Scholarly article on 4-[3-(4-methylphenyl)-4-oxo-2-thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile 1220429-79-7 from Journal of Medicinal Chemistry p. 2779

J. Med. Chem. 2010, 53, 2779–2796 2779

DOI: 10.1021/jm901488g

Structure-Activity Relationship for Thiohydantoin Androgen Receptor Antagonists for

Castration-Resistant Prostate Cancer (CRPC)

Michael E. Jung,*^,†Samedy Ouk,^†Dongwon Yoo,^†Charles L. Sawyers,^‡,§Charlie Chen,^‡Chris Tran,^‡and John Wongvipat^‡,§

^†Department of Chemistry and Biochemistry, and ^‡Department of Medicine, University of California, Los Angeles, 405 Hilgard Avenue,

Los Angeles, California 90095, and ^§Human Oncology and Pathogenesis Program, Howard Hughes Medical Institute, Memorial Sloan Kettering

Cancer Center, 1275 York Avenue, New York, New York 10065

Received October 7, 2009

A structure-activity relationship study was carried out on a series of thiohydantoins and their

analogues 14 which led to the discovery of 92 (MDV3100) as the clinical candidate for the treatment

of hormone refractory prostate cancer.

Scheme 1

Introduction

Although prostate cancer can be initially treated with either

castration or androgen receptor (AR^a) antagonists such as

bicalutamide 1, nilutamide 2, and flutamide 3a (which is

oxidized to the active metabolite hydroxyflutamide 3b), after

a period of approximately 2-4 years, the cancer becomes

resistant to such treatment (Scheme 1).¹Indeed in this castra-

tion resistant stage (formerly called hormone refractory or

“androgen-independent”), former AR antagonists such as

bicalutamide become partial agonists and their use in cancer

treatment must be discontinued. Sawyers and co-workers

showed that a 3- to 5-fold up-regulation of the androgen

receptor was the likely cause of the resistance to anti-andro-

gens.²They further demonstrated that castration resistant

prostate cancer was still dependent on the ligand binding

domain of AR for growth.²Therefore, we began a research

program aimed at the identification of novel chemical struc-

tures that would be potent androgen receptor antagonists,

especially in its up-regulated state in castration resistant

disease, without any significant agonist effect. We report here

the results of our structure-activity relationship (SAR) study

that led to the choice of 92 as the lead candidate for the

treatment of castration-resistant prostate cancer (CRPC).

This compound, named MDV3100, has completed phase

1-2 clinical trials and has now entered a phase 3 randomized

trial for drug registration.^3,4

Scheme 2

We examined the literature on the binding of various

compounds to the AR⁵and the available crystal structures

of the AR⁶(there were only structures of the AR with

compounds in an agonist binding mode)⁷and binding calcu-

lations.⁸We decided to begin with the structure of one of the

strongest known binders to the AR, namely, the nonsteroidal

AR agonist RU59063 4, the affinity of which for the AR is

nearly equalto thatof thewell-knownsteroidalagonist R1881

5, both of which are slightly higher than that of the natural

ligand dihydrotestosterone 6 (DHT) (Scheme 2).⁹Our plan

was to vary systematically the structural units of this strong-

binding agonist to see if we could obtain a reasonably strong-

binding antagonist. We prepared several series of compounds

in which each of the functional groups of this molecule was

varied, and we measured the binding affinity and both the

agonism and antagonism of each.

*To whom correspondence should be addressed. Phone: (310) 825-

7954. Fax: (310) 206-3722. E-mail: jung@chem.ucla.edu.

^aAbbreviations: AR, androgen receptor; CRPC, castration-resistant

prostate cancer; DHT, dihydrotestosterone; DMSO, dimethyl sulfoxide;

FBS, fetal bovine serum; HR, hormone refractory; PK, pharmacoki-

netic; PSA, prostate specific antigen; SAR, structure-activity relation-

ship; SCID, severe combined immunodeficient.

Synthesis

The syntheses of the compounds varied somewhat but

usually involved three general routes. The first (Scheme 3)

r

2010 American Chemical Society

Published on Web 03/10/2010

pubs.acs.org/jmc

2780 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

Scheme 3

several 4-halo aromatic systems 16, e.g., X = F, Z = CN,

NO₂, etc., to give the 4-substituted phenylthiohydantoins 17.

Finally several additional analogues 19 could be prepared by

diazotization of 4-aminophenylthiohydantoins 18 and sub-

stitution with various groups, e.g., halogens, cyano, etc.

(Scheme 5).

Testing Methods

Several systems were utilized to test the activity of the

analogues. We used a prostate specific antigen (PSA) expres-

sion readout for normal LNCaP (hormone sensitive) cells and

in LNCaP/AR cells, which were engineered (using viral

infection with a cDNA encoding for the AR) to express

3- to 5-fold higher levels of the AR to mimic the clinical

setting of CRPC.⁴Tests in LNCaP cells were carried out in the

presence of fetal bovine serum (FBS), whereas tests in

LNCaP/AR cells were carried out in charcoal stripped serum

to mimic the androgen depleted, castration resistant state. We

also developed a luciferase reporter system utilizing ARR₂PB-

Luc, a piece of plasmid DNA that encodes firefly luciferin

with AR binding sites in the natural promoter for probasin of

rat prostate, which provides an easy quantitative assay for AR

activity as a transcription factor.

Scheme 4

Scheme 5

Structure-Activity Relationship

The first set of analogues prepared were analogues with

azidoalkyl and azidoaryl groups at N1, 20-24 and 25,¹⁰with

the hope that the small polar azide group might mimic the

hydroxyl in 4 and give good binding. The activity vs normal

LNCaP (hormone sensitive) cells was measured as relative

prostate specific antigen (PSA) level vs vehicle (DMSO) and

using bicalutamide as a standard for antagonist activity in this

Scheme 6

Scheme 7

was a triply convergent process involving first a Strecker

reaction of a substituted amine or aniline 7 with a ketone 8

and trimethylsilyl cyanide (or the preformed cyanohydrin 9)

