ml, 2M). The second mixture was refluxed for 3 h. After being
cooled to room temperature, the reaction mixture was poured into
cold water (10 ml) and extracted with ethyl acetate (3.times.30
ml). The organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane) to yield 42b (0.594 g, 1.18 mmol,
79%) as a white powder.
##STR00067##
[0227] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60-1.70 (m, 1H),
1.98-2.07 (m, 2H), 2.14-2.26 (m, 1H), 2.40 (t, J=7.4 Hz, 2H),
2.52-2.60 (m, 2H), 2.62-2.68 (m, 2H), 2.74 (t, J=7.4 Hz, 2H), 3.68
(s, 3H), 7.22 (d, J=8.2 Hz, 2H), 7.38 (d, J=8.2 Hz, 2H), 7.86 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (d, J=8.3 Hz, 1H), 7.98
(d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
26.1, 31.4, 33.5, 34.8, 51.7, 67.5, 109.9, 114.9, 121.9 (q, J=272.7
Hz), 127.1 (q, J=4.7 Hz), 129.7, 130.1, 132.3, 133.0, 133.3 (q,
J=33.2 Hz), 135.2, 137.2, 143.5, 173.8, 175.0, 179.9.
42-3)
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-butyric acid, 42c [RD141]
[0228] A mixture of 42b (0.501 g, 1 mmol) in methanol (10 ml) and
solution of sodium hydroxide (10 ml, 2M) was stirred at room
temperature for 5 hours. The methanol was evaporated. The residue
was adjusted to pH=5 by HCl aq. (2M) and then, the medium was
extracted with ethyl acetate (3.times.50 ml). The organic layer was
dried over MgSO.sub.4 and concentrated to dryness to obtain 42c
(0.482 g, 0.99 mmol, 99%), the structure of which is illustrated in
Formula 5.
##STR00068##
[0229] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60-1.70 (m, 1H),
1.98-2.07 (m, 2H), 2.14-2.26 (m, 1H), 2.45 (t, J=7.3 Hz, 2H),
2.51-2.59 (m, 2H), 2.62-2.68 (m, 2H), 2.77 (t, J=7.3 Hz, 2H), 7.23
(d, J=8.1 Hz, 2H), 7.40 (d, J=8.1 Hz, 2H), 7.85 (dd, J=8.3, 1.8 Hz,
1H), 7.95 (d, J=8.3 Hz, 1H), 7.97 (d, J=1.8 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 13.7, 25.9, 31.4, 33.4, 34.7, 67.5,
109.9, 114.9, 121.9 (q, J=272.6 Hz), 127.1 (q, J=4.7 Hz), 129.8,
130.1, 132.3, 133.0, 133.4 (q, J=33.1 Hz), 135.2, 137.2, 143.3,
174.9, 178.9, 179.9.
42-4)
4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-butyramide, 42d [RD130]
[0230] To a suspension of 42c (0.097 g, 0.2 mmol) in THF (10 ml) at
-5.degree. C. was added thionyl chloride (0.019 ml, 0.26 mmol). The
medium was stirred at -5.degree. C. for one hour. Then ammonia was
bubbled into the mixture. The excess of ammonia was condensed by
reflux condenser at -78.degree. C. for 30 minutes and then was
allowed to evaporate. The medium was filtered. The filtrate was
concentrated and chromatographed (dichloromethane:acetone, 70:30)
to yield 42d (0.093 g, 0.19 mmol, 95%) as an off-white powder.
##STR00069##
[0231] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57-1.70 (m, 1H),
2.00-2.08 (m, 2H), 2.16-2.25 (m, 1H), 2.31 (t, J=7.3 Hz, 2H),
2.51-2.59 (m, 2H), 2.62-2.68 (m, 2H), 2.77 (t, J=7.3 Hz, 2H), 5.56
(bs, 1H), 5.65 (bs, 1H), 7.22 (d, J=8.2 Hz, 2H), 7.39 (d, J=8.2 Hz,
2H), 7.85 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (d, J=8.3
Hz, 1H), 7.97 (d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 13.7, 26.5, 31.4, 34.8, 35.0, 67.5, 109.9, 114.9, 121.9 (q,
J=272.7 Hz), 127.1 (q, J=4.7 Hz), 129.8, 130.1, 132.2, 133.0, 133.3
(q, J=33.2 Hz), 135.2, 137.2, 143.5, 173.8, 174.9, 179.9.
42-5)
4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-s-
piro[3.4]oct-5-yl]-phenyl}-N-methyl-butyramide, 42e [RD131]
[0232] To a suspension of 42c (0.097 g, 0.2 mmol) in THF (10 ml) at
-5.degree. C. was added thionyl chloride (0.019 ml, 0.26 mmol). The
medium was stirred at -5.degree. C. for one hour. Then methylamine
was bubbled into the mixture at -5.degree. C. for 30 minutes. The
medium was filtered. The filtrate was concentrated and
chromatographed (dichloromethane:acetone, 75:25) to yield 42e
(0.095 g, 0.19 mmol, 95%) as an off-white powder.
##STR00070##
[0233] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.52-1.64 (m, 1H),
1.94-2.01 (m, 2H), 2.10-2.17 (m, 1H), 2.20 (t, J=7.3 Hz, 2H),
2.46-2.62 (m, 4H), 2.69 (t, J=7.3 Hz, 2H), 2.73 (d, J=4.7 Hz, 3H),
6.09 (bs, 1H), 7.16 (d, J=8.2 Hz, 2H), 7.33 (d, J=8.2 Hz, 2H), 7.82
(dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.91 (d, J=8.3 Hz, 1H),
7.94 (d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta.
13.7, 26.2, 26.8, 31.4, 35.0, 35.7, 67.5, 109.7, 114.9, 121.9 (q,
J=272.7 Hz), 127.1 (q, J=4.7 Hz), 129.7, 130.0, 132.3, 133.8, 133.3
(q, J=33.2 Hz), 135.2, 137.3, 143.7, 173.3, 174.9, 179.8.
42-6)
N-(4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaz-
a-spiro[3.4]oct-5-yl]phenyl}-butanoyl)-methanesulfonamide, 42f
[RD157]
[0234] A mixture of
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[-
3.4]oct-5-yl]phenyl}butanoic acid (42c) (0.049 g, 0.1 mmol),
2,4,6-trichlorobenzoyl chloride (0.244 g, 1 mmol),
4-dimethylaminopyridine (0.122 g, 1 mmol) and methanesulfonamide
(0.019 g, 0.2 mmol) in dichloromethane was stirred at room
temperature for 20 hours. The mixture was concentrated and
chromatographed (dichloromethane:acetone, 80:20) to yield
N-(4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spi-
ro[3.4]oct-5-yl]phenyl}-butanoyl)-methanesulfonamide (420 [RD157]
(0.053 g, 0.094 mmol, 94%), the structure of which is illustrated
in Formula 8, as a white powder.
##STR00071##
[0235] .sup.1H NMR (acetone-d.sub.6, 400 MHz) .delta. 1.51-160 (m,
1H), 1.96-2.11 (m, 3H), 2.49 (t, J=7.3 Hz, 2H), 2.51-2.57 (m, 2H),
2.61-2.67 (m, 2H), 2.75 (t, J=7.5 Hz, 2H), 2.94 (bs, 1H), 3.24 (s,
3H), 7.33 (d, J=8.3 Hz, 2H), 7.43 (d, J=8.2 Hz, 2H), 8.02 (dd,
J=8.3, 1.6 Hz, 1H), 8.02 (d, J=1.6 Hz, 1H), 8.21 (d, J=8.3 Hz, 1H);
.sup.13C NMR (acetone-d.sub.6, 100 MHz) .delta. 13.4, 25.8, 31.2,
34.3, 35.2, 40.6, 67.6, 109.0, 114.8, 122.5 (q, J=271.5 Hz), 127.5
(q, J=4.9 Hz), 129.6, 130.1, 131.9 (q, J=33.6 Hz), 133.3, 133.9,
135.6, 138.4, 143.1, 171.9, 175.0, 180.5.
42-7)
N-methyl-4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-6,8-dioxo-5,7-dia-
zaspiro[3.4]oct-5-yl]-phenyl}butyramide, 42g [RD158]
[0236] Hydrogen peroxide (30%, 0.4) was added dropwise to a
solution of
N-methyl-4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-dia-
za-spiro[3.4]oct-5-yl]phenyl}butanamide (42e) (0.032 g, 0.064 mmol)
in glacial acetic acid (0.5 ml). The mixture was stirred at room
temperature for 5 hours and then washed with water and extracted
with ethyl acetate. The organic layer was dried over magnesium
sulfate, concentrated and chromatographed (dichloromethane:acetone,
80:20) to yield
N-methyl-4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-6,8-dioxo-5,7-diazaspi-
ro[3.4]oct-5-yl]-phenyl}butyramide (42g) [RD158] (0.029 g, 0.06
mmol, 94%), the structure of which is illustrated in Formula 9, as
a white powder.
##STR00072##
[0237] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.71 (m, 1H),
1.93-2.04 (m, 2H), 2.18-2.27 (m, 3H), 2.44-2.53 (m, 2H), 2.57-2.65
(m, 2H), 2.70 (t, J=7.3 Hz, 2H), 2.79 (d, J=4.8 Hz, 3H), 5.79 (bs,
1H), 7.21 (d, J=8.2 Hz, 2H), 7.34 (d, J=8.2 Hz, 2H), 7.92 (d, J=8.4
Hz, 1H), 8.03 (dd, J=8.3, 1.8 Hz, 1H), 8.18 (d, J=1.8 Hz, 1H).
Example 43
43-1) 4-(4-aminophenyl)-piperazine-1-carboxylic acid tert-butyl
ester, 43a
[0238] A mixture of 4-iodoaniline (0.654 g, 3 mmol),
piperazine-1-carboxylic acid tert-butyl ester (0.67 g, 3.6 mmol),
potassium phosphate (1.272 g, 6 mmol), ethylene glycol (0.33 ml)
and copper iodide (0.03 g, 0.15 mmol) in 2-propanol (3 ml) was
placed under argon in a sealed-tube and heated to 80.degree. C. for
30 hours. After being cooled to room temperature, the medium was
washed with water (50 ml) and extracted with ethyl acetate (100
ml). The organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 70:30) to yield 43a (0.36
g, 1.3 mmol, 43%) as a yellow powder.
43-2) 4-[4-(1-cyanocyclobutylamino)phenyl]-piperazine-1-carboxylic
acid tert-butyl ester, 43b
[0239] Trimethylsilyl cyanide (0.3 g, 3 mmol) was added dropwise to
a mixture of 43a (0.415 g, 1.5 mmol), cyclobutanone (0.21 g, 3
mmol) and sodium sulfate (1 g) in dichloromethane (5 ml). The
mixture was stirred for 15 hours. After filtration to eliminate
sodium sulfate, the medium was concentrated under vacuum to obtain
a brown liquid which was subjected to chromatography
(dichloromethane:acetone, 75:25) to yield 43b (0.448 g, 1.26 mmol,
84%) as a yellow solid.
43-3)
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-imino-6-thioxo-5,7-diaza-
spiro[3.4]oct-5-yl]-phenyl}-piperazine-1-carboxylic acid tert-butyl
ester, 43c [RD139] and
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-(4-cyano-3-trifluoromethyl-ph-
enylthiocarbamoylimino)-6-thioxo-5,7-diazaspiro[3.4]oct-5-yl]-phenyl}-pipe-
razine-1-carboxylic acid tert-butyl ester, 43d [RD140]
[0240] A mixture of 1a (0.228 g, 1 mmol) and 43b (0.472 g, 0.63
mmol) in dry DMF (1 ml) was stirred at room temperature for 20
hours. The mixture was concentrated and chromatographed
(dichloromethane:acetone, 90:10) to yield 43c (0.173 g, 0.296 mmol,
47%), the structure of which is illustrated in Formula 10, as a
off-white powder and 43d (0.169 g, 0.21 mmol, 33%), the structure
of which is illustrated in Formula 11, as a yellow powder.
##STR00073##
[0241] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.48, (s, 9H),
1.57-1.67 (m, 1H), 2.01-2.09 (m, 1H), 2.59-2.70 (m, 4H), 3.25 (t,
J=5.1 Hz, 4H), 3.59 (t, J=4.9 Hz, 4H), 7.02 (d, J=8.9 Hz, 2H), 7.20
(d, J=8.9 Hz, 2H), 7.81 (d, J=7.4 Hz, 1H), 7.93 (s, 1H), 7.97 (d,
J=8.1 Hz, 1H).
##STR00074##
[0242] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.48, (s, 9H),
1.57-1.64 (m, 1H), 2.01-2.10 (m, 1H), 2.60-2.89 (m, 4H), 3.24 (t,
J=5.1 Hz, 4H), 3.57 (t, J=4.9 Hz, 4H), 7.02 (d, J=8.9 Hz, 2H), 7.20
(d, J=8.9 Hz, 2H), 7.54-7.98 (m, 4H), 7.97 (d, J=8.1 Hz, 1H).
43-4)
4-[8-Oxo-5-(4-piperazin-1-yl-phenyl)-6-thioxo-5,7-diazaspiro[3.4]oct-
-7-yl]-2-trifluoromethylbenzonitrile, 43e [RD137]
[0243] A mixture of 43c (0.117 g, 0.2 mmol), methanol (5 ml) and
HCl aq. (2 ml, 2M) was refluxed for 2 hours. After being cooled to
room temperature, the reaction mixture was poured into cold water
(10 ml) and extracted with ethyl acetate (3.times.30 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 50:50 and then
methanol:acetone, 50:50) to yield 43e (0.089 g, 0.184 mmol, 92%) as
a white powder.
##STR00075##
[0244] .sup.1H NMR (CD.sub.3OD, 400 MHz) .delta. 1.51-1.61 (m, 1H),
2.01-2.11 (m, 1H), 2.48-2.59 (m, 4H), 2.90-2.97 (m, 4H), 3.25-3.30
(m, 4H), 7.03 (d, J=8.9 Hz, 2H), 7.16 (d, J=8.9 Hz, 2H), 7.86 (dd,
J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 8.02 (d, J=8.3 Hz, 1H), 8.07
(d, J=1.8 Hz, 1H); .sup.13C NMR (CD.sub.3OD, 100 MHz) .delta. 13.2,
30.9, 45.1, 48.9, 67.5, 108.9, 114.8, 115.9, 122.3 (q, J=271.7 Hz),
126.4, 127.3 (q, J=4.7 Hz), 130.4, 132.2 (q, J=33.2 Hz), 133.0,
135.4, 138.1, 152.1, 175.4, 180.4.
43-5)
4-{5-[4-(4-methanesulfonylpiperazin-1-yl)-phenyl]-8-oxo-6-thioxo-5,7-
-diazaspiro[3.4]oct-7-yl}-2-trifluoromethylbenzonitrile, 43f
[RD138]
[0245] A mixture of 43e (0.049 g, 0.1 mmol), methanesulfonyl
chloride (0.012 ml, 0.15 mmol) and triethylamine (0.15 ml) in
dichloromethane was stirred at room temperature for 5 hours. The
medium was filtered. The filtrate was concentrated and
chromatographed (dichloromethane:acetone, 95:5) to yield 43f (0.042
g, 0.074 mmol, 74%) as a white powder.
##STR00076##
[0246] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.62-1.70 (m, 1H),
2.14-2.23 (m, 1H), 2.51-2.58 (m, 2H), 2.61-2.67 (m, 2H), 2.84 (s,
3H), 3.39 (s, 8H), 7.05 (d, J=8.9 Hz, 2H), 7.20 (d, J=8.9 Hz, 2H),
7.84 (dd, J.sub.1=8.3 Hz, J.sub.2=1.8 Hz, 1H), 7.95 (d, J=8.3 Hz,
1H), 7.97 (d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 13.7, 31.4, 34.6, 45.7, 48.4, 67.5, 109.8, 114.9, 117.0,
121.9 (q, J=272.7 Hz), 126.8, 127.1 (q, J=4.7 Hz), 130.7, 132.3,
133.4 (q, J=33.2 Hz), 135.2, 137.3, 151.1, 175.0, 180.2.
Example 44
44-1)
3-{4-[7-(4-Cyano-3-trifluoromethyl-phenyl)-8-oxo-6-thioxo-5,7-diaza--
spiro[3.4]oct-5-yl]-phenyl}-acrylic acid, 44a
[0247] A mixture of 39a (0.025 g, 0.05 mmol) in methanol (2 ml) and
solution of sodium hydroxide (2 ml, 2M) was stirred at room
temperature for 5 hours. Methanol was evaporated. The residue was
adjusted to pH=5 by HCl aq. (2M) and then extracted with ethyl
acetate (3.times.50 ml). The organic layer was dried over
MgSO.sub.4 and concentrated to dryness to obtain 44a (0.02 g, 0.042
mmol, 85%).
44-2)
3-{4-[7-(4-Cyano-3-trifluoromethyl-phenyl)-8-oxo-6-thioxo-5,7-diaza--
spiro[3.4]oct-5-yl]-phenyl}-acrylamide, 44b [RD119]
[0248] To a suspension of 44b (0.02 g, 0.042 mmol) in THF (1 ml) at
-5.degree. C. was added thionyl chloride (0.007 ml, 0.1 mmol). The
medium was stirred at -5.degree. C. for one hour. Then ammonia was
bubbled into the mixture. The excess of ammonia was condensed by
reflux condenser at -78.degree. C. for 30 minutes and then was
allowed to evaporate. The medium was filtered. The filtrate was
concentrated and chromatographed (dichloromethane:acetone, 70:30)
to yield 44b (0.014 g, 0.03 mmol, 71%) as an off-white powder.
##STR00077##
[0249] .sup.1H NMR (DMSO-d.sub.6, 400 MHz) .delta. 1.49-1.52 (m,
1H), 1.88-1.93 (m, 1H), 2.37-2.46 (m, 2H), 2.57-2.62 (m, 2H), 6.66
(d, J=15.9 Hz, 1H), 7.16 (bs, 1H), 7.43 (d, J=8.3 Hz, 2H), 7.47 (d,
J=15.9 Hz, 1H), 7.58 (bs, 1H), 8.03 (dd, J.sub.1=8.3 Hz,
J.sub.2=1.8 Hz, 1H), 8.23 (d, J=1.8 Hz, 1H), 8.34 (d, J=8.3 Hz,
1H).