to generate the cyanoamine 10. The third component, the

isothiocyanate 12, prepared usually in quantitative yield from

the amine 11, was added to 10 to give the thiohydantoin-4-

imine 13 (in which the group on the imine nitrogen could be

either hydrogen of a thiocarbamoyl group derived from a

second equivalent of the isothiocyanate). Hydrolysis of 13

afforded the desired thiohydantoins 14. A second general

method of synthesis (Scheme 4) utilized an N1-unsubstituted

thiohydantoin 15 (prepared from the ketone 8 with ammo-

nium cyanide and hydrolysis) which was added to any of

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2781

Scheme 8

Scheme 9

androgen-dependent assay (Scheme 6). It can be seen that all

six compounds had activity better than bicalutamide itself but

that 25 (the 4-azidophenyl compound) was the best of this

group. We next varied the group at the 4-position of the N1-

phenyl ring, and again all of the analogues, 25-29,¹¹were

active (Scheme 7), both by the luciferase reporter assay and by

relative PSA level. We then kept a methyl group as the

substituent at the 4-position of the phenyl ring and varied

the alkyl groups in the thiohydantoin ring from hydrogen to

methyl, ethyl, and propyl. The activity was measured as

relative luciferase activity vs bicalutamide and 27 as standards

(Scheme 8). All of these derivatives were much less active than

the dimethyl compound 27, without significant differences

between the hydrido analogues 30-33 and the dialkyl analo-

gues 34 and 35.¹¹Analogues with the alternative arrangement

of the dialkyl substituents and the carbonyl (equivalent to

switching the nitrogen substituents), 36 and 37,¹¹also had

reduced activity relative to 27. However, there was little

difference in activity among the analogues. We next prepared

a set of analogues (Scheme 9) featuring cycloalkyl substituents

on the thiohydantoin ring, all of which were made from the

corresponding ketones. Their activity was measured by the

relative PSA expression levels vs bicalutamide and 27 as

standards. All of these derivatives showed good activity with

the cyclobutyl and cyclopentyl analogues 38 and 39,¹¹being

comparable to the dimethyl analogue 27. The six-, seven-, and

eight-membered rings, 40-42,¹¹were slightly less active. The

spiro N-methylpiperidine analogue 43¹¹was distinctly less

active, which may be due to the fact that the nitrogen would

likely be charged at cellular pH. We also tested several other

aromatic rings and substitution patterns on the N1-aryl sub-

stituent (Scheme 10) using 27 as the standard. A compound

with a methyl group at the 2-position (ortho to the thiohy-

dantoin nitrogen), 44,¹¹was active while one with a charged

carboxylic acidgroupatthe 2-position, 45,¹¹was inactive. The

compound with chloromethyl groups at C5, 46,¹¹had de-

creased activity, while the analogue with a fluoromethyl

group, 47,¹¹had good activity. Other functional groups were

placed at the 4-position of the N1-phenyl substituent, and

their activity was assayed using a relative PSA readout vs

standards (Scheme 11). Nearly all of the substituents showed

good activity when compared to the best molecules in their

series; e.g., the 4-phenyl and 4-hydroxy analogues 48 and 49¹¹

were very similar in activity to 38. Similarly the 4-cyano and

4-nitro analogues, 50 and 51,¹¹were nearly as active as the

Scheme 10

methylanalogue 39, whilethe 4-trifluoromethyl analogue52¹¹

was slightly more active than the methyl analogue 27. Thus,

the 4-position of the 1-phenyl ring can bear many different

substituents without losing activity. We also prepared and

tested various imine and thione analogues of the active series

using a relative PSA readout vs 27 and 38 as standards

(Scheme 12). In all cases the thiohydantoins with the thiocar-

bonyl at C2 and the carbonyl at C4 were the most active

although the imines, 53 and 54,¹¹were nearly as active as the

parent compounds 27 and 38 while the dithiohydantoin and

2782 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

Scheme 11

Scheme 12

Scheme 13

the hydantoin analogues, 55 and 56,¹¹were not as active as the

parent thiohydantoin 28.

Other imine derivatives, e.g., 57 and 60,¹¹were also some-

what active as shown in Scheme 13, using both relative PSA

levels and the luciferase assay vs 27 and 38 as standards. The

N-thiocarbamoyl analogues 58-62¹¹were prepared by using

an excess of the isothiocyanate, e.g., 12, in the coupling reac-

tion with the corresponding aniline. They were all active, but

they are hydrolyzed to the corresponding thiohydantoins

under cellular conditions. The condensation of the cyano-

amine 10 with the isothiocyanate 12 gave a small amount of a

new chemical entity, namely, the thiazolidine-2,5-dimines, in

which the 5-imino anion bears a thiocarbamoyl group. These

compounds are presumably formed by attack of the sulfur

atom, rather than the nitrogen atom, of the thioamide anion

on the nitrile of the cyanoamine with subsequent trapping of

that imine anion with another equivalent of the isothiocya-

nate. Two of these new analogues, 63 and 64,¹¹were tested for

activity using both the relative PSA levels and the luciferase

reporter assay (Scheme 14). Both of these new compounds

showed excellent activity comparable to their parent thiohy-

dantoin analogues 27 and 38. As far as we can tell, this is the first

time any compounds with such a structure have been shown

to have antiandrogen activity. Therefore, these compounds

represent a new structural class of antiandrogens. Since the

activity of these two new compounds are extremely similar to

their thiohydantoin counterparts in both the PSA and luciferase

assay systems, it is interesting to speculate whether they are

being converted, under the cellular testing conditions, into the

thiohydantoins. That would require cleavage of the imi-

nothiourea to the imine, opening of the ring via reformation

of the nitrile with ejection of the thiolate anion, and then

recyclization of the thioamide anion on to the nitrile via the

nitrogen atom. Finally in our early set of compounds, we found

several that were essentially devoid of activity (Scheme 15).

Various derivatives of the nitrile, e.g., the amide 65,¹¹

were inactive as were various analogues in which the ortho

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2783

Scheme 14

Scheme 15

Figure 2. Dose response in change in tumor volume of xenografts

with 38.

Figure 3. Serum concentration of 38 after iv injection; n = 3 mice.

Scheme 16

trifluoromethyl group was replaced by halogens, 66-69.¹¹

Acyclic analogues, e.g., the chloromethylamide 70,¹¹were

Figure 1. Fold change in tumor volume of xenografts with bical,

27, and 38 (10 mg/kg).

2784 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

Figure 4. PK curves of 38 and 80.

Scheme 17

inactive as was the 4-oxooxazolidine-2-thione 71.¹¹Com-

pounds with a spacer group between the ring nitrogen and

the aryl group, e.g., the arylsulfonylamide 72 and the benzhy-

dryl analogue 73,¹¹were inactive.

all of which showed good activity in the PSA secretion

assay. The two benzylic alcohol analogues 74 and 76 as well

as the aldehyde 75 exhibited IC₅₀values of 200-300 nM but

were considerably more polar than 38. The extended amide

and alcohol analogues 77 and 78 were even more active with

IC₅₀values of 100-150 nM. A series of analogues with

extended chains and heterocyclic units were prepared, and

their activity using PSA levels in the hormone refractory cell

line LNCaP/AR was evaluated in vitro (Scheme 17). All the

derivatives, with the exception of the (hydroxyethyl)amide

and piperazine analogues 83 and 84, showed good activity,

especially compared to bicalutamide which is inactive in this

hormone-refractory assay. The most active compound was

the N-methylbutyramide analogue 80, which was determined

to have an IC₅₀of 92 nM with a clogP of 3.44. But as the data

show, several other analogues were also quite active with the

N-methylamides being generally more active than the amides

themselves. The corresponding esters and acids were also

preparedaswell as the analogousphenylacetamidederivatives

(two-carbon chain), but the activity of all of these was weaker

(data not shown). Although 80 showed excellent activity, its

PK was also poor (Figure 4), although it was somewhat more

available than the earlier analogue 38. Although hydrolysis of

the N-methylamide to the acid was seen, we postulated that

one reason for the low serum concentration of both 38 and 80

was metabolism via oxidation of the electron-rich aromatic

ring. To try to eliminate this problem, we decided to prepare

While carrying out this SAR study using in vitro assays, we

also decided to test the in vivo activity of lead compounds in

animalsto gauge theirpharmacologic properties and abilityto

impair growth of castration-resistant prostate cancer xeno-

graft models that are also resistant to bicalutamide. Therefore

the ability of compounds 27 and 38 to decrease the growth of

LAPC4/AR cells or LNCaP/AR cells grown as xenografts in

castrate SCID mice was assayed. In a pilot experiment using

the LAPC4/AR xenograft model (Figure 1),¹²both com-

pounds were more effective than bicalutamide with 38 being

slightly superior, with an IC₅₀value of 124 nM for inhibition

of PSA secretion. This thiohydantoin 38 also showed a good

dose response in the castration-resistant xenograft assay

(Figure 2). However, 38 had a short half-life with a very rapid

clearance as shown in Figure 3. This was likely due to both a

rapid metabolism (hydroxylation of the aromatic methyl

group) and its relatively high clogP value of 4.20 (compared

to 2.91 for bicalutamide). Therefore, we decided to prepare

additional analogues of 38 that would be more polar. In

particular we decided to change the substituents on the aryl

ring attached to N1, especially at the 4-position, to see if more

polar and more stable analogues could be prepared. There-

fore, several simple analogues of 38 were prepared (Scheme 16),

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Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2785