Example 45 [RD145]
[0250] Trimethylsilyl cyanide (0.4 g, 4 mmol) was added dropwise to
a mixture of 4-methanesulfonylphenylamine hydrochloride (0.415 g, 2
mmol), cyclobutanone (0.28 g, 4 mmol) and sodium sulfate (1 g) in
DMF (3 ml). The mixture was stirred for 15 hours at 120.degree. C.
After filtration to remove the sodium sulfate, the filtrate was
washed with brine and extracted with ethyl acetate. The organic
layer was concentrated and chromatographed
(dichloromethane:acetone, 90:10) to yield
1-(4-methanesulfonylphenylamino)cyclobutanecarbonitrile (45a)
(0.116 g, 0.44 mmol, 22%) as a yellowish solid.
4-methanesulfonylphenylamine (0.201 g, 1.17 mmol, 59%) was also
recovered.
[0251] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.0.141 g, 0.62 mmol) and
1-(4-methanesulfonylphenylamino)cyclobutanecarbonitrile (45a) (0.11
g, 0.42 mmol) in dry DMF (2 ml) was stirred at room temperature for
3 days. To this mixture were added methanol (10 ml) and aq. 2N HCl
(5 ml). The second mixture was refluxed for 3 h. After being cooled
to room temperature, the reaction mixture was poured into cold
water (10 ml) and extracted with ethyl acetate (3.times.30 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 97:3) to yield
4-[5-(4-methanesulfonylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-yl-
]-2-trifluoromethylbenzonitrile (45b) [RD145] (0.031 g, 0.065 mmol,
15%), the structure of which is illustrated in Formula 14, as a
white powder.
##STR00078##
[0252] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.72 (m, 1H),
2.21-2.28 (m, 1H), 2.46-2.54 (m, 2H), 2.68-2.74 (m, 2H), 3.16 (s,
3H), 7.57 (d, J=8.3 Hz, 2H), 7.85 (dd, J=8.3, 1.8 Hz, 1H), 7.97 (d,
J=1.8 Hz, 1H), 7.98 (d, J=8.3 Hz, 1H), 8.17 (d, J=8.3 Hz, 2H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.6, 31.8, 44.4, 67.5,
110.2, 114.8, 122.4 (q, J=271.5 Hz), 127.0 (q, J=4.9 Hz), 129.4,
131.4, 132.1, 133.6 (q, J=33.3 Hz), 135.3, 136.8, 140.3, 141.8,
174.4, 179.9.
Example 46
[0253] Trimethylsilyl cyanide (0.69 g, 7 mmol) was added dropwise
to a mixture of 4-aminophenylacetic acid (0.755 g, 5 mmol) and
cyclobutanone (0.49 g, 7 mmol) in dioxane (20 ml). The mixture was
stirred for 8 hours at 80.degree. C. The mixture was concentrated
and chromatographed (dichloromethane:acetone, 60:40) to yield
[4-(1-cyanocyclobutylamino)phenyl]acetic acid (46a) (1.138 g, 4.95
mmol, 99%) as a white solid.
46-1) RD146
[0254] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.638 g, 2.8 mmol) and
[4-(1-cyanocyclobutylamino)phenyl]acetic acid (46a) (0.46 g, 2.0
mmol) in DMF (5 ml) was stirred at room temperature for 15 hours.
To this mixture were added methanol (20 ml) and aq. 2N HCl (10 ml).
The second mixture was refluxed for 1 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (10
ml) and extracted with ethyl acetate (3.times.50 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane pure and then dichloromethane:acetone, 95:5) to
yield
{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[3.-
4]oct-5-yl]phenyl}acetic acid methyl ester (46b) [RD146] (0.532 g,
1.124 mmol, 56%), the structure of which is illustrated in Formula
15, as a white powder.
##STR00079##
[0255] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60-1.69 (m, 1H),
2.15-2.25 (m, 1H), 2.50-2.58 (m, 2H), 2.61-2.66 (m, 2H), 3.72 (bs,
5H), 7.27 (d, J=8.3 Hz, 2H), 7.50 (d, J=8.3 Hz, 2H), 7.84 (dd,
J=8.3, 1.8 Hz, 1H), 7.94 (d, J=8.2 Hz, 1H), 7.97 (d, J=1.6 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 31.4, 44.7, 52.3,
67.4, 109.9, 114.9, 122.0 (q, J=272.5 Hz), 127.0 (q, J=4.9 Hz),
130.0, 131.1, 132.3, 133.0 (q, J=33.3 Hz), 134.1, 135.2, 135.9,
137.2, 171.4, 174.9, 179.9.
46-2) RD147
[0256] A mixture of
{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[3.-
4]oct-5-yl]phenyl}acetic acid methyl ester (46b) (0.095 g, 0.2
mmol) and a solution of sodium hydroxide (1 ml, 2M) in methanol (2
ml) was stirred at room temperature for 2 hours. The methanol was
evaporated. The residue was adjusted to pH 5 by aq. 2M HCl and then
the mixture was extracted with ethyl acetate (3.times.10 ml). The
organic layer was dried over MgSO.sub.4 and concentrated to dryness
to obtain
{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[3.-
4]oct-5-yl]phenyl}acetic acid (46c) [RD147] (0.087 g, 0.19 mmol,
95%), the structure of which is illustrated in Formula 16.
##STR00080##
[0257] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60-1.69 (m, 1H),
2.15-2.25 (m, 1H), 2.50-2.64 (m, 4H), 3.73 (s, 2H), 7.26 (d, J=8.3
Hz, 2H), 7.51 (d, J=8.3 Hz, 2H), 7.84 (dd, J=8.3, 1.8 Hz, 1H), 7.95
(d, J=8.2 Hz, 1H), 7.96 (d, J=1.6 Hz, 1H); .sup.13C NMR
(CDCl.sub.3, 100 MHz) .delta. 13.7, 31.4, 40.2, 40.8, 67.4, 109.9,
114.9, 122.0 (q, J=272.5 Hz), 127.0 (q, J=4.9 Hz), 129.9, 131.2,
132.3, 133.3 (q, J=33.3 Hz), 133.9, 135.2, 136.1, 137.2, 174.1,
174.9, 179.9.
46-3) RD148
[0258] Thionyl chloride (0.238 g, 2 mmol) was added dropwise to a
mixture of
{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro-
[3.4]oct-5-yl]phenyl}acetic acid (46c) (0.357 g, 0.777 mmol) in THF
(5 ml) cooled to 0.degree. C. The mixture was stirred for 1 hour at
room temperature and then ammonia was bubbled into the mixture. The
excess ammonia was condensed by a reflux condenser at -78.degree.
C. for 30 minutes and then was allowed to evaporate. The medium was
filtered and the filtrate was concentrated and chromatographed
(dichloromethane:acetone, 70:30) to yield
2-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[-
3.4]oct-5-yl]phenyl}acetamide (46d) [RD148] (0.345 g, 0.75 mmol,
97%), the structure of which is illustrated in Formula 17, as an
off-white powder.
##STR00081##
[0259] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.62-1.66 (m, 1H),
2.18-2.23 (m, 1H), 2.49-2.55 (m, 2H), 2.61-2.66 (m, 2H), 3.63 (s,
2H), 5.91 (bs, 1H), 6.10 (bs, 1H), 7.27 (d, J=8.1 Hz, 2H), 7.50 (d,
J=8.1 Hz, 2H), 7.83 (dd, J=8.3, 1.8 Hz, 1H), 7.95 (d, J=8.2 Hz,
1H), 7.96 (d, J=1.6 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 13.7, 31.5, 42.5, 67.4, 109.9, 114.9, 121.9 (q, J=272.4
Hz), 127.1 (q, J=4.9 Hz), 130.2, 131.1, 132.2, 133.3 (q, J=33.3
Hz), 134.1, 135.2, 136.8, 137.2, 172.8, 174.8, 180.0.
46-4) RD149
[0260] Thionyl chloride (0.238 g, 2 mmol) was added dropwise to a
mixture of
{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro-
[3.4]oct-5-yl]phenyl}acetic acid (46c) (0.357 g, 0.777 mmol) in THF
(5 ml) cooled to 0.degree. C. The mixture was stirred for 1 hour at
room temperature and then methylamine (0.5 ml) was added into the
mixture. The mixture was stirred for an additional 2 hours. The
medium was filtered and the filtrate was concentrated and
chromatographed (dichloromethane:acetone, 80:20) to yield
N-methyl-2-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-dia-
za-spiro[3.4]oct-5-yl]phenyl}acetamide (46e) [RD149] (0.348 g,
0.738 mmol, 95%), the structure of which is illustrated in Formula
18, as an off-white powder.
##STR00082##
[0261] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.61-1.70 (m, 1H),
2.17-2.31 (m, 1H), 2.50-2.56 (m, 2H), 2.61-2.68 (m, 2H), 2.82 (d,
J=4.8 Hz, 3H), 3.62 (s, 2H), 7.27 (d, J=8.3 Hz, 2H), 7.50 (d, J=8.3
Hz, 2H), 7.84 (dd, J=8.3, 1.8 Hz, 1H), 7.95 (d, J=8.2 Hz, 1H), 7.96
(d, J=1.6 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
26.6, 31.5, 43.1, 67.4, 110.0, 114.9, 122.0 (q, J=272.5 Hz), 127.1
(q, J=4.9 Hz), 130.2, 131.0, 132.2, 133.3 (q, J=33.3 Hz), 134.1,
135.2, 137.0, 137.1, 170.1, 174.8, 179.9.
Example 47
N-{4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imid-
azolidin-1-yl]phenyl}methanesulfonamide (47a) [RD150]
[0262] A mixture of
4-[3-(4-aminophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trif-
luoromethylbenzonitrile (2d) (0.02 g, 0.05 mmol), methanesulfonyl
chloride (0.009 g, 0.075 mmol) and pyridine (0.006 g, 0.075 mmol)
in dichloromethane (1 ml) was stirred at room temperature for 15
hours. The medium was washed with water (2 ml) and extracted with
ethyl acetate (5 ml). The organic layer was dried over MgSO.sub.4,
concentrated and chromatographed (HPLC, alumina column) to yield
N-{4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imi-
dazolidin-1-yl]phenyl}methanesulfonamide (47a) [RD150] (0.009 g,
0.018 mmol, 36%), the structure of which is illustrated in Formula
2, as a white powder.
##STR00083##
[0263] .sup.1H NMR (DMSO-d.sub.6, 400 MHz) .delta. 1.46 (s, 6H),
3.07 (s, 3H), 7.32 (s, 4H), 8.05 (dd, J=8.2, 1.2 Hz, 1H), 8.26 (d,
J=1.2 Hz, 1H), 8.35 (d, J=8.2 Hz, 1H), 10.08 (bs, 1H); .sup.13C NMR
(DMSO-d.sub.6, 100 MHz) .delta. 23.3, 40.4, 66.7, 109.0, 115.5,
119.9, 122.6 (q, J=272.2 Hz), 128.5 (q, J=4.7 Hz), 130.8, 131.2,
131.5 (q, J=32.3 Hz), 134.5, 136.6, 138.6, 139.5, 175.4, 180.4.
Example 48
N-{4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imid-
azolidin-1-yl]phenyl}acetamide, 48a, [RD151]
[0264] A mixture of
4-[3-(4-aminophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-trif-
luoromethylbenzonitrile (2d) [RD9] (0.008 g, 0.02 mmol), acetyl
chloride (0.004 g, 0.03 mmol) and triethylamine (0.003 g, 0.03
mmol) in dichloromethane (1 ml) was stirred at 0.degree. C. for 2
hours. The mixture was concentrated and chromatographed
(dichloromethane:acetone, 90:10) to yield
N-{4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imi-
dazolidin-1-yl]phenyl}acetamide, 48a, [RD151] (0.007 g, 0.016 mmol,
80%), the structure of which is illustrated in Formula 3, as a
white powder.
##STR00084##
[0265] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.58 (s, 6H), 2.21
(s, 3H), 7.24 (d, J=8.6 Hz, 2H), 7.48 (bs, 1H), 7.69 (d, J=8.6 Hz,
2H), 7.83 (dd, J=8.2, 1.9 Hz, 1H), 7.96 (d, J=1.2 Hz, 1H), 7.97 (d,
J=8.2 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 23.6,
53.4, 66.4, 110.0, 114.8, 120.7, 122.6 (q, J=272.2 Hz), 127.1 (q,
J=4.7 Hz), 129.1, 130.2, 132.2, 133.5 (q, J=32.3 Hz), 135.2, 137.1,
139.2, 168.1, 175.0, 180.0.
Example 49
[0266] Concentrated sulfuric acid was slowly added to a mixture of
4-aminobenzoic acid (4 g, 29.2 mmol) in methanol cooled to
0.degree. C. After the addition, the mixture was stirred at room
temperature for 5 hours. The mixture was washed with a saturated
solution of sodium bicarbonate and extracted with ethyl acetate.
The organic layer was dried over MgSO.sub.4 and concentrated under
vacuum to obtain 4-aminobenzoic acid methyl ester (49a) (4.22 g,
27.9 mmol, 96%) as an off-white solid.
[0267] A mixture of 4-aminobenzoic acid methyl ester (0.32 g, 2.12
mmol), acetonecyanohydrin (3 ml) and sodium sulfate (1 g) was
refluxed for 15 hours. After filtration to remove the sodium
sulfate, the filtrate was washed with brine and extracted with
ethyl acetate. The organic layer was concentrated and
chromatographed (dichloromethane:acetone, 60:40) to yield
4-[(cyanodimethylmethyl)-amino]-benzoic acid methyl ester (49b)
(0.398 g, 1.95 mmol, 92%) as a white solid.
49-1) RD152
[0268] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.228 g, 1 mmol) and 4-[(cyanodimethylmethyl)-amino]-benzoic
acid methyl ester (49b) (0.14 g, 0.64 mmol) in DMF (2 ml) was
heated under microwave irradiation at 60.degree. C. for 12 hours.
To this mixture were added methanol (6 ml) and aq. 2N HCl (2 ml).
The second mixture was refluxed for 4 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (10
ml) and extracted with ethyl acetate (3.times.30 ml). The organic
layer was dried over MgSO.sub.4, concentrated and chromatographed
(dichloromethane; dichloromethane:acetone, 75:25) to yield
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imidaz-
olidin-1-yl]benzoic acid methyl ester (49c) [RD152] (0.18 g, 0.4
mmol, 63%), the structure of which is illustrated in Formula 19, as
a white powder.
##STR00085##
[0269] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.60 (s, 6H), 3.95
(s, 3H), 7.40 (d, J=8.6 Hz, 2H), 7.84 (dd, J=8.2, 1.9 Hz, 1H), 7.96
(d, J=1.2 Hz, 1H), 7.97 (d, J=8.2 Hz, 1H), 8.21 (d, J=8.6 Hz, 2H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 23.8, 52.6, 66.6, 110.3,
114.8, 121.9 (q, J=272.7 Hz), 127.1 (q, J=4.7 Hz), 129.8, 131.2,
131.4, 132.2, 133.5 (q, J=32.3 Hz), 135.3, 137.0, 139.2, 165.9,
174.7, 179.7.
49-2) RD153
[0270] A mixture of
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imidaz-
olidin-1-yl]benzoic acid methyl ester (49c) (0.02 g, 0.0435 mmol)
and methylamine (2 ml distilled from its 40% aqueous solution) was
kept at -20.degree. C. for 15 hours. After evaporation of the
methylamine, the mixture was chromatographed
(dichloromethane:acetone, 80:20) to yield
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imidaz-
olidin-1-yl]-N-methylbenzamide (49d) [RD153] (0.01 g, 0.0224, 51%),
the structure of which is illustrated in Formula 20. The ester
4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2-thioxo-imidaz-
olidin-1-yl]benzoic acid methyl ester (49c) (0.08 g, 0.0179 mmol,
41%) was also recovered.
##STR00086##
[0271] .sup.1H NMR (Acetone-d.sub.6, 400 MHz) .delta. 1.60 (s, 6H),
2.90 (d, J=4.6 Hz, 3H), 7.48 (d, J=8.6 Hz, 2H), 7.80 (bs, 1H), 7.99
(d, J=8.6 Hz, 2H), 8.06 (dd, J=8.2, 1.8 Hz, 1H), 8.18 (d, J=1.8 Hz,
1H), 8.25 (d, J=8.2 Hz, 1H); .sup.13C NMR (Acetone-d.sub.6, 100
MHz) .delta. 23.8, 54.0, 66.5, 110.3, 114.8, 121.9 (q, J=272.7 Hz),
127.1 (q, J=4.7 Hz), 128.2, 129.9, 133.5 (q, J=32.3 Hz), 135.7,
135.8, 138.2, 138.3, 139.2, 166.0, 174.9, 179.7.
Example 50
50-1) RD154
[0272] A mixture of
4-[8-(4-hydroxymethylphenyl)-5-oxo-7-thioxo-6-azaspiro[3.4]oct-6-yl]-2-tr-
ifluoromethyl-benzonitrile (36b) (0.086 g, 0.2 mmol) and
methanesulfonyl anhydride (0.07 g, 0.4 mmol) in dichloromethane (1
ml) was stirred at room temperature for 15 hours. The mixture was
concentrated and chromatographed (dichloromethane:acetone, 98:2) to
yield Methanesulfonic acid
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]oct-5-yl]phenylmethyl ester (50a) [RD154] (0.089 g, 0.175
mmol, 88%), the structure of which is illustrated in Formula 22, as
a white powder.
##STR00087##
[0273] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.70 (m, 1H),
2.17-2.31 (m, 1H), 2.48-2.57 (m, 2H), 2.64-2.70 (m, 2H), 3.04 (s,
3H), 5.30 (s, 2H), 7.37 (d, J=8.3 Hz, 2H), 7.62 (d, J=8.3 Hz, 2H),
7.84 (dd, J=8.3, 1.8 Hz, 1H), 7.97 (d, J=8.2 Hz, 1H), 7.98 (d,
J=1.6 Hz, 1H).