Scheme 18

Figure 5. Concentration of 91 after iv (blue) or oral (pink) admin-

istration.

Scheme 19

Figure 6. Effect of bicalutamide and 91 on LNCaP/AR (HR)

tumor size at 10 and 50 mg/kg once a day.

analogues that had the electron-withdrawing group attached

directly to the aromatic ring (Scheme 18), which yielded several

very active compounds based on evaluation in the hormone

refractory LNCaP/AR assay. Thus, the sulfone 86, the ester 87,

and the two amides 88 and 89 showed good activity as did the

fluorophenol 90. However, we found that the 3-fluoroamide

analogue 91 (also called RD162)^4,13had not only excellent

activity (Scheme 19) but also a superb pharmacokinetic (PK)

profile. Its IC₅₀was 122 nM, and it had a clogP of 3.20. However,

the measure of its excellent PK profile was its steady state

concentration as shown in Table 1. Compound 91 has almost

the exact same exposure after a 10 mg/kg dose as bicalutamide,

e.g., 9.9 mM vs 10 mM. And the IC₅₀of 91 is nearly 8 times lower

than that of bicalutamide, 122 nM vs 1 mM.

Figure 7. Dose response in tumor volume change of xenografts

with 91 at 0.1, 1, and 10 mg/kg once a day.

The concentration of 91 after iv and oral administration is

shown in Figure 5. With this excellent PK profile, we decided

to choose 91 as our lead drug candidate. Its activity on

LNCaP/AR (HR) tumor size at 10 and 50 mg/kg once a day

Figure 8. Effect of change in tumor volume of xenografts with 91 at

10 mg/kg once a day.

Table 1

compd

IC₅₀(nM)

clogP

C_ss10 mpk (mM)

vs bicalutamide at the same dose (Figure 6) shows it to be very

active. It is cytostatic atthese doses. The dose response of 91 in

LNCaP xenografts¹⁴(Figure 7) shows that at least 1 (mg/kg)/

day is required and that 10 (mg/kg)/day is optimal. We also

assayed the activity of 91 on LNCaP xenografts over an

bicalutamide (1)

1000

124

92

2.91

4.20

3.44

3.20

10.0

NA

0.39

9.9

38

80

91

122

2786 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

Scheme 20. Effect of 38, 80, and 91 on Hormone Sensitive

LNCaP Cells

Figure 9. Dose response study of 91 and 92 (nM) on castration

resistant LNCaP AR cells.

Scheme 21

Conclusion

We have described the structure-activity relationship

study that led to the choice of 92 as a clinical candidate for

the treatment of castration-resistant prostate cancer. Many

analogous diarylthiohydantoins in this series showed good

androgen receptor antagonism with essentially no agonism,

but the pharmacokinetic properties of 91 and its close analo-

gues 92 and 93 led to the choice of 92 as the clinical candidate.

Experimental Section

General. All reactions were carried out under an argon atmo-

sphere unless otherwise specified. Tetrahydrofuran (THF) and

diethyl ether were distilled from benzoquinone ketyl radical

under an argon atmosphere. Dichloromethane, toluene, benzene,

pyridine, triethylamine, and diisopropylethylamine (DIPEA)

were distilled from calcium hydride under an argon atmosphere.

Dimethyl sulfoxide (DMSO) was distilled over calcium hydride

˚

and stored over 4 A molecular sieves. All other solvents or

reagents were purified according to literature procedures. ¹H

NMR and ¹³C NMR spectra were obtained on ARX-400,

ARX-500, or Avance-500 spectrometers. The chemical shifts are

reported in parts per million (ppm, δ). The coupling constants are

reported in hertz (Hz), and the resonance patterns are reported

with notations as the following: br (broad), s (singlet), d (double),

t (triplet), q (quartet), and m (multiplet). Infrared spectra were

recorded on Nicolet 501 or Nicolet AVATAR 370 instrument

using liquid films (neat) or in CDCl₃solution on NaCl plates, and

only the significant absorption bands are recorded (in cm^-1).

Thin-layer chromatography (TLC) was carried out using pre-

coated silica gel sheets (Merck 60 F₂₅₄). Visual detection was

performed with ultraviolet light, p-anisaldehyde stain, potassium

permanganate stain, or iodine. Flash chromatography was per-

formed using SilicaFlash P60 (60 Α, 40-63 μm) silica gel from

SiliCycle, Inc., with compressed air. HPLC was performed on a

Waters HPLC using either a C18 reverse phase column or a

normal silica gel column as appropriate. The purity of all final

compounds was established to be at least 95% pure by a combina-

tion of TLC R_fvalues in several solvent systems and HPLC.

Additionally the absence of any extraneous peaks in the proton

NMR spectrum confirmed the high level of purity.

Synthesis of 20-24. 4-Isothiocyanato-2-trifluoromethylben-

zonitrile, 12a. 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 21 °C. Stirring was continued for an additional

1 h. The reaction medium was extracted with chloroform (3 ꢀ

15 mL). The combined organic phase was dried over MgSO₄and

evaporated to dryness under reduced pressure to yield desired

product as brownish solid and was used as such for the next step

(2.72 g, 11.9 mmol, 99%).

extended period (Figure 8) which showed that it retains

activity at 10 (mg/kg)/day for 31 days. Since 91 was initially

screened in hormone-refractory models, we also looked at its

effect in hormone sensitive cells¹⁵(Scheme 20) and found good

activity vs LNCaP cells, albeit not as good as 80. But this relative

liability is counterbalanced by its excellent PK properties. Thus,

it is possible that 91 might be able to be used for treatment of

both types of prostate cancer, hormone sensitive and castration-

resistant, but one must wait for data from clinical trials.

Several additional analogues of both 80 and 91 were

prepared and tested (Scheme 21). We made both the dimethyl-

amide and the nitrile analogues of 80 (94 and 95,¹³res-

pectively) in order to try to identify a strong-binding analogue

with better PK and, in particular, a longer half-life. Both of

these compounds had quite good activity compared to earlier

compounds. Two additional analogues of 91 were prepared

and tested for their activity on castration resistant prostate

cancer, namely, the analogues with the cyclobutyl unit replaced

by dimethyl unit, 92, and cyclopentyl unit, 93.¹³Both of these

new analogues were very active in hormone-refractory LNCaP/

AR with essentially the same activity as 91. A dose-response

study (Figure 9) showed 92 to be a little more active than

91. Since the dimethyl analogue 92 offers the great advantage

of an inexpensive starting material, acetone or its cyanohydrin,

for its production, it was chosen as the drug candidate and

subjected to metabolic stability, toxicology, and further animal

studies. This compound 92 (also called RD162⁰) was licensed

by Medivation, Inc. It has now entered phase 3 clinical trials

for the treatment of castration-resistant prostate cancer.⁴

Further details on this compound will be reported in due

course.

1,4-Diazidobutane, 20a. To a mixture of 1,4-dibromobutane

(21.6 g, 100 mmol) in DMF (100 mL) was added an aqueous

solution of sodium azide (13.65 g, 210 mmol in 50 mL of water).

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2787

The mixture was stirred and heated to 80 °C for 20 h, and then

the medium was washed with brine (200 mL) and extracted with

hexane (3 ꢀ 300 mL). The combined organic layer was dried

over MgSO₄and concentrated to yield 1,4-diazidobutane as a

liquid (13.72 g, 9.8 mmol, 98%).

4-Azidobutylamine, 20b. To a mixture of 1,4-diazidobutane

20a (4.20 g, 30 mmol), aqueous 1 M HCl (60 mL), diethyl ether

(20 mL), and ethyl acetate (20 mL) cooled to 0 °C was added

triphenylphosphine portionwise during 1 h. The mixture was

warmed to 21 °C and stirred for an additional 20 h, and then the

organic layer was separated from the aqueous layer. The aqueous

phase was washed with ethyl ether (2 ꢀ 50 mL) to remove

triphenylphosphine oxide residual. The aqueous phase was basi-

fied to a pH 13 by aqueous NaOH and then was extracted with

dichloromethane (3 ꢀ 100 mL). The combined dichloromethane

layer was dried over MgSO₄and concentrated to yield 4-azido-

butylamine (2.74 g, 24 mmol, 80%) as a liquid.

2-(4-Azidobutylamino)-2-methylpropionitrile, 20c. A mixture

of 4-azidobutylamine 20b (0.57 g, 5 mmol), acetone cyanohydrin

(0.425 g, 5 mmol), and Na₂SO₄(0.2 g) was stirred at 21 °C for

12 h. The mixture was diluted with hexane and filtered off. The

filtrate was concentrated to yield 2-(4-azidobutylamino)-2-

methylpropionitrile (0.896 g, 4.95 mmol, 99%) as a liquid.

4-[3-(4-Azidobutyl)-5-imino-4,4-dimethyl-2-thioxoimidazoli-

din-1-yl]-2-trifluoromethylbenzonitrile, 20d. A mixture of iso-

thiocyanate 12a (0.684 g, 3 mmol), 20c (0.543 g, 3 mmol), and

triethylamine (0.04 g, 0.4 mmol) in THF (6 mL) was refluxed for

1 h. The medium was concentrated and chromatographed

(dichloromethane/acetone, 6:1) to obtain 20d (0.834 g, 2.04

mmol, 68%) as an off-white solid.

4-[3-(4-Azidobutyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-

1-yl]-2-trifluoromethylbenzonitrile, 20. A mixture of 20d (0.818

g, 2.0 mmol), aqueous 1 M HCl (5 mL), and methanol (20 mL)

was heated to reflux for 1 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (25 mL) and

extracted with dichloromethane (50 mL). The organic layer

was dried over MgSO₄, concentrated, and chromatographed

(dichloromethane/acetone, 9:1) to yield 20 (0.803 g, 1.96 mmol,

98%) as an off-white solid. ¹H NMR (400 MHz, CDCl₃) δ 1.58

(s, 6H), 1.63-1.71 (m, 2H), 1.88-1.96 (m, 2H), 3.37 (t, J = 6.6

Hz, 2H), 3.71 (t, J = 8.1 Hz, 2H), 7.77 (dd, J = 8.2, 1.8 Hz, 1H),

7.89 (d, J = 1.8 Hz, 1H), 7.94 (d, J = 8.2 Hz, 1H); ¹³C NMR

(100 MHz, CDCl₃) δ 23.2, 25.5, 26.4, 43.7, 50.9, 65.1, 109.9,

114.9, 121.9 (q, J = 272.6 Hz), 127.0 (q, J = 4.9 Hz), 132.1,

133.4 (q, J = 33.0 Hz), 135.1, 137.1, 175.2, 178.4.

The same procedure was applied for the synthesis of 21-24.

4-[3-(3-Azidopropyl)-4,4-dimethyl-5-oxo-2-thioxoimidazoli-

din-1-yl]-2-trifluoromethylbenzonitrile, 21. ¹H NMR (400 MHz,

CDCl₃) δ 1.51 (s, 6H), 2.01-2.08 (m, 2H), 3.38 (t, J = 6.4 Hz,

2H), 3.71 (t, J = 7.8 Hz, 2H), 7.74 (dd, J = 8.2, 1.8 Hz, 1H), 7.87

(d, J = 1.8 Hz, 1H), 7.89 (d, J = 8.2 Hz, 1H); ¹³C NMR (100

MHz, CDCl₃) δ 22.8, 27.4, 41.6, 49.0, 65.2, 109.6, 114.9, 120.0

(q, J = 272.4 Hz), 127.0 (q, J = 4.9 Hz), 132.3, 132.9 (q, J = 33.0

Hz), 135.2, 137.3, 175.1, 178.5.

4-[3-(5-Azidopentyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-

yl]-2-trifluoromethylbenzonitrile, 22. ¹H NMR (400 MHz, CDCl₃)

δ 1.44-1.50 (m, 2H), 1.56 (s, 6H), 1.61-1.67 (m, 2H), 1.80-1.86

(m, 2H), 3.27 (t, J = 6.7 Hz, 2H), 3.67 (t, J = 8.2 Hz, 2H), 7.77

(dd, J = 8.2, 1.8 Hz, 1H), 7.88 (d, J = 1.8 Hz, 1H), 7.92 (d, J = 8.2

Hz, 1H); ¹³C NMR (100 MHz, CDCl₃) δ 23.1, 24.2, 27.6, 28.3,

44.0, 51.2, 65.1, 109.8, 114.9, 121.9 (q, J = 272.5 Hz), 127.0 (q, J =

4.9 Hz), 132.2, 133.2 (q, J = 33.0 Hz), 135.1, 137.2, 175.2, 178.3.

4-[3-(6-Azidohexyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-

1-yl]-2-trifluoromethylbenzonitrile, 23. ¹H NMR (400 MHz,

CDCl₃) δ 1.31-1.41 (m, 4H), 1.51 (s, 6H), 1.52-1.59 (m, 2H),

1.74-1.81 (m, 2H), 3.20 (t, J = 6.7 Hz, 2H), 3.62 (t, J = 8.2 Hz,

2H), 7.75 (dd, J = 8.2, 1.8 Hz, 1H), 7.88 (d, J = 1.8 Hz, 1H), 7.90 (d,

J=8.2Hz,1H);¹³C NMR (100 MHz, CDCl₃) δ22.9, 26.1, 26.5, 27.8,

28.6, 44.1, 51.2, 65.1, 109.5, 114.9, 122.0 (q, J = 272.5 Hz), 127.0 (q,

J = 4.9 Hz), 132.2, 133.2 (q, J = 33.0 Hz), 135.1, 137.3, 175.2, 178.1.