50-2) RD155
[0274] Methylamine (0.5 ml) was bubbled into a mixture of
Methanesulfonic acid
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]oct-5-yl]phenylmethyl ester (50a) (0.059 g, 0.115 mmol) in THF
(3 ml) cooled to -78.degree. C. After 1 hour of reaction at
-78.degree. C., the mixture was concentrated and chromatographed
(dichloromethane:acetone, 95:5; methanol) to yield
4-[5-(4-methylaminomethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7--
yl]-2-trifluoromethylbenzonitrile (50b) [RD155] (0.042 g, 0.095
mmol, 82%), the structure of which is illustrated in Formula 23, as
a white powder.
##STR00088##
[0275] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57-1.70 (m, 1H),
2.16-2.24 (m, 1H), 2.52 (s, 3H), 2.53-2.57 (m, 2H), 2.60-2.68 (m,
2H), 3.85 (s, 2H), 7.27 (d, J=8.3 Hz, 2H), 7.55 (d, J=8.3 Hz, 2H),
7.84 (dd, J=8.3, 1.8 Hz, 1H), 7.95 (d, J=8.2 Hz, 1H), 7.97 (d,
J=1.6 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
31.5, 36.4, 55.6, 67.4, 110.0, 114.9, 122.0 (q, J=272.5 Hz), 127.0
(q, J=4.9 Hz), 129.1, 129.6, 129.8, 132.2, 133.3 (q, J=33.3 Hz),
133.7, 135.2, 142.4, 174.8, 179.9.
50-3) RD156
[0276] A mixture of Methanesulfonic acid
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]-
oct-5-yl]phenylmethyl ester (50a) (0.02 g, 0.039 mmol) and
dimethylamine (0.5 ml; distilled from its 40% aqueous solution) in
THF (1 ml) was stirred for 2 hours at -78.degree. C. The mixture
was concentrated and chromatographed (dichloromethane:acetone,
95:5; acetone) to yield
4-[5-(4-dimethylaminomethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct--
7-yl]-2-trifluoromethylbenzonitrile (50c) [RD156] (0.017 g, 0.037
mmol, 95%), the structure of which is illustrated in Formula 24, as
a white powder.
##STR00089##
[0277] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.57-1.70 (m, 1H),
2.16-2.24 (m, 1H), 2.32 (s, 6H), 2.55-2.60 (m, 2H), 2.63-2.69 (m,
2H), 3.53 (s, 2H), 7.27 (d, J=8.3 Hz, 2H), 7.55 (d, J=8.3 Hz, 2H),
7.84 (dd, J=8.3, 1.8 Hz, 1H), 7.95 (d, J=8.2 Hz, 1H), 7.97 (d,
J=1.6 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7,
31.5, 45.5, 63.7, 67.4, 110.0, 114.9, 122.0 (q, J=272.5 Hz), 127.0
(q, J=4.9 Hz), 129.1, 129.6, 129.8, 132.2, 133.3 (q, J=33.3 Hz),
133.7, 135.2, 142.4, 174.8, 179.9.
Example 51
[0278] Sodium cyanide (0.245 g, 5 mmol) was added to a mixture of
4-aminobenzoic acid (0.274 g, 2 mmol) and cyclobutanone (0.21 g, 3
mmol) in 90% acetic acid (4.5 ml). The reaction mixture was stirred
at room temperature for 15 hours. The mixture was washed with
aqueous HCl (pH 2) and extracted with ethyl acetate. The organic
layer was dried over magnesium sulfate and concentrated to dryness
under vacuum to yield 4-(1-cyanocyclobutylamino)benzoic acid (51a)
(0.426 g, 1.97 mmol, 99%) as a white solid.
51-1) RD159 and RD160
[0279] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.51 g, 2.22 mmol) and 4-(1-cyanocyclobutylamino)benzoic acid
(51a) (0.343 g, 1.59 mmol) in DMF (2 ml) was heated under microwave
irradiation at 60.degree. C. and stirred for 16 hours. To this
mixture were added methanol (10 ml) and aq. 2M HCl (5 ml). The
second mixture was refluxed for 12 hours. After being cooled to
room temperature, the reaction mixture was poured into cold water
(20 ml) and extracted with ethyl acetate (3.times.30 ml). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 95:5) to yield
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]-
oct-5-yl]-benzoic acid methyl ester (51b) [RD159] (0.09 g, 0.196
mmol, 12%), the structure of which is illustrated in Formula 25, as
a white powder and
N-(3-cyano-4-trifluoromethylphenyl)-4-[7-(4-cyano-3-trifluoromethylphenyl-
)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-5-yl]benzamide (51b')
[RD160] (0.28 g, 0.45 mmol, 29%), the structure of which is
illustrated in Formula 26, as a white powder.
##STR00090##
[0280] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.67-1.71 (m, 1H),
2.20-2.26 (m, 1H), 2.49-2.57 (m, 2H), 2.66-2.73 (m, 2H), 3.96 (s,
3H), 7.42 (d, J=8.4 Hz, 2H), 7.85 (dd, J=8.3, 1.7 Hz, 1H), 7.97 (d,
J=8.3 Hz, 1H), 7.98 (d, J=1.7 Hz, 1H), 8.26 (d, J=8.3 Hz, 2H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.7, 31.6, 52.6, 67.5,
110.1, 114.8, 121.8 (q, J=272.7 Hz), 127.0 (q, J=4.7 Hz), 130.2,
131.4, 131.5, 132.2, 133.4 (q, J=33.2 Hz), 135.2, 137.0, 139.2,
165.9, 174.6, 179.7.
##STR00091##
[0281] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.67-1.71 (m, 1H),
2.18-2.26 (m, 1H), 2.50-2.58 (m, 2H), 2.68-2.74 (m, 2H), 7.47 (d,
J=8.5 Hz, 2H), 7.83 (d, J=8.7 Hz, 1H), 7.84 (dd, J=8.3, 1.9 Hz,
1H), 7.96 (d, J=8.0 Hz, 1H), 9.97 (d, J=1.9 Hz, 1H), 8.10-8.14 (m,
3H), 8.21 (d, J=1.9 Hz, 1H), 8.88, (s, 1H).
51-2) RD161
[0282] A mixture of
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]-
oct-5-yl]-benzoic acid methyl ester (51b) (0.046 g, 0.1 mmol) and
methylamine (1 ml distilled from its 40% aqueous solution) was kept
at -20.degree. C. for 15 hours. After evaporation of the
methylamine, the mixture was chromatographed
(dichloromethane:acetone, 80:20) to yield
N-methyl-4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diazas-
piro[3.4]oct-5-yl]benzamide (51c) [RD161] (0.041 g, 0.085, 84%),
the structure of which is illustrated in Formula 27.
##STR00092##
[0283] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.63-1.70 (m, 1H),
2.18-2.26 (m, 1H), 2.48-2.56 (m, 2H), 2.65-2.71 (m, 2H), 3.05 (d,
J=4.8 Hz, 3H), 6.32 (bs, 1H), 7.39 (d, J=8.3 Hz, 2H), 7.84 (dd,
J=8.3, 1.7 Hz, 1H), 7.95-7.98 (m, 4H); .sup.13C NMR (CDCl.sub.3,
100 MHz) .delta. 13.6, 27.0, 31.6, 67.4, 110.3, 114.8, 121.8 (q,
J=272.7 Hz), 127.0 (q, J=4.7 Hz), 128.7, 130.3, 132.1, 133.3 (q,
J=33.2 Hz), 135.2, 136.3, 137.0, 137.8, 167.2, 174.6, 179.8.
Example 52 [RD162]
[0284] Thionyl chloride (2.38 g, 20 mmol) was added slowly to a
solution of 2-fluoro-4-nitrobenzoic acid (2.97 g, 16 mmol) in DMF
(50 ml) cooled at -5.degree. C. The mixture was stirred for an
additional 1 hour at -5.degree. C. Methylamine (0.62 g, 20 mmol;
freshly distilled from its 40% aqueous solution) was added to the
reaction medium. The second mixture was stirred for an additional 1
hour. Ethyl acetate (300 ml) was added to the mixture, which was
washed with brine (3.times.150 ml). The organic layer was dried
over MgSO.sub.4, and concentrated to yield
N-methyl-2-fluoro-4-nitrobenzamide (52a) (2.89 g, 14.6 mmol, 91%)
as a yellow solid. .sup.1H NMR (Acetone d6, 400 MHz) .delta. 3.05
(d, J=4.3 Hz, 3H), 6.31 (dd, J=13.5, 2.1 Hz, 1H), 6.40 (dd, J=8.5,
2.1 Hz, 1H), 7.64 (dd, J=8.6, 8.6 Hz, 1H).
[0285] A mixture of N-methyl-2-fluoro-4-nitrobenzamide (52a) (2.89
g, 14.6 mmol) and iron (5.04 g, 90 mmol) in ethyl acetate (40 ml)
and acetic acid (40 ml) was refluxed for 1 hour. The solid
particles were filtered off. The filtrate was washed with water and
extracted with ethyl acetate. The organic layer was dried over
MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 95:5) to yield
N-methyl-2-fluoro-4-aminobenzamide (52b) (2.3 g, 13.7 mmol, 94%) as
an off-white solid. .sup.1H NMR (acetone-d.sub.6, 400 MHz) .delta.
2.86 (d, J=4.3 Hz, 3H), 5.50 (bs, 2H), 6.37 (dd, J.sub.1=14.7 Hz,
J.sub.2=2.1 Hz, 1H), 6.50 (dd, J=8.5, 2.1 Hz, 1H), 7.06 (bs, 1H),
7.68 (dd, J=8.8 8.8 Hz, 1H); .sup.13C NMR (acetone-d.sub.6, 100
MHz) .delta. 25.8, 99.6 (d, J=13.8 Hz), 109.2 (d, J=12.8 Hz), 110.0
(d, J=1.6 Hz), 132.5 (d, J=4.8 Hz), 153.5 (d, J=12.6 Hz), 162.2 (d,
J=242.5 Hz), 164.0 (d, J=3.1 Hz).
[0286] Sodium cyanide (1.47 g, 30 mmol) was added to a mixture of
N-methyl-2-fluoro-4-aminobenzamide (52b) (1.68 g, 10 mmol) and
cyclobutanone (1.4 g, 20 mmol) in 90% acetic acid (20 ml). The
reaction mixture was stirred at 80.degree. C. for 24 hours. The
mixture was washed with water and extracted with ethyl acetate. The
organic layer was dried over magnesium sulfate and concentrated to
dryness under vacuum. The solid was washed with a 50:50 mixture of
ethyl ether and hexane (10 ml) to remove cyclobutanone cyanohydrin
to afford after filtration
N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide (52c) (2.19
g, 8.87 mmol, 89%). .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta.
1.87-1.95 (m, 1H), 2.16-2.27 (m, 1H), 2.35-2.41 (m, 2H), 2.76-2.83
(m, 2H), 2.97 (d, J=4.4 Hz, 3H), 4.68 (bs, 1H), 6.29 (dd, J=14.3,
1.8 Hz, 1H), 6.48 (dd, J=8.3, 1.8 Hz, 1H), 6.75 (q, J=4.4 Hz, 1H),
7.90 (dd, J=8.3, 8.3 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz)
.delta. 15.7, 26.7, 33.9, 49.4, 100.2 (d, J=29.5 Hz), 110.6, 111.0
(d, J=11.8 Hz), 133.1 (d, J=4.2 Hz), 148.4 (d, J=12.0 Hz), 162.0
(d, J=244.1 Hz), 164.4 (d, J=3.6 Hz).
[0287] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (2.16 g, 9.47 mmol) and
N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide (52c) (1.303
g, 5.27 mmol) in DMF (20 ml) was heated under microwave irradiation
at 80.degree. C. for 16 hours. To this mixture was added methanol
(50 ml) and aq. 2N HCl (20 ml). The second mixture was refluxed for
3 hours. After being cooled to room temperature, the reaction
mixture was poured into cold water (100 ml) and extracted with
ethyl acetate (150 ml). The organic layer was dried over
MgSO.sub.4, concentrated and chromatographed
(dichloromethane:acetone, 95:5) to yield
N-methyl-4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza--
spiro[3.4]oct-5-yl]-2-fluorobenzamide (52d) [RD162] (1.43 g, 3.0
mmol, 57%), the structure of which is illustrated in Formula 28, as
a yellow powder.
##STR00093##
[0288] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.65-1.75 (m, 1H),
2.18-2.30 (m, 1H), 2.49-2.57 (m, 2H), 2.67-2.73 (m, 2H), 3.07 (d,
J=4.4 Hz, 3H), 6.75 (q, J=4.6 Hz, 1H), 7.17 (dd, J=11.5, 1.9 Hz,
1H), 7.26 (dd, J=8.3, 1.9 Hz, 1H), 7.83 (dd, J=8.2, 2.0 Hz, 1H),
7.95 (d, J=1.8 Hz, 1H), 7.97 (d, J=8.3 Hz, 1H) 8.30 (dd, J=8.3, 8.3
Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.6, 27.0,
31.7, 67.4, 110.3, 114.8, 118.2, 118.5, 121.9 (q, J=272.7 Hz),
126.6, 127.0 (q, J=4.8 Hz), 132.1, 133.3 (q, J=33.2 Hz), 133.8,
135.3, 136.8, 139.1 (d, J=10.9 Hz), 160.5 (d, J=249.1 Hz), 162.7
(d, J=3.3 Hz), 174.3, 179.8; .sup.19F NMR (CDCl.sub.3, 100 MHz)
.delta. -111.13, -62.58.
Example 53 [RD163]
[0289] A mixture of 4-nitro-3-fluorophenol (0.314 g, 2 mmol) and
iron (0.56 g, 10 mmol) in ethyl acetate (4 ml) and acetic acid (2
ml) was refluxed for 3 hour. The solid particles were filtered off.
The filtrate was washed with water and extracted with ethyl
acetate. The organic layer was dried over MgSO.sub.4, concentrated
to yield 4-amino-3-fluorophenol (53a) (0.25 g, 19.6 mmol, 98%) as a
brown solid. .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 6.48-6.58
(m, 2H), 6.61-6.70 (m, 1H), 7.87 (bs, 3H).
[0290] Sodium cyanide (0.194 g, 4 mmol) was added to a mixture of
4-amino-3-fluorophenol (0.29 g, 2.28 mmol) and cyclobutanone (0.175
g, 2.5 mmol) in 90% acetic acid (3 ml). The reaction mixture was
stirred at room temperature for 15 hours. The medium was washed
with water and extracted with ethyl acetate. The organic layer was
dried over magnesium sulfate, concentrated and chromatographed
(dichloromethane:acetone, 90:10) to yield
1-(2-fluoro-4-hydroxyphenylamino)-cyclobutanecarbonitrile (53b)
(0.271 g, 1.31 mmol, 58%) as an off-white solid. .sup.1H NMR
(CDCl.sub.3, 400 MHz) .delta. 2.13-2.20 (m, 2H), 2.36-2.41 (m, 2H),
2.70-2.75 (m, 2H), 4.00 (bs, 1H), 6.46 (bs, 1H), 6.52 (ddd,
J.sub.1=2.2 Hz, J.sub.2=0.65 Hz, J.sub.3=0.22 Hz, 1H), 6.57 (d,
J=2.3 Hz), 6.62 (dd, J.sub.1=3.0 Hz, J.sub.2=0.67 Hz, 1H); NMR
(CDCl.sub.3, 100 MHz) .delta. 15.7, 34.1, 50.9, 104.0 (d, J=21.9
Hz), 111.0 (d, J=3.4 Hz), 115.8 (d, J=3.7 Hz), 121.8, 125.3 (d,
J=12.3 Hz), 150.1 (d, J=10.4 Hz), 152.8 (d, J=239.3 Hz).
[0291] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.228 g, 1.0 mmol) and
1-(2-fluoro-4-hydroxyphenylamino)-cyclobutanecarbonitrile (53b)
(0.145 g, 0.7 mmol) in dry DMF (2 ml) was stirred at room
temperature for 24 hours. To this mixture were added methanol (10
ml) and aq. 2M HCl (2 ml). The second mixture was refluxed for 1
hour. After being cooled to room temperature, the reaction mixture
was poured into cold water (10 ml) and extracted with ethyl acetate
(50 ml). The organic layer was dried over MgSO.sub.4, concentrated
and chromatographed (dichloromethane pure and then
dichloromethane:acetone, 90:10) to yield
4-[5-(2-fluoro-4-hydroxyphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-y-
l]-2-trifluoromethylbenzonitrile (53c) [RD163] (0.17 g, 0.39 mmol,
56%), the structure of which is illustrated in Formula 29, as a
off-white powder.
##STR00094##
[0292] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.66-1.75 (m, 1H),
2.18-2.28 (m, 1H), 2.42-2.50 (m, 1H), 2.54-2.67 (m, 3H), 6.76 (d,
J=2.2 Hz, 2H), 7.15 (t, J=2.1 Hz, 1H), 7.35 (bs, 1H), 7.87 (dd,
J.sub.1=8.2 Hz, J.sub.2=1.8 Hz, 1H), 7.97 (d, J=8.2 Hz, 1H), 7.98
(d, J=1.8 Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 13.8,
31.0, 67.6, 104.8 (d, J=22.3 Hz), 109.8, 112.6, 114.4 (d, J=13.1
Hz), 114.9, 121.9 (q, J=272.8 Hz), 127.1 (q, J=4.8 Hz), 132.0,
132.3, 133.5 (q, J=33.3 Hz), 135.3, 137.2, 159.3 (d, J=11.2 Hz),
159.6 (d, J=249.7 Hz), 175.2, 180.5; .sup.19F NMR (CDCl.sub.3, 100
MHz) .delta. -117.5, -62.49.
Example 54 [RD168]
[0293] A mixture of 4-nitro-2-fluorobenzonitrile (1.83 g, 5 mmol)
and iron (1.68 g, 6 mmol) in a mixture of acetic acid (40 ml) and
ethyl acetate (40 ml) was refluxed for 2 hours. The solid was
filtered off and the filtrate was washed with water and extracted
with ethyl acetate. The organic layer was dried over magnesium
sulfate, concentrated and chromatographed (dichloromethane:acetone,
95:5) to yield 4-amino-2-fluorobenzonitrile (54a) (0.653 g, 4.8
mmol, 96%).
[0294] Sodium cyanide (0.74 g, 15 mmol) was added to a mixture of
4-amino-2-fluorobenzonitrile (1.36 g, 10 mmol) and cyclopentanone
(1.26 g, 15 mmol) in 90% acetic acid (10 ml). The reaction mixture
was stirred at room temperature for 3 hours and then the medium was
hearted to 80.degree. C. and stirred for an additional 5 hours. The
medium was washed with water and extracted with ethyl acetate. The
organic layer was dried over magnesium sulfate, concentrated and
chromatographed (dichloromethane:acetone, 97:3) to yield
4-(1-cyanocyclopentylamino)-2-fluorobenzonitrile (54b) (2.07 g,
9.03 mmol, 90%) as a yellow solid. .sup.1H NMR (CDCl.sub.3, 400
MHz) .delta. 1.69-1.91 (m, 4H), 2.13-2.18 (m, 2H), 2.37-2.42 (m,
2H), 5.08 (bs, 1H), 6.54-6.62 (m, 2H), 7.39 (t, J=7.3 Hz, 1H);
.sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 23.7, 39.8, 56.8, 89.6
(d, J=15.8 Hz), 101.2 (d, J=23.8 Hz), 110.9, 115.2, 120.8, 134.1
(d, J=2.4 Hz), 150.3 (d, J=11.2 Hz), 164.5 (d, J=254.1 Hz).
[0295] A mixture of 4-isothiocyanato-2-trifluoromethylbenzonitrile
(1a) (0.171 g, 0.75 mmol) and
4-(1-cyanocyclopentylamino)-2-fluorobenzonitrile (54b) (0.115 g,
0.5 mmol) in dry DMF (1 ml) was heated under microwave irradiation
at 60.degree. C. for 48 hours. To this mixture were added methanol
(3 ml) and aq 2M HCl (2 ml). The second mixture was refluxed for 1
hour. After being cooled to room temperature, the reaction mixture
was poured into cold water (10 ml) and extracted with ethyl acetate
(15 ml). The organic layer was dried over MgSO.sub.4, concentrated
and chromatographed (dichloromethane:acetone, 98:2) to yield
4-[1-(4-cyano-3-fluorophenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl]-
-2-trifluoromethylbenzonitrile (54c) [RD168] (0.017 g, 0.037 mmol,
7%), of which the structure is illustrated in Formula 30, as an
off-white powder.
##STR00095##
[0296] .sup.1H NMR (CDCl.sub.3, 400 MHz) .delta. 1.53-1.63 (m, 2H),
1.89-2.00 (m, 2H), 2.09-2.16 (m, 2H), 2.35-2.42 (m, 2H), 7.27-7.37
(m, 2H), 7.78-7.90 (m, 3H), 7.95 (d, J=1.8 Hz, 1H), 7.97 (d, J=8.3
Hz, 1H); .sup.13C NMR (CDCl.sub.3, 100 MHz) .delta. 25.2, 36.5,
75.3, 103.2 (d, J=15.3 Hz), 110.4, 112.8, 114.7, 119.2 (d, J=20.7
Hz), 121.9 (q, J=272.8 Hz), 127.0 (q, J=4.8 Hz), 132.1, 133.7 (q,
J=33.2 Hz), 134.6, 135.3, 135.8, 136.8, 141.8 (d, J=9.5 Hz), 163.4
(d, J=261.5 Hz), 175.3, 180.1.
Example 55 [RD136 and RD142]
[0297] Additional diarylhydantoin compounds can be synthesized,
including the following compounds illustrated in Formulas 35 and
36.
##STR00096##
Example 56 [RD162']
[0298] In the following, air or moisture sensitive reactions were
conducted under argon atmosphere using oven-dried glassware and
standard syringe/septa techniques. The reactions were monitored
with a SiO.sub.2 TLC plate under UV light (254 nm) followed by
visualization with a p-anisaldehyde or ninhydrin staining solution.
Column chromatography was performed on silica gel 60. .sup.1H NMR
spectra were measured at 400 MHz in CDCl.sub.3 unless stated
otherwise and data were reported as follows in ppm (.delta.) from
the internal standard (TMS, 0.0 ppm): chemical shift (multiplicity,
integration, coupling constant in Hz.).
##STR00097##
[0299] Periodic acid (1.69 g, 7.41 mmol) was dissolved in
acetonitrile (25 mL) by vigorous stirring, and then chromium
trioxide (0.16 g, 1.60 mmol) was dissolved into the solution.
2-Fluoro-4-nitrotoluene (0.33 g, 2.13 mmol) was added to the above
solution with stirring. A white precipitate formed immediately with
exothermic reaction. After 1 h of stirring, the supernatant liquid
of the reaction mixture was decanted to a flask, and the solvent
was removed by evaporation. The residues were extracted with
methylene chloride (2.times.30 mL) and water (2.times.30 mL). The
organic layer was dried over MgSO.sub.4, and concentrated to give
2-Fluoro-4-nitrobenzoic acid (Formula 37) (0.32 mg, 81%) as a white
solid. .sup.1H NMR .delta. 8.06 (ddd, 1H, J=9.9, 2.2 and 0.3), 8.13
(ddd, 1H, J=8.6, 2.2 and 0.9), 8.25 (ddd, 1H, J=8.6, 7.0 and
0.3).
##STR00098##
[0300] Thionyl chloride (0.15 g, 1.30 mmol) was added slowly to a
solution of 2-fluoro-4-nitrobenzoic acid (Formula 37) (0.20 g, 1.10
mmol) in DMF (5 mL) cooled at -5.degree. C. The mixture was stirred
for an additional 1 hour at -5.degree. C. Excess methylamine
(freshly distilled from its 40% aqueous solution) was added to the
reaction medium. The second mixture was stirred for an additional 1
hour. Ethyl acetate (50 mL) was added to the mixture, which was
washed with brine (2.times.50 ml). The organic layer was dried over
MgSO.sub.4, and concentrated to yield
N-Methyl-2-fluoro-4-nitrobenzamide (Formula 38) (0.18 g, 85%) as a
yellowish solid. .sup.1H NMR (acetone-d.sub.6) .delta. 3.05 (d, 3H,
J=4.3), 6.31 (dd, 1H, J=13.5 and 2.1), 6.40 (dd, 1H, J=8.6 and
2.1), 7.64 (dd, 1H, J=8.6 and 8.6).
##STR00099##
[0301] A mixture of N-Methyl-2-fluoro-4-nitrobenzamide (Formula 38)
(0.18 g, 0.91 mmol) and iron (0.31 g, 5.60 mmol) in ethyl acetate
(5 mL) and acetic acid (5 mL) was refluxed for 1 h. The solid
particles were filtered off. The filtrate was washed with water and
extracted with ethyl acetate. The organic layer was dried over
MgSO.sub.4, concentrated and the residue was purified with
SiO.sub.2 column chromatography (dichloromethane:acetone, 95:5) to
give N-Methyl-2-fluoro-4-aminobenzamide (Formula 39) (0.14 g, 92%)
as an off-white solid. .sup.1H NMR (acetone-d.sub.6) .delta. 2.86
(d, 3H, J=4.3), 5.50 (br s, 2H), 6.37 (dd, 1H, J=14.7 and 2.1),
6.50 (dd, 1H, J=8.6 and 2.1), 7.06 (br s, 1H), 7.68 (dd, 1H, J=8.8
and 8.8).
##STR00100##
[0302] A mixture of N-Methyl-2-fluoro-4-aminobenzamide (Formula 39)
(96 mg, 0.57 mmol), acetone cyanohydrin (0.3 mL, 3.14 mmol) and
magnesium sulfate (50 mg) was heated to 80.degree. C. and stirred
for 12 h. To the medium was added ethyl acetate (25 mL) and then
washed with water (2.times.25 mL). The organic layer was dried over
MgSO.sub.4 and concentrated and the residue was purified with
SiO.sub.2 column chromatography (dichloromethane:acetone, 95:5) to
give N-Methyl-2-fluoro-4-(1,1-dimethyl-cyanomethyl)-aminobenzamide
(Formula 40) (101 mg, 75%) as a white solid. .sup.1H NMR .delta.
1.74 (s, 6H), 2.98 (dd, 3H, J=4.8 and 1.1), 6.58 (dd, 1H, J=14.6
and 2.3), 6.63 (dd, 1H, J=8.7 and 2.3), 6.66 (br s, 1H), 7.94 (dd,
1H, J=8.7 and 8.7).
##STR00101##
[0303] 4-Amino-2-trifluoromethylbenzonitrile (2.23 g, 12 mmol) was
added portionwise over 15 min into a well-stirred heterogeneous
mixture of thiophosgene (1 mL, 13 mmol) in water (22 mL) at room
temperature. Stirring was continued for an additional 1 h. The
reaction medium was extracted with chloroform (3.times.15 ml). The
combined organic phase was dried over MgSO.sub.4 and evaporated to
dryness under reduced pressure to yield desired product
4-Isothiocyanato-2-trifluoromethylbenzonitrile (Formula 41) as
brownish solid and was used as such for the next step (2.72 g, 11.9
mmol, 99%). .sup.1H NMR .delta. 7.49 (dd, 1H, J=8.3 and 2.1), 7.59
(d, 1H, J=2.1), 7.84 (d, 1H, J=8.3).
##STR00102##
56-1) RD162'
[0304] A mixture of
N-Methyl-2-fluoro-4-(1,1-dimethyl-cyanomethyl)-aminobenzamide
(Formula 40) (30 mg, 0.13 mmol) and
4-Isothiocyanato-2-trifluoromethylbenzonitrile (Formula 41) (58 mg,
0.26 mmol) in DMF (1 mL) was heated under microwave irradiation at
100.degree. C. for 11 hours. To this mixture was added methanol (20
mL) and aq. 1 N HCl (5 mL). The second mixture was refluxed for 1.5
h. After being cooled to room temperature, the reaction mixture was
poured into cold water (50 mL) and extracted with ethyl acetate (50
mL). The organic layer was dried over MgSO.sub.4, concentrated and
the residue was purified with SiO.sub.2 column chromatography
(dichloromethane:acetone, 95:5) to give RD162' (Formula 42) (15 mg,
25%) as a colorless crystal. .sup.1H NMR .delta. 1.61 (s, 6H), 3.07
(d, 3H, J=4.1), 6.71 (m, 1H), 7.15 (dd, 1H, J=11.7 and 2.0), 7.24
(dd, 1H, J=8.4 and 2.0), 7.83 (dd, 1H, J=8.2 and 2.1), 7.95 (d, 1H,
J=2.1), 7.99 (d, 1H, J=8.2), 8.28 (dd, 1H, J=8.4 and 8.4).
Example 57
##STR00103##
[0306] A mixture of N-Methyl-2-fluoro-4-aminobenzamide (Formula 39)
(62 mg, 0.37 mmol), cyclopentanone (0.07 mL, 0.74 mmol) and TMSCN
(0.1 mL, 0.74 mmol) was heated to 80.degree. C. and stirred for 13
h. To the medium was added ethyl acetate (2.times.20 mL) and then
washed with water (2.times.20 mL). The organic layer was dried over
MgSO.sub.4 and concentrated and the residue was purified with
silica gel column chromatography (dichloromethane:acetone, 95:5) to
give N-Methyl 2-fluoro-4-(1-cyanocyclopentyl)aminobenzamide
(Formula 43) (61 mg, 63%) as a white solid. .sup.1H NMR .delta.
7.95 (dd, 1H, J=8.8, 8.8 Hz), 6.65 (br s, 1H), 6.59 (dd, 1H, J=8.8,
2.3 Hz), 6.50 (dd, 1H, J=14.6, 2.3 Hz), 4.60 (br s, 1H), 2.99 (dd,
3H, J=4.8, 1.1 Hz), 2.36-2.45 (m, 2H), 2.10-2.18 (m, 2H), 1.82-1.95
(m, 4H).
##STR00104##
57-1) RD162''
[0307] A mixture of N-Methyl
2-fluoro-4-(1-cyanocyclopentyl)aminobenzamide (Formula 43) (57 mg,
0.22 mmol) and 4-isothiocyanato-2-trifluoromethyl benzonitrile
(0.15 g, 0.65 mmol) in DMF (3 mL) was heated under microwave
irradiation (open vessel) at 130.degree. C. for 12 hours. To this
mixture was added methanol (20 mL) and aq. 1 N HCl (5 mL). The
second mixture was refluxed for 1.5 h. After being cooled to room
temperature, the reaction mixture was poured into cold water (50
mL) and extracted with ethyl acetate (50 mL). The organic layer was
dried over MgSO.sub.4, concentrated and the residue was purified
with silica gel column chromatography (dichloromethane:acetone,
95:5) to give
4-(3-(4-Cyano-3-(trifluoromethyl)phenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.-
4]nonan-1-yl)-2-fluoro-N-methylbenzamide, RD162'' (Formula 44) (8
mg, 7%) as a pale yellowish solid. .sup.1H NMR .delta. 8.28 (dd,
1H, J=8.4, 8.4 Hz), 7.98 (d, 1H, J=8.3 Hz), 7.96 (d, 1H, J=1.8 Hz),
7.84 (dd, 1H, J=8.3, 1.8 Hz), 7.27 (dd, 1H, J=8.4, 1.8 Hz), 7.17
(dd, 1H, J=11.7, 1.8 Hz), 6.67-6.77 (m, 1H), 3.07 (d, 3H, J=4.3
Hz), 2.32-2.41 (m, 2H), 2.13-2.21 (m, 2H), 1.85-1.96 (m, 2H),
1.49-1.59 (m, 2H).
Example 58
##STR00105##
[0309] Trifluoroacetic anhydride (0.85 mL, 6.14 mmol) was added to
a solution of 4-(4-aminophenyl)butyric acid (0.5 g, 2.79 mmol) in
chloroform (10 mL) at 0.degree. C. The mixture was warmed to room
temperature and stirred for 3 hours. The mixture was partitioned
with chloroform (20 mL) and water (20 mL). The organic layer was
dried over MgSO.sub.4, concentrated and the residue was purified
with silica gel column chromatography (dichloromethane:acetone,
9:1) to give 4-[4-(2,2,2-Trifluoroacetylamino)phenyl]butanoic acid
(Formula 45) (0.53 g, 69%). .sup.1H NMR .delta. 7.81 (br s, 1H),
7.48 (d, 2H, J=8.5 Hz), 7.22 (d, 2H, J=8.5 Hz), 2.68 (t, 2H, J=7.5
Hz), 2.38 (t, 2H, J=7.5 Hz), 1.96 (p, 2H, J=7.5 Hz).
##STR00106##
[0310] Thionyl chloride (71 mg, 0.60 mmol) was added slowly to a
solution of 4-[4-(2,2,2-Trifluoroacetylamino)phenyl]butanoic acid
(Formula 45) (0.15 g, 0.55 mmol) in DMF (5 mL) cooled at -5.degree.
C. The mixture was stirred for an additional 1 hour at -5.degree.
C. Excess dimethylamine (freshly distilled from its 40% aqueous
solution) was added to the reaction medium. The second mixture was
stirred for an additional 1 hour. Ethyl acetate (50 mL) was added
to the mixture, which was washed with brine (2.times.50 ml). The
organic layer was dried over MgSO.sub.4, and concentrated to yield
N,N-Dimethyl 4-[4-(2,2,2-Trifluoroacetylamino)phenyl]butanamide
(Formula 46) (0.17 g, quant.) as a yellowish solid. .sup.1H NMR
.delta. 9.70 (br s, 1H), 7.55 (d, 2H, J=8.6 Hz), 7.11 (d, 2H, J=8.6
Hz), 2.91 (s, 3H), 2.89 (s, 3H), 2.60 (t, 2H, J=7.7 Hz), 2.27 (t,
2H, J=7.7 Hz), 1.89 (p, 2H, J=7.7 Hz).
##STR00107##
[0311] 1 N NaOH solution (3 mL) was added to a solution of
N,N-Dimethyl 4-[4-(2,2,2-Trifluoroacetylamino)phenyl]butanamide
(Formula 46) (0.17 g, 0.55 mmol) in methanol (2 mL) at room
temperature. The mixture was stirred for 14 hour. The mixture was
partitioned with chloroform (25 mL) and water (25 mL). The organic
layer was dried over MgSO.sub.4, and concentrated and the residue
was purified with silica gel column chromatography
(dichloromethane:acetone, 9:1) to give N,N-Dimethyl
4-(4-aminophenyl)butanamide (Formula 47) (74 mg, 66%) as a white
solid. .sup.1H NMR .delta. 6.97 (d, 2H, J=8.3 Hz), 6.61 (d, 2H,
J=8.3 Hz), 3.56 (br s, 2H), 2.92 (s, 6H), 2.56 (t, 2H, J=7.7 Hz),
2.28 (t, 2H, J=7.7 Hz), 1.91 (p, 2H, J=7.7 Hz).
##STR00108##
[0312] A mixture of N,N-Dimethyl 4-(4-aminophenyl)butanamide
(Formula 47) (74 mg, 0.36 mmol), cyclobutanone (54 mg, 0.78 mmol)
and TMSCN (77 mg, 0.78 mmol) was heated to 80.degree. C. and
stirred for 15 h. To the medium was added ethyl acetate (2.times.20
mL) and then washed with water (2.times.20 mL). The organic layer
was dried over MgSO.sub.4 and concentrated and the residue was
purified with silica gel column chromatography
(dichloromethane:acetone, 9:1) to give N,N-Dimethyl
4-[4-(1-cyanocyclobutylamino)phenyl]butanamide (Formula 48) (58 mg,
57%) as a white solid. .sup.1H NMR .delta. 7.07 (d, 2H, J=8.5 Hz),
6.59 (d, 2H, J=8.5 Hz), 3.94 (br s, 1H), 2.94 (s, 3H), 2.93 (s,
3H), 2.75-2.83 (m, 2H), 2.60 (t, 2H, J=7.6 Hz), 2.33-2.42 (m, 2H),
2.30 (t, 2H, J=7.6 Hz), 2.11-2.28 (m, 2H), 1.93 (p, 2H, J=7.6
Hz).
##STR00109##
[0313] A mixture of N,N-Dimethyl
4-[4-(1-cyanocyclobutylamino)phenyl]butanamide (Formula 48) (58 mg,
0.20 mmol) and 4-isothiocyanato-2-trifluoromethyl benzonitrile (74
mg, 0.32 mmol) in DMF (3 mL) was heated under reflux for 2 hours.