4-[3-(7-Azidoheptyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-

yl]-2-trifluoromethylbenzonitrile, 24. ¹H NMR (400 MHz, CDCl₃)

δ 1.30-1.42 (m, 6H), 1.53 (s, 6H), 1.54-1.59 (m, 2H), 1.74-1.81

(m, 2H), 3.21(t, J= 6.7 Hz, 2H), 3.64 (t, J= 8.2 Hz, 2H), 7.76 (dd,

J = 8.2, 1.8 Hz, 1H), 7.88 (d, J = 1.8 Hz, 1H), 7.91 (d, J = 8.2 Hz,

1H); ¹³C NMR (100 MHz, CDCl₃) δ 23.0, 26.5, 26.6, 26.8, 27.9,

28.6, 44.2, 51.3, 65.1, 109.6, 114.9, 122.0 (q, J = 272.5 Hz), 127.0 (q,

J= 4.9 Hz), 132.2, 133.2 (q, J= 33.0 Hz), 135.1, 137.3, 175.3, 178.1.

Synthesis of 26. 2-Methyl-2-phenylaminopropanenitrile, 26a.

A mixture of aniline (0.931 g, 10 mmol) and acetone cyanohy-

drin (2 mL) was heated to reflux and stirred for 20 h. After being

cooled to 21 °C, the reaction mixture was poured into ethyl

acetate (40 mL) and washed with cold water (2 ꢀ 30 mL). The

organic layer was dried over MgSO₄and concentrated under

vacuum to dryness to yield 26a (1.51 g, 9.4 mmol, 94%) as a

brown slurry liquid.

4-[3-Phenyl-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-1-yl]-2-

trifluoromethylbenzonitrile, 26. A mixture of isothiocyanate 12a

(0.274 g, 1.2 mmol) and 26a (0.160 g, 1 mmol) in DMF (0.2 mL)

was stirred for 48 h. To this mixture were added methanol

(10 mL) and 2 N HCl (3 mL). The second mixture was refluxed

for 6 h. After being cooled to 21 °C, the reaction mixture was

poured into cold water (20 mL) and extracted with ethyl acetate

(20 mL). The organic layer was dried over MgSO₄, concen-

trated, and chromatographed (dichloromethane) to yield 26

(0.276 g, 0.71 mmol, 71%) as a white powder. ¹H NMR (CDCl₃,

400 MHz) δ 1.60 (s, 6H), 7.28-7.31 (m, 2H), 7.50-7.58 (m, 3H),

7.85 (dd, J = 8.3, 1.8 Hz, 1H), 7.96-7.99 (m, 2H); ¹³C NMR

(CDCl₃, 100 MHz) δ 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.

Synthesis of 27. 2-Methyl-2-(4-methylphenyl)aminopropane-

nitrile, 27a. A mixture of p-toluidine (1.07 g, 10 mmol) and

acetone cyanohydrin (10 mL) was heated to 80 °C and stirred for

4 h. The medium was concentrated and dried under vacuum to

yield 27a (1.72 g, 9.9 mmol, 99%) as a brown solid.

4-[3-(4-Methylphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazoli-

din-1-yl]-2-trifluoromethylbenzonitrile, 27. A mixture of isothio-

cyanate 12a (0.547 g, 2.4 mmol) and 27a (0.348 g, 2 mmol) in

DMF (0.6 mL) was stirred for 36 h. To this mixture were added

methanol (20 mL) and 2 N HCl (5 mL). The second mixture was

refluxed for 6 h. After being cooled to 21 °C, the reaction

mixture was poured into cold water (30 mL) and extracted with

ethyl acetate (40 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane) to yield

27 (0.596 g, 1.48 mmol, 74%) as a white powder. ¹H NMR

(CDCl₃, 400 MHz) δ 1.61 (s, 6H), 2.44 (s, 3H), 7.17-7.20 (m,

2H), 7.33-7.36 (m, 2H), 7.86 (dd, J = 8.3, 1.8 Hz, 1H), 7.96-

7.98 (m, 2H); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

Synthesis of 28. 2-(4-Hydroxyphenylamino)-2-methylpropa-

nenitrile, 28a. A mixture of 4-aminophenol (1.09 g, 10 mmol),

acetone cyanohydrin (10 mL), and MgSO₄(2 g) was heated to

80 °C and stirred for 4 h. After concentration of the reaction

medium under vacuum, compound 28a was crystallized from

water (20 mL). The solid was filtered and dried to yield 28a

(1.69 g, 9.6 mmol, 96%).

4-[3-(4-Hydroxyphenyl)-5-imino-4,4-dimethyl-2-thioxoimida-

zolidin-1-yl]-2-trifluoromethylbenzonitrile, 28b. Triethylamine

(0.101 g, 1 mmol) was added to a solution of isothiocyanate

12a (0.456 g, 2 mmol) and 28a (0.352 g, 2 mmol) in THF (5 mL).

The reaction mixture was stirred at 0 °C for 48 h and then

concentrated to yield a dark residue which was subjected to flash

chromatography (dichloromethane/acetone, 85:15) to afford

28b (0.274 g, 0.68 mmol, 34%).

4-[3-(4-Hydroxyphenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazo-

lidin-1-yl]-2-trifluoromethylbenzonitrile, 28. A mixture of 28b

(0.202 g, 0.5 mmol) in aqueous 2 N HCl (2 mL) and methanol

2788 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

(5 mL) was heated to reflux for 2 h. After being cooled to 21 °C,

the reaction mixture was poured into cold water (10 mL) and

extracted with ethyl acetate (10 mL). The organic layer was dried

over MgSO₄, concentrated, and chromatographed (dichloro-

methane/acetone, 9:1) to yield 28 (0.198 g, 0.49 mmol, 98%) as a

white powder. ¹H NMR (CDCl₃, 400 MHz) δ 1.57 (s, 6H), 6.26

(s, OH), 6.90-6.93 (m, 2H), 7.11-7.14 (m, 2H), 7.84 (dd, J =

8.3, 1.8 Hz, 1H), 7.95-7.98 (m, 2H); ¹³C NMR (CDCl₃, 100

MHz) δ 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.

Synthesis of 29. 4-Aminophenyl)carbamic Acid tert-Butyl Es-

ter, 29a. 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 a mixture of THF

(30 mL) and DMF (10 mL). To this mixture was added di-tert-

butyl pyrocarbonate, Boc₂O (2.18 g, 10 mmol), dropwise over

0.5 h. The reaction mixture was stirred for an additional 4 h at 21

°C. The mixture was then poured into cold water (40 mL) and

extracted with chloroform (3 ꢀ 50 mL). The combined organic

phase was dried over MgSO₄and concentrated to yield a brown

residue which was subjected to flash chromatography (dichloro-

methane/acetone, 4:1) to afford 29a as a yellow solid (1.98 g, 9.5

mmol, 95%) (yield based on Boc₂O).

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 C₁₉H₁₃F₃N₆OS, calculated 430.4, found 430.1.

Synthesis of 30-37. N-(4-Cyano-3-trifluoromethylphenyl)-

N⁰-(4-methylphenyl)thiourea, 30a. Addition of a solution of

4-methylaniline in DMF to a solution of the isothiocyanate

12a in DMF solution at 21 °C for 1 h afforded the desired

unsymmetrical thiourea 30a in 99% yield.

4-[3-(4-Methylphenyl)-5-oxo-2-thioxoimidazolidin-1-yl]-2-tri-

fluoromethylbenzonitrile, 30. Treatment of a solution of the

thiourea 30a in THF at 0 °C with a solution of chloroacetyl

chloride in THF for 1 h gave a 95% yield of a 1:1 mixture of the

two thiohydantoins, the 4-oxo and the 5-oxo regioisomers 30

and 30b. These were easily separated by column chromatogra-

phy, using silica gel (dichloromethane), to give pure 30.

4-[3-(4-Methylphenyl)-4-methyl-5-oxo-2-thioxoimidazolidin-1-

yl]-2-trifluoromethylbenzonitrile, 31. To a suspension of NaH in

THF cooled to 0 °C was added the unsubstituted thiohydantoin

30, and the mixture was stirred for 30 min. A solution of methyl

iodide in THF was added and the mixture stirred for 5 h. Normal

aqueous workup and extraction afforded the crude product

which was purified by column chromatography, using silica gel

(dichloromethane), to give in 5% yield the pure 4-methylthiohy-

dantoin 31.

4-[3-(4-Methylphenyl)-4-ethyl-5-oxo-2-thioxoimidazolidin-1-yl]-

2-trifluoromethylbenzonitrile, 32. To a suspension of NaH in THF

cooled to 0 °C was added the unsubstituted thiohydantoin 30, and

the mixture was stirred for 30 min. A solution of ethyl iodide in

THF was added and the mixture stirred for 5 h. Normal aqueous

workup and extraction afforded the crude product which was

purified by column chromatography, using silica gel (dichloro-

methane), to give in 5% yield the pure 4-ethylthiohydantoin 32.

4-[3-(4-Methylphenyl)-5-oxo-4-propyl-2-thioxoimidazolidin-1-yl]-

2-trifluoromethylbenzonitrile, 33. To a suspension of NaH in THF

cooled to 0 °C was added the unsubstituted thiohydantoin 30,

and the mixture was stirred for 30 min. A solution of propyl

iodide in THF was added and the mixture stirred for 5 h. Normal

aqueous workup and extraction afforded the crude product

which was purified by column chromatography, using silica gel

(dichloromethane), to give in 5% yield the pure 4-propylthio-

hydantoin 33.

4-[3-(4-Methylphenyl)-4,4-diethyl-5-oxo-2-thioxoimidazolidin-1-

yl]-2-trifluoromethylbenzonitrile, 34. To a suspension of NaH in

THF cooled to 0 °C was added the 4-ethylthiohydantoin 32, and

the mixture was stirred for 30 min. A solution of ethyl iodide in

THF was added and the mixture stirred for 5 h. Normal

aqueous workup and extraction afforded the crude product which

was purified by column chromatography, using silica gel

(dichloromethane), to give in 5% yield the pure 4,4-diethylthiohy-

dantoin 34.

4-[3-(4-Methylphenyl)-4-ethyl-4-methyl-5-oxo-2-thioxoimida-

zolidin-1-yl]-2-trifluoromethylbenzonitrile, 35. To a suspension

of NaH in THF cooled to 0 °C was added the 4-methylthiohy-

dantoin 31, and the mixture was stirred for 30 min. A solution of

ethyl iodide in THF was added and the mixture stirred for 5 h.

Normal aqueous workup and extraction afforded the crude

product which was purified by column chromatography, using

silica gel (dichloromethane), to give in 5% yield the pure 4,4-

diethylthiohydantoin 35.

4-[3-(4-Methylphenyl)-5-ethyl-5-methyl-4-oxo-2-thioxoimida-

zolidin-1-yl]-2-trifluoromethylbenzonitrile, 36. To a suspension

of NaH in THF cooled to 0 °C was added the unsubstituted

4-oxothiohydantoin 30b, and the mixture was stirred for 30 min.

A solution of methyl iodide in THF was added and the mixture

stirred for 5 h. Normal aqueous workup and extraction afforded

the crude product which was purified by column chromatogra-

phy, using silica gel (dichloromethane), to give in 5% yield the

5-methyl-4-oxothiohydantoin 36a. To a suspension of NaH in

THF cooled to 0 °C was added the monomethyl 4-oxothiohy-

dantoin 36a, and the mixture was stirred for 30 min. A solution

{4-[(1-Cyano-1-methylethyl)amino]phenyl}carbamic Acid tert-

Butyl Ester, 29b. A mixture of 29a (0.83 g, 4 mmol) and acetone

cyanohydrin (4 mL) and MgSO₄(2 g) was heated to 80 °C and

stirred over 2.5 h. After the mixture was cooled to 21 °C,

compound 29b was crystallized from water (30 mL). The solid

was filtered and dried to yield 29b (1.08 g, 3.9 mmol, 98%).

{4-[3-(4-Cyano-3-trifluoromethylphenyl)-4-imino-5,5-dimethyl-

2-thioxoimidazolidin-1-yl]phenyl}carbamic Acid tert-Butyl Es-

ter, 29c. Triethylamine (0.202 g, 2 mmol) was added to a solution

of isothiocyanate 12a (0.456 g, 2 mmol) and 29b (0.57 g, 2 mmol)

in dry THF (5 mL). The reaction mixture was stirred at 21 °C for

15 h and then concentrated to yield a dark residue which was

subjected to flash chromatography (ethyl ether/acetone, 97:3) to

afford 29c (0.15 g, 0.3 mmol, 15%).

4-[3-(4-Aminophenyl)-4,4-dimethyl-5-oxo-2-thioxoimidazolidin-

1-yl]-2-trifluoromethylbenzonitrile, 29. Amixtureof29c (0.15 g, 0.3

mmol) in aqueous 3 N HCl (1 mL) and methanol (4 mL) was

heated to reflux for 2 h. After being cooled to 21 °C, the reaction

mixture was poured into cold water (5 mL) and extracted with

dichloromethane (8 mL). The organic layer was dried over

MgSO₄, concentrated, and chromatographed (dichloromethane/

acetone, 9:1) to yield 29 (0.118 g, 0.29 mmol, 97%) as a yellow

solid. ¹H NMR (400 MHz, CDCl₃) δ 1.54 (s, 6H), 6.73-6.75 (m,

2H), 7.00-7.03 (m, 2H), 8.02 (dd, J₁= 8.2 Hz, J₂= 1.8 Hz, 1H),

8.16 (d, J = 1.8 Hz, 1H), 8.20 (d, J = 8.2 Hz, 1H); ¹³C NMR (100

MHz, CDCl₃) δ22.7, 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.