To this mixture was added methanol (20 mL) and aq. 1 N HCl (5 mL).
The second mixture was refluxed for 1.5 h. After being cooled to
room temperature, the reaction mixture was poured into cold water
(50 mL) and extracted with ethyl acetate (50 mL). The organic layer
was dried over MgSO.sub.4, concentrated and the residue was
purified with silica gel column chromatography
(dichloromethane:acetone, 95:5) to give
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)-N,N-dimethylbutanamide, RD169 (Formula 49)
(44 mg, 42%) as a pale yellowish solid. .sup.1H NMR .delta. 7.98
(s, 1H), 7.97 (d, 1H, J=8.2 Hz), 7.86 (d, 1H, J=8.2 Hz), 7.42 (d,
2H, J=8.3 Hz), 7.22 (d, 2H, J=8.3 Hz), 2.99 (s, 3H), 2.96 (s, 3H),
2.78 (t, 2H, J=7.5 Hz), 2.62-2.70 (m, 2H), 2.52-2.63 (m, 2H), 2.40
(t, 2H, J=7.5 Hz), 2.15-2.30 (m, 1H), 2.04 (p, 2H, J=7.5 Hz),
1.62-1.73 (m, 1H).
Example 59
##STR00110##
[0315] A mixture of 4-(4-aminophenyl)butyric acid (0.20 g, 1.12
mmol), cyclobutanone (0.17 mL, 2.23 mmol) and TMSCN (0.30 mL, 2.23
mmol) was heated to 80.degree. C. and stirred for 13 h. To the
medium was added ethyl acetate (2.times.30 mL) and then washed with
water (2.times.30 mL). The organic layer was dried over MgSO.sub.4
and concentrated and the residue was purified with silica gel
column chromatography (dichloromethane:acetone, 9:1) to give
4-[4-(1-Cyanocyclobutylamino)phenyl]butanoic acid (Formula 50)
(0.21 g, 74%) as a yellowish solid. .sup.1H NMR .delta. 7.06 (d,
2H, J=8.6 Hz), 6.59 (d, 2H, J=8.6 Hz), 2.75-2.83 (m, 2H), 2.59 (t,
2H, J=7.5 Hz), 2.37 (t, 2H, J=7.5 Hz), 2.33-2.42 (m, 2H), 2.11-2.28
(m, 2H), 1.92 (p, 2H, J=7.5 Hz).
##STR00111##
[0316] A mixture of 4-[4-(1-Cyanocyclobutylamino)phenyl]butanoic
acid (Formula 50) (0.21 g, 0.83 mmol) and
4-isothiocyanato-2-trifluoro benzonitrile (0.25 g, 1.08 mmol) in
toluene (10 mL) was heated under reflux for 1 hours. To this
mixture was added aq. 1 N HCl (5 mL). The second mixture was
refluxed for 1.5 h. After being cooled to room temperature, the
reaction mixture was poured into cold water (50 mL) and extracted
with ethyl acetate (50 mL). The organic layer was dried over
MgSO.sub.4, concentrated and the residue was purified with silica
gel column chromatography (dichloromethane:acetone, 95:5) to give
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)butanoic acid, RD141 (Formula 51) (60 mg,
15%). .sup.1H NMR .delta. 7.98 (d, 1H, J=1.8 Hz), 7.97 (d, 1H,
J=8.3 Hz), 7.86 (dd, 1H, J=8.3, 1.8 Hz), 7.42 (d, 2H, J=8.5 Hz),
7.24 (d, 2H, J=8.5 Hz), 2.79 (t, 2H, J=7.5 Hz), 2.62-2.68 (m, 2H),
2.51-2.59 (m, 2H), 2.47 (t, 2H, J=7.5 Hz), 2.14-2.26 (m, 1H), 2.06
(p, 2H, J=7.5 Hz), 1.60-1.70 (m, 1H).
Example 60
##STR00112##
[0318] To a solution of
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)butanoic acid, RD141 (Formula 51) (60 mg,
0.12 mmol) in DMF (3 mL) was added thionyl chloride (0.01 mL, 0.15
mmol) at 0.degree. C. The mixture was stirred at 0.degree. C. for 1
hour. Then ammonia was bubbled into the mixture. The mixture was
partitioned with ethyl acetate (25 mL) and water (25 mL). The
organic layer was dried over MgSO.sub.4, concentrated and
chromatographed (dichloromethane:acetone, 70:30) to yield
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)butanamide, RD130 (Formula 52) (37 mg, 61%)
as a white powder. .sup.1H NMR .delta. 7.97 (d, 1H, J=1.8 Hz), 7.95
(d, 1H, J=8.3 Hz), 7.85 (dd, 1H, J=8.3 Hz), 7.39 (d, 2H, J=8.3 Hz),
7.22 (d, 2H, J=8.3 Hz), 5.59 (br s, 2H), 2.77 (t, 2H, J=7.5 Hz),
2.62-2.68 (m, 2H), 2.51-2.59 (m, 2H), 2.31 (t, 2H, J=7.5 Hz),
2.16-2.25 (m, 1H), 2.05 (p, 2H, J=7.5 Hz), 1.57-1.70 (m, 1H).
Example 61
##STR00113##
[0320] A solution of DMSO (0.01 mL, 0.12 mmol) in dry
dichloromethane (1 mL) was added to a stirred solution of oxalyl
chloride (0.01 mL, 0.09 mmol) in dry dichloromethane (2 mL) at
-78.degree. C. After 15 min, a dichloromethane solution of
4-(4-(7-(4-Cyano-3-(trifluoromethyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro-
[3.4]octan-5-yl)phenyl)butanamide, RD130 (Formula 52) (35 mg, 0.07
mmol) was added to the reaction mixture. Stirring was continued for
20 min at -78.degree. C., and then triethylamine (0.03 mL, 0.22
mmol) was added. After 30 min at -78.degree. C., the reaction
mixture was warmed to room temperature and then reaction was
quenched with saturated aq. NH.sub.4Cl solution. The reaction
mixture was diluted with dichloromethane, and extracted with
dichloromethane. The organic layer was dried over MgSO.sub.4,
concentrated and chromatographed (dichloromethane:acetone, 95:5) to
yield
4-(5-(4-(3-Cyanopropyl)phenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]octan-7--
yl)-2-(trifluoromethyl)benzonitrile, RD170 (Formula 53) (29 mg,
87%) as a viscous oil. .sup.1H NMR .delta. 7.98 (d, 1H, J=1.8 Hz),
7.98 (d, 1H, J=8.3 Hz), 7.86 (dd, 1H, J=8.3, 1.8 Hz), 7.43 (d, 2H,
J=8.4 Hz), 7.27 (d, 2H, J=8.4 Hz), 2.90 (t, 2H, J=7.3 Hz),
2.63-2.73 (m, 2H), 2.52-2.62 (m, 2H), 2.42 (t, 2H, J=7.3 Hz),
2.18-2.30 (m, 1H), 2.07 (p, 2H, J=7.3 Hz), 1.63-1.73 (m, 1H).
[0321] One skilled in the art could modify and/or combine the
syntheses described herein to make other diarylhydantoin
compounds.
[0322] Inventive compounds also include those with the following
formulas.
##STR00114##
[0323] Where R is selected from hydrogen, aryl, substituted aryl,
alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,
substituted alkynyl, halogenated alkyl, halogenated alkenyl,
halogenated alkynyl, arylalkyl, arylalkenyl, arylalkynyl,
heterocyclic aromatic or non-aromatic, substituted heterocyclic
aromatic or non-aromatic, cycloalkyl, substituted cycloalkyl,
halogen, SO.sub.2R.sub.11, NR.sub.11R.sub.12,
NR.sub.12(CO)OR.sub.11, NH(CO)NR.sub.11R.sub.12,
NR.sub.12(CO)R.sub.11, O(CO)R.sub.11, O(CO)OR.sub.11,
O(CS)R.sub.11, NR.sub.12(CS)R.sub.11, NH(CS)NR.sub.11R.sub.12,
NR.sub.12(CS)OR.sub.11.
[0324] R.sub.1 and R.sub.2 are independently selected from
hydrogen, aryl, alkyl, substituted alkyl, alkenyl, substituted
alkenyl, alkynyl, substituted alkynyl, halogenated alkyl,
halogenated alkenyl, halogenated alkynyl, arylalkyl, arylalkenyl,
arylalkynyl, heterocylic aromaric or non-aromatic, substituted
heterocyclic aromatic or non-aromatic, cycloalkyl, substituted
cycloalkyl.
[0325] R.sub.1 and R.sub.2 can be connected to form a cycle which
can be heterocyclic, substituted heterocyclic, cycloakyl,
substituted cycloalkyl.
[0326] R.sub.3 is selected from aryl, substituted aryl, alkyl,
substituted alkyl, alkenyl, substituted alkenyl, alkynyl,
substituted alkynyl, arylalkyl, arylalkenyl, arylalkynyl,
heterocyclic aromatic or non-aromatic, substituted heterocyclic
aromatic or non-aromatic, cycloalkyl, substituted cycloalkyl,
SO.sub.2R.sub.11, NR.sub.11R.sub.12, (CO)OR.sub.11,
(CO)NR.sub.11R.sub.12, (CO)R.sub.11, (CS)R.sub.11, (CS)R.sub.11,
(CS)NR.sub.11R.sub.12, (CS)OR.sub.11.
[0327] R.sub.5 is CN or NO.sub.2 or SO.sub.2R.sub.11
[0328] R.sub.6 is CF.sub.3, alkyl, substituted alkyl, alkenyl,
substituted alkenyl, alkynyl, substituted alkynyl, halogenated
alkyl, halogenated alkenyl, halogenated alkynyl, halogen.
[0329] A is sulfur atom (S) or oxygen atom (O).
[0330] B is O or S or NR.sub.3
[0331] X is carbon or nitrogen and can be at any position in the
ring.
[0332] R.sub.11 and R.sub.12 are independently selected from
hydrogen, aryl, aralkyl, substituted aralkyl, alkyl, substituted
alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl,
halogenated alkyl, halogenated alkenyl, halogenated alkynyl,
arylalkyl, arylalkenyl, arylalkynyl, heterocyclic aromatic or
non-aromatic, substituted heterocyclic aromatic or non-aromatic,
cycloalkyl, substituted cycloalkyl.
[0333] R.sub.11 and R.sub.12 can be connected to form a cycle which
can be heterocyclic aromatic or non-aromatic, substituted
heterocyclic aromatic, cycloakyl, substituted cycloalkyl.
Pharmacological Examination of the Compounds
[0334] Compounds for which synthetic routes are described above
were identified through screening on hormone refractory prostate
cancer cells for antagonistic and agonistic activities against AR
utilizing screening procedures similar to those in PCT applications
US04/42221 and US05/05529, which are hereby incorporated by
reference. A number of compounds exhibited potent antagonistic
activities with minimal agonistic activities for over expressed AR
in hormone refractory prostate cancer.
In Vitro Biological Assay
Effect of Compounds on AR by a Reporter Assay
[0335] The compounds were subjected to tests using an artificial AR
response reporter system in a hormone refractory prostate cancer
cell line. In this system, the prostate cancer LNCaP cells were
engineered to stably express about 5-fold higher level of AR than
endogenous level. The exogenous AR has similar properties to
endogenous AR in that both are stabilized by a synthetic androgen
R1881. The AR-over expressed cells were also engineered to stably
incorporate an AR response reporter and the reporter activity of
these cells shows features of hormone refractory prostate cancer.
It responds to low concentration of a synthetic androgen R1881, is
inhibited only by high concentrations of bicalutamide (see Table
1), and displays agonistic activity with bicalutamide (FIG. 1 and
Table 2). Consistent with published data, bicalutamide inhibited AR
response reporter and did not have agonistic activity in hormone
sensitive prostate cancer cells (FIG. 2).
[0336] We examined the antagonistic activity of the compounds for
which the synthesis is described above in the presence of 100 pM of
R1881. Engineered LNCaP cells (LNCaP-AR, also abbreviated LN-AR)
were maintained in Iscove's medium containing 10% fetal bovine
serum (FBS). Two days prior to drug treatment, the cells were grown
in Iscove's medium containing 10% charcoal-stripped FBS (CS-FBS) to
deprive of androgens. The cells were split and grown in Iscove's
medium containing 10% CS-FBS with 100 pM of R1881 and increasing
concentrations of test compounds. After two days of incubation,
reporter activities were assayed.
[0337] Table 1 lists the IC50 of these compounds to inhibit AR in
hormone refractory prostate cancer. The control substance
bicalutamide has an IC50 of 889 nM. Most of the compounds
identified (diarylthiohydantoins) have IC50s between 100 to 200 nM
in inhibiting AR in hormone refractory prostate cancer. In
contrast, antiandrogenic compounds listed as examples in U.S. Pat.
No. 5,705,654, such as examples 30-2, 30-3, 31-2, 31-3, and 24-3
(RD73-RD77) have no inhibitory activities on AR in this system.
TABLE-US-00001 TABLE 1 Antagonistic activities against AR in
hormone refractory prostate cancer, measured by an AR response
reporter and by endogenous PSA expression. IC50 (nM) IC50 (nM)
Example Name Reporter PSA Bicalutamide
N-[4-cyano-3-(trifluoromethyl)phenyl]-3-[(4- 889 >1000
Comparative fluorophenyl)sulfonyl]-2-hydroxy-2- methylpropanamide
29 4-[3-(4-hydroxybutyl)-4,4-dimethyl-5-oxo-2- .sup. No(*) No
Comparative thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile
6-2 4-[3-phenyl-4,4-dimethyl-5-oxo-2-thioxoimidazolidin- 149 n/a
(**) (6b) 1-yl]-2-trifluoromethylbenzonitrile [RD10] 5-3b
4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2- 125 132 (5c)
thioxoimidazolidin-1-yl]-2-trifluoromethyl- [RD7] benzonitrile 3-3
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-5-oxo-2- 137 122 (3c)
thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile [RD8] 2-4
4-[3-(4-aminophenyl)-4,4-dimethyl-5-oxo-2- 273 n/a (2d)
thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile [RD9] 4
Chloroacetic acid 4-[3-(4-cyano-3- 131 n/a (4a)
trifluoromethylphenyl)-5,5-dimethyl-4-oxo-2- [RD13]
thioxoimidazolidin-1-yl]phenyl ester 8-2
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3- 147 n/a (8b)
diazaspiro[4.4]non-3-yl)-2-trifluoromethylbenzonitrile [RD35] 7-3b
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7- 124 128 (7c)
diazaspiro[3.4]oct-7-yl)-2-trifluoromethylbenzonitrile [RD37] 9-3
4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3- 194 n/a (9c)
diazaspiro[4.5]dec-3-yl)-2-trifluoromethylbenzonitrile [RD48] 10-3
4-(4-oxo-2-thioxo-1-(4-methylphenyl)-1,3- 232 n/a (10c)
diazaspiro[4.5]undec-3-yl)-2- [RD49] trifluoromethylbenzonitrile 28
4-(8-methyl-4-oxo-2-thioxo-1,3,8-triazaspiro[4.5]dec- No n/a
Comparative 3-yl)-2-trifluoromethylbenzonitrile (28a) [RD52] 27-3
4-(8-methyl-4-oxo-2-thioxo-1-(4-methylphenyl)-1,3,8- 638 n/a (27c)
triazaspiro[4.5]dec-3-yl)-2-trifluoromethylbenzonitrile [RD53] 26
4-[1-(4-cyanophenyl)-4-oxo-2-thioxo-l,3- 469 n/a (26a)
diazaspiro[4.4]non-3-yl]-2-trifluoromethylbenzonitrile [RD54] 25
4-[1-(4-nitrophenyl)-4-oxo-2-thioxo-1,3- 498 n/a (25a)
diazaspiro[4.4]non-3-yl]-2-trifluoromethylbenzonitrile [RD55] 12-2
4-(8-oxo-6-thioxo-5-(4-biphenyl)-5,7- 283 n/a (12b)
diazaspiro[3.4]oct-7-yl)-2-trifluoromethylbenzonitrile [RD57] 11-2
4-(8-oxo-6-thioxo-5-(4-hydroxyphenyl)-5,7- 162 n/a (11b)
diazaspiro[3.4]oct-7-yl)-2-trifluoromethylbenzonitrile [RD58] 17
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-2,5- 278 287 (17a)
dithioxoimidazolidin-1-yl]-2- [RD59] trifluoromethylbenzonitrile 18
4-[3-(4-hydroxyphenyl)-4,4-dimethyl-2,5- 369 511 (18a)
dioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile [RD60] 22-2
2-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimethyl- 523 >500
(22b) 4-oxo-2-thioxoimidazolidin-1-yl]benzoic acid [RD65] 20-2
4-(4,4-dimethyl-5-oxo-2-thioxo-3-(4- 143 144 (20b)
trifluoromethylphenyl)imidazolidin-1-yl)-2- [RD66]
trifluoromethylbenzonitrile 21-2
4-(4,4-bischloromethyl-5-oxo-2-thioxo-3-(4- 521 >500 (21b)
methylphenyl)imidazolidin-1-yl)-2- [RD67]
trifluoromethylbenzonitrile 19-2
4-(4-fluoromethyl-4-methyl-5-oxo-2-thioxo-3-(4- 126 129 (19b)
methylphenyl)imidazolidin-1-yl)-2- [RD68]
trifluoromethylbenzonitrile 23-2
4-(8-oxo-6-thioxo-5-(2-methylphenyl)-5,7- 258 232 (23b)
diazaspiro[3.4]oct-7-yl)-2-trifluoromethylbenzonitrile [RD71] 30-2
4-(5-methyl-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7- No No
Comparative yl)-2-trifluoromethylenzonitrile (30b) [RD73] 30-3
4-(5-methyl-6,8-dioxo-5,7-diazaspiro[3.4]oct-7-yl)-2- No No
Comparative trifluoromethylbenzonitrile (30c) [RD74] 31-2
4-(1-methyl-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3- No No
Comparative yl)-2-trifluoromethylbenzonitrile (31b) [RD75] 31-3
4-(1-methyl-2,4-dioxo-1,3-diaza-spiro[4.4]non-3-yl)- No No
Comparative 2-trifluoromethylbenzonitrile (31c) [RD76] 24-3
4-(4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2- No No Comparative
trifluoromethylbenzonitrile (24c) [RD77] 15-2
4-[4,4-dimethyl-3-(4-pyridin-2-yl)-5-oxo-2- 723 n/a (15b)
thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile [RD82] 14-2
4-[4,4-dimethyl-3-(4-methylpyridin-2-yl)-5-oxo-2- 457 n/a (14b)
thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile [RD83] 16-2
4-[5-(5-methyl-2H-pyrazol-3-yl)-8-oxo-6-thioxo-5,7- >1000 n/a
Comparative diaza-spiro[3.4]oct-7-yl]-2-trifluoromethyl- (16b)
benzonitrile [RD84] 13-2 4-(8-oxo-6-thioxo-5-(4-biphenyl)-5,7-
>1000 n/a (12b)
diazaspiro[3.4]oct-7-yl)-2-trifluoromethylbenzonitrile [RD85] 32
4-(8-methylimino-6-thioxo-5-p-tolyl- 222 421 (32a)
5,7-diazaspiro[3.4]oct-7-yl)-2-trifluoromethyl-benzonitrile [RD90]
33 1-[3-(4-cyano-3-trifluoromethyl-phenyl)-5,5-dimethyl- 157 239
(33a) 2-thioxo-1-p-tolyl-imidazolidin-4-ylidene]-3-ethyl- [RD91]
thiourea 34 1-[7-(4-cyano-3-trifluoromethyl-phenyl)-6-thioxo-5-p-
176 276 (34a) tolyl-5,7-diaza-spiro[3.4]oct-8-ylidene]-3-phenyl-
[RD92] thiourea 35
1-(4-Cyano-3-trifluoromethyl-phenyl)-3-[7-(4-cyano- 144 158 (35a)
3-trifluoromethyl-phenyl)-6-thioxo-5-p-toly1-5,7- [RD93]
diaza-spiro[3.4]oct-8-ylidene]-thiourea 36-2
4-[8-(4-hydroxymethyl-phenyl)-5-oxo-7-thioxo-6-aza- 311 337 (36b)
spiro[3.4]oct-6-yl]-2-trifluoromethyl-benzonitrile [RD110] 37
4-[5-(4-formylphenyl)-8-oxo-6-thioxo-5,7- n/a 263 (37a)
diazaspiro[3.4]oct-7-yl]-2-trifluoromethyl-benzonitrile [RD114] 38
4-{5-[4-(1-hydroxyethyl)-phenyl]-8-oxo-6-thioxo-5,7- n/a 187 (38a)
diazaspiro[3.4]oct-7-yl}-2-trifluoromethyl-benzonitrile [RD116] 39
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6- n/a 197 (39a)
thioxo-5,7-diazaspiro[3.4]oct-5-yl]-phenyl}-acrylic [RD117] acid
ethyl ester 40 4-{5-[4-(3-hydroxypropenyl)-phenyl]-8-oxo-6-thioxo-
n/a 114 (40a) 5,7-diazaspiro[3.4]oct-7-yl}-2- [RD120]
trifluoromethylbenzonitrile 41-2
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6- No n/a (41b)
thioxo-5,7-diazaspiro[3.4]oct-5-yl]-phenyl}-propionic [RD128] acid
methyl ester 41-4
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6- 224 n/a (41d)
thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}- [RD133] propionamide
41-5 3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6- 234 n/a
(41e) thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}-N- [RD134]
methyl-propionamide 41-6
3-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6- 732 n/a (41f)
thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}-N-(2- [RD135]
hydroxyethyl)-propionamide 42-2
4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6- 432 n/a (42b)
thioxo-5,7-diazaspiro[3.4]oct-5-yl]-phenyl}-butyric [RD129] acid
methyl ester 42-4
4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6- 112 n/a (42d)
thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}- [RD130] butyramide
42-5 4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6- 92 n/a
(42e) thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}-N- [RD131]
methyl-butyramide 43-4
4-[8-Oxo-5-(4-piperazin-1-yl-phenyl)-6-thioxo-5,7- 718 n/a (43e)
diazaspiro[3.4]oct-7-yl]-2-trifluoromethylbenzonitrile [RD137] 43-5
4-{5-[4-(4-methanesulfonylpiperazin-1-yl)-phenyl]-8- 138 n/a (43f)
oxo-6-thioxo-5,7-diazaspiro[3.4]oct-7-yl}-2- [RD138]
trifluoromethylbenzonitrile 44-2 44-2)
3-{4-[7-(4-Cyano-3-trifluoromethyl-phenyl)-8- 113 (44b)
oxo-6-thioxo-5,7-diaza-spiro[3.4]oct-5-yl]-phenyl}- [RD119]
acrylamide, (*)No: the compound did not inhibit AR response
reporter; (**) n/a: the compound was not examined in this
assay.