Synthesis of 25. 4-[3-(4-Azidophenyl)-4,4-dimethyl-5-oxo-2-

thioxoimidazolidin-1-yl]-2-trifluoromethylbenzonitrile, 25. An

aqueous solution of sulfuric acid (25 wt %, 1 mL) was added

to a solution of 29 (0.10 g, 0.25 mmol) in acetone (1 mL) cooled

to -5 °C. An aqueous solution of NaNO₂(0.024 g, 0.35 mmol,

in 0.5 mL of water) was added slowly to the above mixture over

0.1 h. The reaction mixture was allowed to stir at -5 °C for an

additional 1 h, and then an aqueous solution of NaN₃(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 21 °C and stirred for an additional 3 h. The product was

extracted with dichloromethane (3 ꢀ 5 mL). The combined

organic layer was dried over MgSO₄, concentrated, and chro-

matographed (dichloromethane) to yield 25 (0.08 g, 0.18 mmol,

72%) as a yellowish solid. ¹H NMR (400 MHz, CDCl₃) δ 1.54 (s,

6H), 7.17-7.20 (m, 2H), 7.27-7.30 (m, 2H), 7.84 (dd, J₁= 8.3,

1.8 Hz, 1H), 7.96 (d, J = 1.8 Hz, 1H), 7.97 (d, J = 8.3 Hz, 1H);

¹³C NMR (100 MHz, CDCl₃) δ 23.7, 66.4, 110.1, 114.8, 120.4,

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2789

of ethyl iodide in THF was added and the mixture stirred for 5 h.

Normal aqueous workup and extraction afforded the crude

product which was purified by column chromatography, using

silica gel (dichloromethane), to give in 5% yield the 5-ethyl-5-

methyl-4-oxothiohydantoin 36.

Synthesis of 40. 1-(4-Methylphenyl)aminocyclohexanecar-

bonitrile, 40a. 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 90% acetic acid (3 mL). The reaction

mixture was stirred at 21 °C for 12 h, and then 20 mL of ethyl

acetate was added. The organic layer was washed with water

(3 ꢀ 10 mL), dried over magnesium sulfate, and concentrated

under vacuum to dryness to yield 40a (0.398 g, 1.86 mmol, 93%)

as a brown solid.

4-(4-Imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-

3-yl)-2-trifluoromethylbenzonitrile, 40b. Triethylamine (0.05 g, 0.5

mmol) was added to a solution of isothiocyanate 12a (0.228 g,

1 mmol) and 40a (0.214 g, 1 mmol) in THF (2 mL). The reaction

mixture was stirred at 21 °C for 2 days and then concentrated to

yield a dark residue which was subjected to flash chromatography

(dichloromethane/acetone, 95:5) to afford 40b (0.035 g, 0.08

mmol, 8%).

4-[3-(4-Methylphenyl)-5,5-diethyl-4-oxo-2-thioxoimidazolidin-1-

yl]-2-trifluoromethylbenzonitrile, 37. To a suspension of NaH in

THF cooled to 0 °C was added the unsubstituted 4-oxothiohy-

dantoin 30b, and the mixture was stirred for 30 min. A solution of

ethyl iodide in THF was added and the mixture stirred for 5 h.

Normal aqueous workup and extraction afforded the crude

product which was purified by column chromatography, using

silica gel (dichloromethane), to give in 5% yield the 5-ethyl-4-

oxothiohydantoin 37a. To a suspension of NaH in THF cooled to

0 °C was added the monoethyl 4-oxothiohydantoin 37a, and the

mixture was stirred for 30 min. A solution of ethyl iodide in THF

was added and the mixture stirred for 5 h. Normal aqueous workup

and extraction afforded the crude product which was purified by

column chromatography, using silica gel (dichloromethane), to

give in 5% yield the 5,5-diethyl-4-oxothiohydantoin 37.

Synthesis of 39. 1-(4-Methylphenyl)aminocyclopentanecarbo-

nitrile, 39a. 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 21 °C for 6 h and then concentrated under vacuum

to obtain a brown liquid which was subjected to chromatogra-

phy (dichloromethane) to yield 39a (0.981 g, 4.9 mmol, 98%) as

a yellowish solid.

4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.5]dec-

3-yl)-2-trifluoromethylbenzonitrile, 40. A mixture of 40b (0.035

g, 0.08 mmol) in aqueous 2 N HCl (1 mL) and methanol (3 mL)

was heated to reflux for 2 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (5 mL) and

extracted with ethyl acetate (6 mL). The organic layer was dried

over MgSO₄, concentrated, and chromatographed (dichloro-

methane) to yield 40 (0.034 g, 0.076 mmol, 95%) as a

1

white powder. H NMR (CDCl₃, 400 MHz) δ 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 = 8.2, 1.8

Hz, 1H), 7.96 (d, J = 8.3 Hz, 1H), 7.97 (d, J = 1.8 Hz, 1H); ¹³

C

4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.4]non-

3-yl)-2-trifluoromethylbenzonitrile, 39. A mixture of isothiocya-

nate 12a (0.296 g, 1.3 mmol) and 39a (0.2 g, 1 mmol) in DMF (0.2

mL) was stirred for 48 h. To this mixture were added methanol

(10 mL) and aqueous 2 N HCl (3 mL). The second mixture was

refluxed for 6 h. After being cooled to 21 °C, the reaction mixture

was poured into cold water (20 mL) and extracted with ethyl

acetate (30 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane) to yield

39 (0.3 g, 0.7 mmol, 70%) as a white powder. ¹H NMR (CDCl₃,

400 MHz) δ 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 = 8.2, 1.8 Hz, 1H), 7.96 (d, J =

8.2 Hz, 1H), 7.98 (d, J = 1.8 Hz, 1H); ¹³C NMR (CDCl₃, 100

MHz) δ 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.

Synthesis of 38. 1-(4-Methylphenyl)aminocyclobutanecarbo-

nitrile, 38a. 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 21 °C for 6 h and then concentrated under vacuum

to obtain a brown liquid which was subjected to chromatogra-

phy (dichloromethane) to yield 38a (0.912 g, 4.9 mmol, 98%) as

a yellowish solid.

NMR (CDCl₃, 100 MHz) δ 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.

Synthesis of 41. 1-(4-Methylphenyl)aminocycloheptanecar-

bonitrile, 41a. 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 21 °C for 12 h, and then 20 mL of ethyl

acetate was added. The organic layer was washed with water

(3 ꢀ 10 mL), dried over magnesium sulfate, and concentrated

under vacuum to dryness to yield 41a (0.438 g, 1.92 mmol, 96%)

as a brown solid.

4-(4-Imino-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.6]undec-

3-yl)-2-trifluoromethylbenzonitrile, 41b. Triethylamine (0.05 g, 0.5

mmol) was added to a solution of isothiocyanate 12a (0.228 g,

1 mmol) and 41a (0.228 g, 1 mmol) in THF (2 mL). The reaction

mixture was stirred at 21 °C for 2 days and then concentrated to

yield a dark residue which was subjected to flash chromatogra-

phy (dichloromethane/acetone, 95:5) to afford 41b (0.036 g, 0.08

mmol, 8%).