[0338] One previously unrecognized property of AR overexpression in
hormone refractory prostate cancer is its ability to switch
antagonists to agonists. Therefore, only those compounds with
minimal or no agonistic activities are qualified to be
anti-androgens for this disease. To determine agonistic activities
of different compounds, we examined their stimulating activities on
AR using the AR response reporter as the measure in the LN-AR
system in the absence of R1881. Table 2 lists the agonistic
activities of different compounds. Consistent with previous
results, bicalutamide activated AR in hormone refractory prostate
cancer. The diarylthiohydantoin derivatives such as examples 7-3b
(RD37), 33 (RD91), 34 (RD92), and 35 (RD93) have no agonistic
activity. In contrast, RU59063, and other anti-androgenic compounds
listed as examples in U.S. Pat. No. 5,705,654, such as examples
30-2, 30-3, 31-2, 31-3, and 24-3 (RD73-RD77) strongly activated AR
in hormone refractory prostate cancer.
TABLE-US-00002 TABLE 2 Agonistic activities of selective test
substances on AR response reporter in hormone refractory prostate
cancer Fold induction by increasing concentrations of compounds
Example Name 0.1 .mu.M 1 .mu.M 10 .mu.M DMSO Dimethyl sulfoxide
1.00 (*) 1.00 1.00 R1881 methyltrienolone 44.33 n/a(**) n/a
Bicalutamide N-[4-cyano-3-(trifluoromethyl)phenyl]-3-[(4- 1.66 3.04
10.40 fluorophenyl)sulfonyl]-2-hydroxy-2- methylpropanamide 29
4-[3-(4-hydroxybutyl)-4,4-dimethyl-5-oxo-2- 10.99 20.84 34.62 Comp.
thioxoimidazolidin-1-yl]-2- trifluoromethylbenzonitrile 7-3b
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7- 0.87 1.19 0.89 (7c)
diazaspiro[3.4]oct-7-yl)-2- [RD37] trifluoromethylbenzonitrile 33
1-[3-(4-cyano-3-trifluoromethyl-phenyl)-5,5- 1.30 1.18 1.28 (33a)
dimethyl-2-thioxo-1-p-tolyl-imidazolidin-4- [RD91]
ylidene]-3-ethyl-thiourea 34
1-[7-(4-cyano-3-trifluoromethyl-phenyl)-6- 1.19 1.41 1.17 (34a)
thioxo-5-p-tolyl-5,7-diaza-spiro[3.4]oct-8- [RD92]
ylidene]-3-phenyl-thiourea 35
1-(4-Cyano-3-trifluoromethyl-phenyl)-3-[7-(4- 1.26 1.10 1.30 (35a)
cyano-3-trifluoromethyl-phenyl)-6-thioxo-5-p- [RD93]
tolyl-5,7-diaza-spiro[3.4]oct-8-ylidene]-thiourea 30-2
4-(5-methyl-8-oxo-6-thioxo-5,7- 14.88 19.41 35.22 Comp.
diazaspiro[3.4]oct-7-yl)-2- (30b) trifluoromethylenzonitrile [RD73]
30-3 4-(5-methyl-6,8-dioxo-5,7-diazaspiro[3.4]oct-7- 11.39 14.26
30.63 Comp. yl)-2-trifluoromethylbenzonitrile (30c) [RD74] 31-2
4-(1-methyl-4-oxo-2-thioxo-1,3- 17.03 16.63 33.77 Comp.
diazaspiro[4.4]non-3-yl)-2- (31b) trifluoromethylbenzonitrile
[RD76] 31-3 4-(1-methyl-2,4-dioxo-1,3-diaza-spiro[4.4]non- 11.99
19.77 38.95 Comp. 3-yl)-2-trifluoromethylbenzonitrile (31c) [RD76]
24-3 4-(4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)- 14.88 22.48
37.09 Comp. 2-trifluoromethylbenzonitrile (24c) [RD77] (*) Fold
induction: activities induced by a specific test substance over
activities in DMSO vehicle; (**)n/a: the compound was not examined
in this assay.
[0339] To examine the specificity of AR inhibitors, selective
compounds were tested in LNCaP cells with an over expression of
glucocorticoid receptor (GR), the closest member of AR in the
nuclear receptor family. These cells also carry a GR response
reporter and the reporter activity was induced by dexamethasone, a
GR agonist and the induction was blocked by RU486, a GR inhibitor.
Example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethyl benzonitrile) had no effect on GR in this system.
Effect of Compounds on AR by Measuring Secreted Levels of Prostate
Specific Antigen (PSA)
[0340] It is well established that PSA levels are indicators of AR
activities in prostate cancer. To examine if the compounds affect
AR function in a physiological environment, we determined secreted
levels of endogenous PSA induced by R1881 in the AR-overexpressed
LNCaP cells (LNCaP-AR, also abbreviated LN-AR). The LNCaP-AR cells
are a line of lymph node carcinoma of prostate cells transduced
with a plasmid that makes express androgen receptors. LNCaP-AR
cells were maintained in Iscove's medium containing 10% FBS. Two
days prior to drug treatment, the cells were grown in Iscove's
medium containing 10% CS-FBS to deprive of androgens. The cells
were split and grown in Iscove's medium containing 10% CS-FBS with
appropriate concentrations of R1881 and the test compounds. After
four days incubation, secreted PSA levels were assayed using PSA
ELISA kits (American Qualex, San Clemente, Calif.)
[0341] The secreted PSA level of LNCaP-AR cells was strongly
induced by 25 pM of R1881. In contrast, PSA was not induced in the
parental LNCaP cells until concentration of R1881 reached 100 pM.
This is consistent with our previous report that the AR in hormone
refractory prostate cancer is hyper-sensitive to androgens. A
dose-dependent inhibition on AR activity was carried out to
determine the IC50s of different compounds in inhibiting PSA
expression, and the results were listed in Table 1. IC50s of the
selective compounds on PSA expression closely resemble those
measured by the reporter assay, confirming that the diarylhydantoin
derivatives are strong inhibitors of AR in hormone refractory
prostate cancer.
[0342] We also examined agonistic activities of selective compounds
on AR in hormone refractory prostate cancer using secreted PSA as
the surrogate marker. To do this, androgen-starved AR over
expressed LNCaP cells were incubated with increasing concentrations
of the compounds for which a synthesis is described above in the
absence of R1881 and secreted PSA in the culture medium was
measured 4 days later.
[0343] Table 3 lists the agonistic activities of the selective
compounds. Consistent with the results obtained from the reporter
assay, the diarylthiohydantoin derivatives such as examples 7-3b
(RD37), 33 (RD91), 34 (RD92), and 35 (RD93) have no agonistic
activities. In contrast, RU59063, and other antiandrogenic
compounds listed as examples in U.S. Pat. No. 5,705,654, such as
examples 30-2 (RD73), 30-3 (RD74), and 31-2 (RD75) stimulated PSA
expression in hormone refractory prostate cancer.
TABLE-US-00003 TABLE 3 Agonistic activities of selective test
substances on endogenous PSA in hormone refractory prostate cancer
Fold induction by increasing concentrations of compounds Example
Name 0.1 .mu.M 1 .mu.M 10 .mu.M DMSO Dimethyl sulfoxide 1.00 (*)
1.00 1.00 R1881 methyltrienolone 20.69 n/a(**) n/a Bicalutamide
N-[4-cyano-3-(trifluoromethyl)phenyl]-3-[(4- 2.00 2.55 5.55
fluorophenyl)sulfonyl]-2-hydroxy-2- methylpropanamide 29
4-[3-(4-hydroxybutyl)-4,4-dimethyl-5-oxo-2- 6.88 11.50 21.50 Comp.
thioxoimidazolidin-1-yl]-2- trifluoromethylbenzonitrile 7-3b
4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7- 1.25 1.20 1.15 (7c)
diazaspiro[3.4]oct-7-yl)-2- [RD37] trifluoromethylbenzonitrile 33
1-[3-(4-cyano-3-trifluoromethyl-phenyl)-5,5- 1.06 1.30 0.85 (33a)
dimethyl-2-thioxo-1-p-tolyl-imidazolidin-4- [RD91]
ylidene]-3-ethyl-thiourea 34
1-[7-(4-cyano-3-trifluoromethyl-phenyl)-6- 1.31 1.05 0.90 (34a)
thioxo-5-p-tolyl-5,7-diaza-spiro[3.4]oct-8- [RD92]
ylidene]-3-phenyl-thiourea 35
1-(4-Cyano-3-trifluoromethyl-phenyl)-3-[7-(4- 1.44 1.30 1.05 (35a)
cyano-3-trifluoromethyl-phenyl)-6-thioxo-5-p- [RD93]
tolyl-5,7-diaza-spiro[3.4]oct-8-ylidene]-thiourea 30-2
4-(5-methyl-8-oxo-6-thioxo-5,7- 6.25 17.95 25.65 Comp.
diazaspiro[3.4]oct-7-yl)-2- (30b) trifluoromethylenzonitrile [RD73]
30-3 4-(5-methyl-6,8-dioxo-5,7-diazaspiro[3.4]oct-7- 7.50 15.20
23.75 Comp. yl)-2-trifluoromethylbenzonitrile (30c) [RD74] 31-2
4-(1-methyl-4-oxo-2-thioxo-1,3- 8.13 18.20 17.50 Comp.
diazaspiro[4.4]non-3-yl)-2- (31b) trifluoromethylbenzonitrile
[RD75] (*) Fold induction: activities induced by a specific test
substance over activities in DMSO vehicle; (**)n/a: the compound
was not examined in this assay.
Effect of Compounds on AR Mitochondrial Activity by MTS Assay
[0344] LNCaP-AR cells were maintained in Iscove's medium containing
10% FBS. The compounds were examined for their effect on growth of
hormone refractory prostate cancer cells. Overexpressed LNCaP cells
were used because these cells behave as hormone refractory prostate
cancer cells in vitro and in vivo (1). We measured mitochondria
activity by MTS assay, a surrogate for growth. LNCaP cells with
overexpressed AR (LN-AR) were maintained in Iscove's medium
containing 10% FBS. Two days prior to drug treatment, the cells
were grown in Iscove's medium containing 10% CS-FBS to deprive of
androgens. The cells were then split and grown in Iscove's medium
containing 10% CS-FBS with appropriate concentrations of R1881 and
increasing concentrations of the test compounds. After four days
incubation, cell growth was monitored by MTS (Promega, Madison,
Wis.).
[0345] Consistent with the reporter assay and PSA assay, growth of
the AR-overexpressed LNCaP was stimulated by 25 microM of R1881,
but the parental cells were not stimulated until R1881
concentration reached 100 microM. FIG. 2 shows the inhibitory
effect of selected compounds on growth of hormone refractory
prostate cancer in the presence of 100 pM of R1881. The current
clinical drug bicalutamide did not inhibit hormone refractory
prostate cancer. In contrast, example 5-3b (RD7)
(4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-tr-
ifluoromethyl-benzonitrile) and example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) inhibited hormone refractory prostate
cancer with high potency.
[0346] We examined if growth inhibition in the MTS assay occurs by
targeting AR, example 5-3b (RD7)
(4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-tr-
ifluoromethyl-benzonitrile) and example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) were tested in DU-145 cells, a prostate
cancer cell line that lacks AR expression. These compounds had no
growth inhibitory effect on DU-145 cells. The compounds did not
inhibit cells other than AR-expressed prostate cancer cells, as
they had no growth effect on MCF7 and SkBr3, two commonly used
breast cancer cells, or 3T3, a normal mouse fibroblast cell
line.
[0347] Examples of in vitro biological activity of
diarylthiohydantoin derivatives are shown in the FIGS. 3, 4 and 5.
For example, based on relative luciferase activity, FIG. 3
indicates that at a concentration of 500 nM the compounds ranked,
in order of most active to least active as follows:
RD152>RD153>RD145>RD163>RD161=RD162>bicalutamide.
For example, based on relative PSA level, FIG. 4 indicates that at
a concentration of 500 nM the compounds ranked, in order of most
active to least active as follows:
RD138>RD131>RD37>RD133>RD134>RD137>RD138>RD135>bi-
calutamide. For example, based on relative MTS units, FIG. 5
indicates that at a concentration of 500 nM the compounds ranked,
in order of most active to least active as follows:
RD168>RD37>RD141>RD162>bicalutamide.
Inhibitory Effect on Hormone Refractory Prostate Cancer Xenograft
Tumors
[0348] Example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) was used to examine if the diarylhydantoin
derivatives have in vivo effects on hormone refractory prostate
cancer. First we examined this compound on xenograft tumors
established from AR-overexpressed LNCaP cells. The engineered cells
in Matrigel (Collaborative Biomedical) were injected subcutaneously
into the flanks of the castrated male SCID mice. Tumor size was
measured weekly in three dimensions using calipers. After xenograft
tumors established (tumor size reached at least 40 mm.sup.3), mice
with tumors were randomized and treated with different doses of
compounds orally once daily. Consistent with clinical observation,
current clinical drug bicalutamide did not inhibit growth of
hormone refractory prostate cancer (same as vehicle) (FIG. 7A). In
contrast, example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) strongly inhibited growth of these tumors
(FIG. 7A) and the inhibition is dose-dependent (FIG. 7B).
Furthermore, example 7-3b (RD37) inhibited PSA expression (FIG. 8),
the clinical marker for hormone refractory prostate cancer.
[0349] Example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) was also tested in another xenograft model
of hormone refractory prostate cancer, hormone refractory LAPC4.
This model was established from passaging of hormone sensitive
prostate cancer in castrated mice, which mimics the clinical
progression of prostate cancer (2). Similar to the finding using
AR-overexpressed LNCaP xenograft model, current clinical drug
bicalutamide did not inhibit growth and PSA expression in hormone
refractory LAPC4 xenograft model (same as vehicle) (FIGS. 9A and
9B). In contrast, example 7-3b (RD37) strongly inhibited growth and
PSA expression of these tumors (FIGS. 9A and 9B).
Inhibitory Effect on Growth of Hormone Sensitive Prostate Cancer
Cells
[0350] To determine if the diarylthiahydantoin derivatives also
inhibit hormone sensitive prostate cancer cells, we tested some
selective compounds on growth of LNCaP cells by measuring MTS of
mitochondria activities. In contrast to have no effect on growth of
hormone refractory prostate cancer, the current clinical drug
bicalutamide mildly inhibited hormone sensitive LNCaP cells in a
dose-dependent manner Example 5-3b (RD7)
(4-[3-(4-methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl-
]-2-trifluoromethyl-benzonitrile) and example 7-3b (RD37)
(4-(8-oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-7-yl)-2-trif-
luoromethylbenzonitrile) inhibited hormone sensitive prostate
cancer with a 10-fold higher potency than bicalutamide (FIG.
10).
In Vivo Biological Assay
[0351] All animal experiments were performed in compliance with the
guidelines of the Animal Research Committee of the University of
California at Los Angeles. Animals were bought from Taconic and
maintained in a laminar flow tower in a defined flora colony.
LNCaP-AR and LNCaP-vector cells were maintained in RPMI medium
supplemented with 10% FBS. 10.sup.6 cells in 100 .mu.l of 1:1
Matrigel to RPMI medium were injected subcutaneously into the
flanks of intact or castrated male SCID mice. Tumor size was
measured weekly in three dimensions
(length.times.width.times.depth) using calipers. Mice were
randomized to treatment groups when tumor size reached
approximately 100 mm.sup.3. Drugs were given orally every day at 10
mg/kg and 50 mg/kg. To obtain pharmacodynamic readout, the animals
were imaged via an optical CCD camera, 3 hours after last dose of
the treatment. A ROI is drawn over the tumor for luciferase
activity measurement in photon/second. The right panels (FIGS. 12C
and 12D) were a representation of the ROIs measurements. Data are
shown in FIGS. 11 and 12A through 12D. Over 18 days RD162 was
effective to prevent tumor growth and even to cause tumor
shrinkage, and was distinctly more effective than bicalutamide.
[0352] The pharmacokinetics of bicalutamide,
4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-spiro[3.4-
]oct-5-yl]-toluene [RD37],
N-methyl-4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-dia-
za-spiro[3.4]oct-5-yl]phenyl}butanamide [RD131], and
N-methyl-4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza--
spiro[3.4]oct-5-yl]-2-fluorobenzamide (52d) [RD162] were evaluated
in vivo using 8 week-old FVB mice which were purchased from Charles
River Laboratories. Mice were divided into groups of three for each
time points. Two mice were not treated with drug and two other mice
were treated with vehicle solution. Each group was treated with 10
mg per kilogram of body weight.
[0353] The drug was dissolved in a mixture 1:5:14 of
DMSO:PEG400:H.sub.2O. (Vehicle solution) and was administered into
mice through the tail vein. The animals are warmed under a heat
lamp for approximately 20 minutes prior to treatment to dilate
their tail vein. Each mouse was placed into a mouse restrainer
(Fisher Sci. Cat#01-288-32A) and was injected with 200 .mu.l of
drug in vehicle solution into the dilated tail vein. After drug
administration, the animals were euthanized via CO.sub.2 inhalation
at different timepoints: 5 mn, 30 mn, 2 h, 6 h, 16 h. Animals were
immediately bleed after exposure to CO.sub.2 via cardiac puncture
(1 ml BD syringe +27 G 5/8 needle). For oral dosage, the drug was
dissolved in a mixture 50:10:1:989 of
DMSO:Carboxymethylcellulose:Tween80:H2O before oral administration
via a feeding syringe.
[0354] The serum samples were analyzed to determine the drug's
concentration by the HPLC which (Waters 600 pump, Waters 600
controller and Waters 2487 detector) was equipped with an Alltima
C18 column (3.mu., 150 mm.times.4.6 mm). The RD37, RD131, and RD162
compounds were detected at 254 nm wave length and bicalutamide was
detected at 270 nm wave length.
[0355] The samples for HPLC analysis were prepared according to the
following procedure: [0356] Blood cells were separated from serum
by centrifugation. [0357] To 400 .mu.l of serum were added 80 .mu.l
of a 10 .mu.M solution of an internal standard and 520 .mu.l of
acetonitrile. Precipitation occurred. [0358] The mixture was
vortexed for 3 minutes and then placed under ultrasound for 30
minutes. [0359] The solid particles were filtered off or were
separated by centrifugation. [0360] The filtrate was dried under an
argon flow to dryness. The sample was reconstructed to 80 .mu.l
with acetonitrile before analyzing by HPLC to determine the drug
concentration. [0361] Standard curve of drug was used to improve
accuracy.
[0362] The concentration of RD162 in plasma as a function of time
resulting from intravenous and from oral administration is shown in
FIG. 13. The steady state concentration (Css) of bicalutamide,
RD131, and RD162 is shown in Table 4. The concentration at steady
state of RD162 is essentially as good as that of bicalutamide, and
substantially better than RD131.
TABLE-US-00004 TABLE 4 Steady-state concentration of bicalutamide,
RD131, and RD162 in mice plasma. IC50 Css, 10 mg/kg Css, 25 mg/kg
Css, 50 mg/kg Name [nM] LogP [.mu.M] [.mu.M] [.mu.M] Bic. 1000 2.91
10.0 11.4 11.9 RD131 92 3.44 0.39 0.43 0.40 RD162 122 3.20 9.9 10.7
10.2
Ranking of Compounds in Tiers
[0363] Tables 5-10 present diarylhydantoin compounds grouped into
Tiers 1-6. Table 11 presents diarylhydantoin compounds which have
not been placed into a tier. The placement of compounds into tiers
was based on available data coupled with analytical judgment. Data
considered included in vitro assays (AR response reporter system in
LNCaP cell line, PSA level measurement, MTS mitochondrial assay)
and in vivo experiments (tumor size measured directly or by
emission induced by luciferase reporter gene, pharmacokinetic
assays based on blood plasma levels). Not every compound was
subjected to each assay. Not all data that was generated is shown.
Judgment was applied in ranking compounds relative to each other
for their utility in treating prostate cancer, in particular when
ranking two compounds for which the same experiments were not
performed. Characteristics considered in establishing the ranking
include AR antagonism activity, lack of AR agonism in hormone
refractory cells, prevention of tumor growth, tumor shrinkage, and
pharmacokinetic behavior, with a longer residence time in blood
being advantageous.
Tier 1
[0364] Generally, Tier 1 compounds are diarylthiohydantoins with a
disubstituted left hand aryl ring that are disubstituted on the
right hydantoin carbon, and have either an oxygen or N substituent
on the left hydantoin carbon. It is expected that the amido
substituent hydrolyzes to an oxygen in aqueous solutions such as
encountered in biological systems, in vitro and in vivo. RD100 has
good activity with an iodine instead of a CF.sub.3 substituent on
the left hand aryl ring.
[0365] Tier 1 compounds (see Table 5) were judged to be much better
than bicalutamide for treating prostate cancer. However, RD37 and
RD131 were found to metabolize fast, that is, have a short
residence time in blood. RD162 had desirable pharmacokinetics.
[0366] FIG. 17 shows that under treatment with bicalutamide, PSA
levels for LNCaP cells stayed the same or increased relative to
treatment with vehicle solution, whereas under treatment with
RD162, PSA levels decreased. FIG. 18 illustrates that under
treatment with vehicle solution, tumors continued to increase in
size. By contrast, under treatment with RD162 at a dose of 1 mg per
kg body weight per day, the rate of tumor increase decreased, and
the size of the tumor appeared to be stabilizing after about 17
days. Under treatment with RD162 at a dose of 10 mg per kg body
weight per day, tumor size decreased with time. FIGS. 19A through
19C that under treatment with RD162 at a dose of 10 mg per kg body
weight per day, photon emission associated with luciferase activity
decreased. FIGS. 20A through 20C show that treatment with RD162 at
this dose resulted in a decrease or stabilization of tumor size and
a decrease in photon emission associated with luciferase
activity.
[0367] FIGS. 21A through 21C show that under treatment with RD162,
RD162', RD162'', RD169, and RD170 at doses of 100, 200, 500, and
1000 nM, PSA levels of LN-AR cells decreased. Moreover, the higher
the dose, the lower the PSA level. FIGS. 23A and 23B present
urogenital tract weight and rate of photon emission associated with
luciferase activity initially and after 14 days of treatment with
bicalutamide or with RD162 for intact and castrated mice. The
weight and rate of photon emission increased for both intact and
castrated mice. Treatment of castrated mice with RD162 resulted in
a decrease in weight and photon emission with respect to the
untreated castrated mice, as did treatment with bicalutamide.
[0368] Thus, Tier 1 compounds are particularly advantageous for use
as AR antagonists, and as therapeutic agents for hormone refractory
prostate cancer. They may be useful to treat other AR related
diseases or conditions such as benign prostate hyperplasia, hair
loss, and acne. These and related compounds may also be useful as
modulators of other nuclear receptors, such as glucocorticoid
receptor, estrogen receptor, and peroxisome proliferator-activated
receptor, and as therapeutic agents for diseases in which nuclear
receptors play a role, such as breast cancer, ovarian cancer,
diabetes, cardiac diseases, and metabolism related diseases. They
may be useful in assays e.g. as standards, or as intermediates or
prodrugs.
TABLE-US-00005 TABLE 5 TIER 1 COMPOUNDS ##STR00115## RD7
##STR00116## RD8 ##STR00117## RD10 ##STR00118## RD35 ##STR00119##
RD36 ##STR00120## RD37 ##STR00121## RD57 ##STR00122## RD58
##STR00123## RD90 ##STR00124## RD91 ##STR00125## RD92 ##STR00126##
RD93 ##STR00127## RD94 ##STR00128## RD95 ##STR00129## RD96
##STR00130## RD97 ##STR00131## RD100 ##STR00132## RD102
##STR00133## RD119 ##STR00134## RD120 ##STR00135## RD130
##STR00136## RD131 ##STR00137## RD145 ##STR00138## RD152
##STR00139## RD153 ##STR00140## RD163 ##STR00141## RD162
##STR00142## RD162' ##STR00143## RD162'' ##STR00144## RD168
##STR00145## RD169 ##STR00146## RD170
Tier 2
[0369] Tier 2 compounds (see Table 6) were significantly better
than bicalutamide for treating prostate cancer, although there were
indications that RD54 could act as an agonist. FIG. 3 illustrates
that compounds RD145, RD152, RD153, RD162, and RD163 in Tier 1 and
RD161 in Tier 2 dosed at concentrations ranging from 125 nM to 1000
nM acted to reduce luciferase activity in LNCaP-AR cells whereas
control solutions of DMSO and of bicalutamide had little or no
effect. FIG. 4 illustrates, for example, that at concentrations of
1000 nM, compounds RD37 and RD131, in Tier 1, caused a greater
decrease in PSA level of LNCaP-AR cells than RD133, RD134, and
RD138 in Tier 2. FIG. 11 presents tumor volume over time, and
illustrates that under treatment with bicalutamide or vehicle
solution, tumors continued to grow, whereas under treatment with
RD162, in Tier 1, tumors decreased in size. FIGS. 12A through 12D
illustrate that photon emission associated with luciferase activity
remained about the same or increased under treatment with
bicalutamide relative to treatment with vehicle solution, whereas
photon emission decreased under treatment with RD162. FIG. 14
illustrates that under treatment with bicalutamide, there was
little or no decrease in PSA levels, whereas under treatment with
RD131 and RD162, PSA levels decreased. FIG. 15A illustrates that
the IC.sub.50 for RD37, RD 131, and RD162, in Tier 1, was much
lower than the IC.sub.50 for bicalutamide.
[0370] Generally, Tier 2 compounds are structurally similar to Tier
1 compounds, but with different substituents on the right hand aryl
ring. Tier 2 compounds are advantageous for use as AR antagonists,
and as therapeutic agents for hormone refractory prostate cancer.
They may be useful to treat other AR related diseases or conditions
such as benign prostate hyperplasia, hair loss, and acne. These and
related compounds may also be useful as modulators of other nuclear
receptors, such as estrogen receptor and peroxisome
proliferator-activated receptor, and as therapeutic agents for
diseases in which nuclear receptors play a role, such as breast
cancer, ovarian cancer, diabetes, cardiac diseases, and metabolism
related diseases. They may be useful in assays e.g. as standards,
or as intermediates or prodrugs.
TABLE-US-00006 TABLE 6 TIER 2 COMPOUNDS ##STR00147## RD6
##STR00148## RD13 ##STR00149## RD48 ##STR00150## RD49 ##STR00151##
RD51 ##STR00152## RD53 ##STR00153## RD54 ##STR00154## RD55
##STR00155## RD63 ##STR00156## RD66 ##STR00157## RD68 ##STR00158##
RD71 ##STR00159## RD87 ##STR00160## RD103 ##STR00161## RD110
##STR00162## RD111 ##STR00163## RD114 ##STR00164## RD116
##STR00165## RD133 ##STR00166## RD134 ##STR00167## RD138
##STR00168## RD161
Tier 3
[0371] Tier 3 compounds (see Table 7) were judged to be slightly
better than bicalutamide for treating prostate cancer. RD133,
RD134, and RD138 (in Tier 2) caused a greater decrease in PSA level
of LNCaP-AR cells than RD135 and RD137, in Tier 3. All of these
compounds caused a greater decrease in PSA level than
bicalutamide.
[0372] Other Tier 3 compounds (not shown) were not
diarylthiohydantoins, and were comparable in activity to prior art
monoarylhydantoin compounds RD2, RD4, and RD5.
[0373] Thus, Tier 3 compounds are useful as AR antagonists, and as
therapeutic agents for hormone refractory prostate cancer. They may
be useful to treat other AR related diseases or conditions such as
benign prostate hyperplasia, hair loss, and acne. These and related
compounds may also be useful as modulators of other nuclear
receptors, such as estrogen receptor and peroxisome
proliferator-activated receptor, and as therapeutic agents for
diseases in which nuclear receptors play a role, such as breast
cancer, ovarian cancer, diabetes, cardiac diseases, and metabolism
related diseases. They may be useful in assays e.g. as standards,
or as intermediates or prodrugs.
TABLE-US-00007 TABLE 7 TIER 3 COMPOUNDS ##STR00169## RD3
##STR00170## RD4 ##STR00171## RD5 ##STR00172## RD69 ##STR00173##
RD127 ##STR00174## RD128 ##STR00175## RD129 ##STR00176## RD135
##STR00177## RD137
Tier 4
[0374] Tier 4 compounds (see Table 8) were judged to be no better
than bicalutamide for treating prostate cancer. Tier 4 RD 39 and
RD40 and Tier 1 RD37, for example, differ only in the substituent
on the lower right carbon of the hydantoin ring. The substituents
on the right hand aryl ring may also affect activity.
[0375] Some Tier 4 compounds (including those shown and others that
are not shown) were not diaryl compounds (lacking the right hand
aryl ring), were not thiohydantoins, were not disubstituted on the
carbon on the lower right hand of the hydantoin ring, and/or had
substituents other than oxygen or amido on the lower left hand
carbon of the hydantoin ring. This provides evidence of the
surprising advantages of diarylthiohydantoins that are
disubstituted on the lower right hand carbon of the hydantoin ring
and have oxygen or amido on the lower left hand carbon of the
hydantoin ring.
[0376] Thus, Tier 4 compounds may be useful as AR antagonists, and
as therapeutic agents for hormone refractory prostate cancer, at
least to the extent that they are comparable to bicalutamide. They
may be useful to treat other AR related diseases or conditions such
as benign prostate hyperplasia, hair loss, and acne. These and
related compounds may also be useful as modulators of other nuclear
receptors, such as estrogen receptor and peroxisome
proliferator-activated receptor, and as therapeutic agents for
diseases in which nuclear receptors play a role, such as breast
cancer, ovarian cancer, diabetes, cardiac diseases, and metabolism
related diseases. They may be useful in assays e.g. as standards,
or as intermediates or prodrugs.
TABLE-US-00008 TABLE 8 TIER 4 COMPOUNDS ##STR00178## RD2
##STR00179## RD9 ##STR00180## RD21 ##STR00181## RD22 ##STR00182##
RD23 ##STR00183## RD24 ##STR00184## RD25 ##STR00185## RD26
##STR00186## RD27 ##STR00187## RD30 ##STR00188## RD31 ##STR00189##
RD39 ##STR00190## RD40 ##STR00191## RD44 ##STR00192## RD59
##STR00193## RD60 ##STR00194## RD67 ##STR00195## RD82 ##STR00196##
RD83 ##STR00197## RD117 ##STR00198## RD118 ##STR00199## RD148
##STR00200## RD149 ##STR00201## RD150 ##STR00202## RD151
Tier 5
[0377] Tier 5 compounds (see Table 9) were inactive or nearly
inactive, and thus, were worse than bicalutamide for treating
prostate cancer. The substituents on the right hand aryl ring are
important to determining activity.
[0378] Some Tier 5 compounds (some of which are shown and some that
are not shown) were not diaryl compounds (lacking the right hand
aryl ring), were not thiohydantoins, were not disubstituted on the
carbon on the lower right hand of the hydantoin ring, and/or had
substituents other than oxygen or amido on the lower left hand
carbon of the hydantoin ring. This provides evidence of the
surprising advantages of diarylthiohydantoins that are
disubstituted on the lower right hand carbon of the hydantoin ring
and have oxygen or amido on the lower left hand carbon of the
hydantoin ring. In particular, the terminal substituent in RD155,
RD 156, and 158 (CH.sub.2NR.sub.xR.sub.y, where R.sub.x,y=H or
methyl) is not seen as contributing to activity in these
compounds.
[0379] Tier 5 compounds would not be desirable for treatment of
prostate cancer or as AR antagonists, although these and related
compounds may be useful as modulators of other nuclear receptors,
such as estrogen receptor and peroxisome proliferator-activated
receptor, and as therapeutic agents for diseases in which nuclear
receptors play a role, such as breast cancer, ovarian cancer,
diabetes, cardiac diseases, and metabolism related diseases. They
may be useful in assays e.g. as standards, or as intermediates or
prodrugs.
TABLE-US-00009 TABLE 9 TIER 5 COMPOUNDS ##STR00203## RD32
##STR00204## RD33 ##STR00205## RD65 ##STR00206## RD84 ##STR00207##
RD85 ##STR00208## RD155 ##STR00209## RD156 ##STR00210## RD157
##STR00211## RD158
Tier 6
[0380] Tier 6 compounds (see Table 10) were inactive or nearly
inactive, and furthermore were strong agonists, and thus were much
worse than bicalutamide for treating prostate cancer. The
comparative compounds ranked very poor relative to the inventive
compounds. Notably, RD72 had very poor activity, with a chlorine
substituent on the left hand aryl ring, whereas RD7, with a
trifluoromethane, and RD100, with iodine, ranked in Tier 1. The
results for the Tier 6 compounds provide evidence of the surprising
advantages of diarylthiohydantoins that are disubstituted on the
lower right hand carbon of the hydantoin ring and have oxygen or
amido on the lower left hand carbon of the hydantoin ring, and have
certain substituents on the left hand aryl ring.
[0381] Tier 6 compounds would not be desirable for treatment of
prostate cancer or as AR antagonists.
TABLE-US-00010 TABLE 10 TIER 6 COMPOUNDS ##STR00212## RD72
##STR00213## RD73 ##STR00214## RD74 ##STR00215## RD75 ##STR00216##
RD76 ##STR00217## RD77
Untiered Compounds
[0382] For several compounds, there was insufficient experimental
data to rank them. These untiered compounds are presented in Table
11.
[0383] Based on the data and methods of the invention, and applying
judgment based on review of many compounds, including some not
shown here, one can make some observations about the untiered
compounds. Comparative example RD1 is expected to be in Tier 3 with
comparative examples RD3-RD5. RD89 is expected to hydrolyze to RD37
(Tier 1), and should therefore have comparable activity. RD104 is
expected to hydrolyze to RD58 (Tier 1), and should therefore have
comparable activity. RD105 is expected to hydrolyze to RD8 (Tier
1), and RD 139 and RD140 are expected to hydrolyze to RD138 (Tier
2), and they should therefore have comparable activity.
TABLE-US-00011 TABLE 11 UNTIERED COMPOUNDS ##STR00218## RD1
##STR00219## RD19 ##STR00220## RD52 ##STR00221## RD79 ##STR00222##
RD80 ##STR00223## RD81 ##STR00224## RD89 ##STR00225## RD104
##STR00226## RD105 ##STR00227## RD106 ##STR00228## RD115
##STR00229## RD132 ##STR00230## RD136 ##STR00231## RD139
##STR00232## RD140 ##STR00233## RD141 ##STR00234## RD142
##STR00235## RD146 ##STR00236## RD147 ##STR00237## RD154
[0384] In short, novel compounds which show evidence of being far
superior to bicalutamide in treating prostate cancer were
identified and produced.
Sensitivity of Anti-Cancer Activity of Compounds to Structural
Differences
[0385] The inventors have determined that what might appear to be a
small change in the structure of hydantoin compounds may result in
a large change in that compound's performance in treating prostate
cancer. For example, RD161 and RD162 differ only by a single
fluorine substituent on an aryl ring, and RD162 is in Tier 1, while
RD161 is in Tier 2, both being better than bicalutamide for the
treatment of prostate cancer, but RD162 being superior. However,
RD149, which differs from RD 161 only in having an additional
carbon atom between the methylcarbamoyl group and the aryl ring, is
no better than bicalutamide for the treatment of prostate cancer
and is ranked in Tier 4. The effect of RD161, RD162, and RD149 on
luciferase activity can be seen in FIG. 24. At a given
concentration of compound, the luciferase activity upon exposure to
RD161 and RD162 is less than the luciferase activity upon exposure
to RD149.
[0386] RD9 differs from RD8 only in that an amino group is
substituted for a hydroxyl group. However, whereas RD8 is in Tier
1, much better than bicalutamide for the treatment of prostate
cancer, RD9 is in Tier 4, no better than bicalutamide. The effect
of RD8 and RD9 on luciferase activity in the 1AR cell line can be
seen in FIG. 27. For a given dose, the luciferase activity upon
exposure to RD8 is less than the luciferase activity upon exposure
to RD9. The effect of RD8 and RD9 on luciferase activity in the 4AR
cell line can be seen in FIG. 26. For a given dose, the luciferase
activity upon exposure to RD8 is less than the luciferase activity
upon exposure to RD9. The effect of RD8 and RD9 on PSA levels in
the LN/AR cell line can be seen in FIG. 25. For a given dose, the
PSA level upon exposure to RD8 is less than the PSA level upon
exposure to RD9.
[0387] RD130 and RD131 differ from each other only by a methyl
substituent on the end of a carbamoyl group and both compounds are
ranked in Tier 1, although RD131 has been found to be particularly
advantageous. RD129 is the same as RD130, with the exception of a
methoxy group being substituted for an amino group. However, RD129
is ranked in Tier 3. RD128 is similar to RD129, but has one less
carbon in the chain linking the ester group to the aryl ring; RD128
is ranked in Tier 3. The effect of RD130, RD131, RD128, and RD129
on PSA levels in the LN/AR cell line can be seen in FIG. 28. For a
given concentration, the PSA level upon exposure to RD130 and RD131
is less than the PSA level upon exposure to RD128 and RD129.
[0388] RD153 and RD155 differ from each other in that the former
has a methylcarbamoyl group attached to an aryl ring and a dimethyl
substituent attached to the thiohydantoin group, whereas the latter
has a methylamino group attached to the right hand aryl ring and a
cyclobutyl substituent attached to the thiohydantoin group. Whereas
RD153 is in Tier 1, much better than bicalutamide for the treatment
of prostate cancer, RD155 is in Tier 5, inactive or nearly inactive
in the treatment of prostate cancer. The effect of RD153 and RD155
on luciferase activity in the LN/AR cell line can be seen in FIG.
29. For a given concentration, the luciferase activity upon
exposure to RD153 is less than the luciferase activity upon
exposure to RD155.
[0389] RD58 and RD60 differ from each other in the substitution of
a thio for an oxo group and a dimethyl substituent for a cyclobutyl
substituent. Whereas RD58 is in Tier 1, RD60 is in Tier 4.
Pharmaceutical Compositions and Administration
[0390] The compounds of the invention are useful as pharmaceutical
compositions prepared with a therapeutically effective amount of a
compound of the invention, as defined herein, and a
pharmaceutically acceptable carrier or diluent.
[0391] The diarylhydantoin compounds of the invention can be
formulated as pharmaceutical compositions and administered to a
subject in need of treatment, for example a mammal, such as a human
patient, in a variety of forms adapted to the chosen route of
administration, for example, orally, nasally, intraperitoneally, or
parenterally, by intravenous, intramuscular, topical or
subcutaneous routes, or by injection into tissue.
[0392] Thus, diarylhydantoin compounds of the invention may be
systemically administered, e.g., orally, in combination with a
pharmaceutically acceptable vehicle such as an inert diluent or an
assimilable edible carrier, or by inhalation or insufflation. They
may be enclosed in hard or soft shell gelatin capsules, may be
compressed into tablets, or may be incorporated directly with the
food of the patient's diet. For oral therapeutic administration,
the diarylhydantoin compounds may be combined with one or more
excipients and used in the form of ingestible tablets, buccal
tablets, troches, capsules, elixirs, suspensions, syrups, wafers,
and the like. The diarylhydantoin compounds may be combined with a
fine inert powdered carrier and inhaled by the subject or
insufflated. Such compositions and preparations should contain at
least 0.1% diarylhydantoin compounds. The percentage of the
compositions and preparations may, of course, be varied and may
conveniently be between about 2% to about 60% of the weight of a
given unit dosage form. The amount of diarylhydantoin compounds in
such therapeutically useful compositions is such that an effective
dosage level will be obtained.
[0393] The tablets, troches, pills, capsules, and the like may also
contain the following: binders such as gum tragacanth, acacia, corn
starch or gelatin; excipients such as dicalcium phosphate; a
disintegrating agent such as corn starch, potato starch, alginic
acid and the like; a lubricant such as magnesium stearate; and a
sweetening agent such as sucrose, fructose, lactose or aspartame or
a flavoring agent such as peppermint, oil of wintergreen, or cherry
flavoring may be added. When the unit dosage form is a capsule, it
may contain, in addition to materials of the above type, a liquid
carrier, such as a vegetable oil or a polyethylene glycol. Various
other materials may be present as coatings or to otherwise modify
the physical form of the solid unit dosage form. For instance,
tablets, pills, or capsules may be coated with gelatin, wax,
shellac or sugar and the like. A syrup or elixir may contain the
active compound, sucrose or fructose as a sweetening agent, methyl
and propylparabens as preservatives, a dye and flavoring such as
cherry or orange flavor. Of course, any material used in preparing
any unit dosage form should be pharmaceutically acceptable and
substantially non-toxic in the amounts employed. In addition, the
diarylhydantoin compounds may be incorporated into
sustained-release preparations and devices. For example, the
diarylhydantoin compounds may be incorporated into time release
capsules, time release tablets, and time release pills.
[0394] The diarylhydantoin compounds may also be administered
intravenously or intraperitoneally by infusion or injection.
Solutions of the diarylhydantoin compounds can be prepared in
water, optionally mixed with a nontoxic surfactant. Dispersions can
also be prepared in glycerol, liquid polyethylene glycols,
triacetin, and mixtures thereof and in oils. Under ordinary
conditions of storage and use, these preparations can contain a
preservative to prevent the growth of microorganisms.
[0395] The pharmaceutical dosage forms suitable for injection or
infusion can include sterile aqueous solutions or dispersions or
sterile powders comprising the diarylhydantoin compounds which are
adapted for the extemporaneous preparation of sterile injectable or
infusible solutions or dispersions, optionally encapsulated in
liposomes. In all cases, the ultimate dosage form should be
sterile, fluid and stable under the conditions of manufacture and
storage. The liquid carrier or vehicle can be a solvent or liquid
dispersion medium comprising, for example, water, ethanol, a polyol
(for example, glycerol, propylene glycol, liquid polyethylene
glycols, and the like), vegetable oils, nontoxic glyceryl esters,
and suitable mixtures thereof. The proper fluidity can be
maintained, for example, by the formation of liposomes, by the
maintenance of the required particle size in the case of
dispersions or by the use of surfactants. The prevention of the
action of microorganisms can be brought about by various
antibacterial and antifungal agents, for example, parabens,
chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In
many cases, it will be preferable to include isotonic agents, for
example, sugars, buffers or sodium chloride. Prolonged absorption
of the injectable compositions can be brought about by the use in
the compositions of agents delaying absorption, for example,
aluminum monostearate and gelatin.
[0396] Sterile injectable solutions are prepared by incorporating
the diarylhydantoin compounds in the required amount in the
appropriate solvent with various of the other ingredients
enumerated above, as required, followed by filter sterilization. In
the case of sterile powders for the preparation of sterile
injectable solutions, the preferred methods of preparation are
vacuum drying and freeze drying techniques, which yield a powder of
the active ingredient plus any additional desired ingredient
present in the previously sterile-filtered solutions.
[0397] For topical administration, the diarylhydantoin compounds
may be applied in pure form. However, it will generally be
desirable to administer them to the skin as compositions or
formulations, in combination with a dermatologically acceptable
carrier, which may be a solid or a liquid.
[0398] Useful solid carriers include finely divided solids such as
talc, clay, microcrystalline cellulose, silica, alumina and the
like. Other solid carriers include nontoxic polymeric nanoparticles
or microparticles. Useful liquid carriers include water, alcohols
or glycols or water/alcohol/glycol blends, in which the
diarylhydantoin compounds can be dissolved or dispersed at
effective levels, optionally with the aid of non-toxic surfactants.
Adjuvants such as fragrances and additional antimicrobial agents
can be added to optimize the properties for a given use. The
resultant liquid compositions can be applied from absorbent pads,
used to impregnate bandages and other dressings, or sprayed onto
the affected area using pump-type or aerosol sprayers.
[0399] Thickeners such as synthetic polymers, fatty acids, fatty
acid salts and esters, fatty alcohols, modified celluloses or
modified mineral materials can also be employed with liquid
carriers to form spreadable pastes, gels, ointments, soaps, and the
like, for application directly to the skin of the user.
[0400] Examples of useful dermatological compositions which can be
used to deliver the diarylhydantoin compounds to the skin are known
to the art; for example, see Jacquet et al. (U.S. Pat. No.
4,608,392), Geria (U.S. Pat. No. 4,992,478), Smith et al. (U.S.
Pat. No. 4,559,157) and Wortzman (U.S. Pat. No. 4,820,508), all of
which are hereby incorporated by reference.
[0401] Useful dosages of the compounds of formula I can be
determined by comparing their in vitro activity, and in vivo
activity in animal models. Methods for the extrapolation of
effective dosages in mice, and other animals, to humans are known
to the art; for example, see U.S. Pat. No. 4,938,949, which is
hereby incorporated by reference.
[0402] For example, the concentration of the diarylhydantoin
compounds in a liquid composition, such as a lotion, can be from
about 0.1-25% by weight, or from about 0.5-10% by weight. The
concentration in a semi-solid or solid composition such as a gel or
a powder can be about 0.1-5% by weight, or about 0.5-2.5% by
weight.
[0403] The amount of the diarylhydantoin compounds required for use
in treatment will vary not only with the particular salt selected
but also with the route of administration, the nature of the
condition being treated and the age and condition of the patient
and will be ultimately at the discretion of the attendant physician
or clinician.
[0404] Effective dosages and routes of administration of agents of
the invention are conventional. The exact amount (effective dose)
of the agent will vary from subject to subject, depending on, for
example, the species, age, weight and general or clinical condition
of the subject, the severity or mechanism of any disorder being
treated, the particular agent or vehicle used, the method and
scheduling of administration, and the like. A therapeutically
effective dose can be determined empirically, by conventional
procedures known to those of skill in the art. See, e.g., The
Pharmacological Basis of Therapeutics, Goodman and Gilman, eds.,
Macmillan Publishing Co., New York. For example, an effective dose
can be estimated initially either in cell culture assays or in
suitable animal models. The animal model may also be used to
determine the appropriate concentration ranges and routes of
administration. Such information can then be used to determine
useful doses and routes for administration in humans. A therapeutic
dose can also be selected by analogy to dosages for comparable
therapeutic agents.
[0405] The particular mode of administration and the dosage regimen
will be selected by the attending clinician, taking into account
the particulars of the case (e.g., the subject, the disease, the
disease state involved, and whether the treatment is prophylactic).
Treatment may involve daily or multi-daily doses of compound(s)
over a period of a few days to months, or even years.
[0406] In general, however, a suitable dose will be in the range of
from about 0.001 to about 100 mg/kg, e.g., from about 0.01 to about
100 mg/kg of body weight per day, such as above about 0.1 mg per
kilogram, or in a range of from about 1 to about 10 mg per kilogram
body weight of the recipient per day. For example, a suitable dose
may be about 1 mg/kg, 10 mg/kg, or 50 mg/kg of body weight per
day.
[0407] The diarylhydantoin compounds are conveniently administered
in unit dosage form; for example, containing 0.05 to 10000 mg, 0.5
to 10000 mg, 5 to 1000 mg, or about 100 mg of active ingredient per
unit dosage form.
[0408] The diarylhydantoin compounds can be administered to achieve
peak plasma concentrations of, for example, from about 0.5 to about
75 .mu.M, about 1 to 50 .mu.M, about 2 to about 30 .mu.M, or about
5 to about 25 .mu.M. Exemplary desirable plasma concentrations
include at least or no more than 0.25, 0.5, 1, 5, 10, 25, 50, 75,
100 or 200 .mu.M. For example, plasma levels may be from about 1 to
100 micromolar or from about 10 to about 25 micromolar. This may be
achieved, for example, by the intravenous injection of a 0.05 to 5%
solution of the diarylhydantoin compounds, optionally in saline, or
orally administered as a bolus containing about 1-100 mg of the
diarylhydantoin compounds. Desirable blood levels may be maintained
by continuous infusion to provide about 0.00005-5 mg per kg body
weight per hour, for example at least or no more than 0.00005,
0.0005, 0.005, 0.05, 0.5, or 5 mg/kg/hr. Alternatively, such levels
can be obtained by intermittent infusions containing about
0.0002-20 mg per kg body weight, for example, at least or no more
than 0.0002, 0.002, 0.02, 0.2, 2, 20, or 50 mg of the
diarylhydantoin compounds per kg of body weight.
[0409] The diarylhydantoin compounds may conveniently be presented
in a single dose or as divided doses administered at appropriate
intervals, for example, as two, three, four or more sub-doses per
day. The sub-dose itself may be further divided, e.g., into a
number of discrete loosely spaced administrations; such as multiple
inhalations from an insufflator.
[0410] A number of the above-identified compounds exhibit little or
no agonistic activities with respect to hormone refractory prostate
cancer cells. Because these compounds are strong AR inhibitors,
they can be used not only in treating prostate cancer, but also in
treating other AR related diseases or conditions such as benign
prostate hyperplasia, hair loss, and acne. Because AR belongs to
the family of nuclear receptors, these compounds may serve as
scaffolds for drug synthesis targeting other nuclear receptors,
such as estrogen receptor and peroxisome proliferator-activated
receptor. Therefore, they may be further developed for other
diseases such as breast cancer, ovarian cancer, diabetes, cardiac
diseases, and metabolism related diseases, in which nuclear
receptors play a role.
[0411] The embodiments illustrated and discussed in this
specification are intended only to teach those skilled in the art
the best way known to the inventors to make and use the invention.
Nothing in this specification should be considered as limiting the
scope of the present invention. All examples presented are
representative and non-limiting. The above-described embodiments of
the invention may be modified or varied, without departing from the
invention, as appreciated by those skilled in the art in light of
the above teachings. It is therefore to be understood that, within
the scope of the claims and their equivalents, the invention may be
practiced otherwise than as specifically described.
* * * * *