4-(4-Oxo-2-thioxo-1-(4-methylphenyl)-1,3-diazaspiro[4.6]undec-

3-yl)-2-trifluoromethylbenzonitrile, 41. A mixture of 41b (0.036 g,

0.08 mmol) in aqueous 2 N HCl (1 mL) and methanol (3 mL) was

heated to reflux for 2 h. After being cooled to 21 °C, the reaction

mixture was poured into cold water (5 mL) and extracted with

ethyl acetate (6 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane) to yield

4-(8-Oxo-6-thioxo-5-(4-methylphenyl)-5,7-diazaspiro[3.4]oct-

7-yl)-2-trifluoromethylbenzonitrile, 38. A mixture of isothiocya-

nate 12a (0.912 g, 4 mmol) and 38a (0.558 g, 3 mmol) in DMF

(0.5 mL) was stirred at 21 °C for 24 h. To this mixture were

added methanol (30 mL) and aqueous 2 N HCl (6 mL). The

second mixture was refluxed for 6 h. After being cooled to 21 °C,

the reaction mixture was poured into cold water (50 mL) and

extracted with ethyl acetate (60 mL). The organic layer was dried

over MgSO₄, concentrated, and chromatographed (dichloro-

methane) to yield 38 (0.959 g, 2.31 mmol, 77%) as a white

1

41 (0.034 g, 0.075 mmol, 94%) as a white powder. H NMR

(CDCl₃, 400 MHz) δ 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 = 8.2,

1.8 Hz, 1H), 7.95-7.97 (m, 2H); ¹³C NMR (CDCl₃, 100 MHz) δ

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.

Synthesis of 43. 1-Methyl-4-(4-methylphenylamino)piperi-

dine-4-carbonitrile, 43a. 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 21 °C for 6 h, and

1

powder. H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.96

(d, J = 8.3 Hz, 1H), 7.99 (d, J = 1.8 Hz, 1H); ¹³C NMR (CDCl₃,

100 MHz) δ 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.

2790 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

then 100 mL of dichloromethane was added. The organic layer

was washed with aqueous 2 N NaOH (2 ꢀ 50 mL), dried over

magnesium sulfate, concentrated, and chromatographed

(dichloromethane and then acetone) to obtain 43a (0.722 g,

3.15 mmol, 63%).

4-(4-Imino-8-methyl-2-thioxo-1-(4-methylphenyl)-1,3,8-triazaspiro-

[4.5]dec-3-yl)-2-trifluoromethylbenzonitrile, 43b. Triethylamine (0.02,

0.2 mmol) was added to a solution of isothiocyanate 12a (0.228 g,

1 mmol) and 43a (0.114 g, 0.5 mmol) in THF (2 mL). The reaction

mixture was stirred at 21 °C 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 43b

(0.059 g, 0.13 mmol, 26%).

4-(8-Methyl-4-oxo-2-thioxo-1-(4-methylphenyl)-1,3,8-triazaspiro-

[4.5]dec-3-yl)-2-trifluoromethylbenzonitrile, 43. A mixture of 43b

(0.059 g, 0.13 mmol) in aqueous 2 N HCl (1 mL) and methanol

(3 mL) was heated to reflux for 2 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (5 mL) and extracted

with ethyl acetate (10 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane/acetone,

60:40) to yield 43 (0.055 g, 0.012 mmol, 92%) as a white powder.

¹H NMR (acetone-d₆, 400 MHz) δ 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, J₁=

8.2, 1.8 Hz, 1H), 8.16 (d, J = 1.8 Hz, 1H), 8.22 (d, J = 8.2 Hz, 1H);

¹³C NMR (acetone-d₆, 100 MHz) δ 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.

Synthesis of 49. 1-(4-Hydroxyphenyl)aminocyclobutanecar-

bonitrile, 49a. 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 21 °C for 6 h and then concentrated under

vacuum to obtain a brown liquid which was subjected to

chromatography (dichloromethane/acetone, 98:2) to yield 49a

(0.903 g, 4.8 mmol, 96%) as a yellowish solid.

4-(8-Oxo-6-thioxo-5-(4-hydroxyphenyl)-5,7-diazaspiro[3.4]oct-

7-yl)-2-trifluoromethylbenzonitrile, 49. A mixture of isothiocya-

nate12a(0.57 g, 2.5mmol)and49a (0.376g, 2mmol)inDMF(0.5

mL) was stirred at 21 °C for 40 h. To this mixture were added

methanol (30 mL) and aqueous 2 N HCl (5 mL). The second

mixture was refluxed for 6 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (40 mL) and extracted

with ethyl acetate (50 mL). The organic layer was dried over

MgSO₄, concentrated, and chromatographed (dichloromethane/

acetone, 98:2) to yield 49 (0.659 g, 1.58 mmol, 79%) as a white

powder. ¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 8.14 (d, J =

1.8 Hz, 1H), 8.21 (d, J = 8.3 Hz, 1H); ¹³C NMR (acetone-d₆, 100

MHz) δ 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.

Synthesis of 50 and 51. 1-Aminocyclopentanecarbonitrile, 50a.

Anhydrous ammonia was bubbled into a mixture of cyclopen-

tanone (0.452 g) and trimethylsilyl cyanide (0.66 mL, 5 mmol).

The ammonia was refluxed with a dry ice-acetone condenser.

After 1 h of reflux, the ammonia was allowed to evaporate from

the medium and then the remaining mixture was concentrated

under vacuum to yield 50a (0.522 g, 4.75 mmol, 95%) as a

colorless liquid.

4-(4-Oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2-trifluoro-

methylbenzonitrile, 50c. A mixture of 50b (0.741 g, 2.19 mmol) in

aqueous 2 N HCl (4 mL) and methanol (20 mL) was heated to

reflux for 1 h. After being cooled to 21 °C, the reaction mixture

was poured into cold water (20 mL) and extracted with ethyl

acetate (40 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane) to yield

50c (0.72 g, 2.12 mmol, 97%) as a white powder.

4-[1-(4-Cyanophenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-

3-yl]-2-trifluoromethylbenzonitrile, 50. A mixture of 50c (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 di-

methylformamide (0.5 mL) was placed in a sealed tube under

argon and heated to 140 °C for 5 days. The reaction mixture

was poured into ethyl acetate (5 mL) and washed with water

(2 ꢀ 10 mL). The organic layer was dried over MgSO₄, con-

centrated, and chromatographed (dichloromethane) to yield 50

(0.023 g, 0.052 mmol, 26%) as a white powder. ¹H NMR

(CDCl₃, 400 MHz) δ 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); ¹³C NMR (CDCl₃, 100

MHz) δ 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.

4-[1-(4-Nitrophenyl)-4-oxo-2-thioxo-1,3-diazaspiro[4.4]non-3-

yl]-2-trifluoromethylbenzonitrile, 51. A mixture of 50c (0.0678 g,

0.2 mmol), 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU, 0.05 g,

0.33 mmol), and 4-fluoronitrobenzene (0.056 g, 0.4 mmol) in

dimethylformamide (0.5 mL) was placed in a sealed tube under

argon and heated to 130 °C for 40 h. The reaction mixture was

poured into ethyl acetate (5 mL) and washed with water (2 ꢀ

10 mL). The organic layer was dried over MgSO₄, concentrated,

and chromatographed (dichloromethane) to yield 51 (0.038 g,

1

0.084 mmol, 42%) as a white powder. H NMR (CDCl₃, 400

MHz) δ 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 = 8.2,

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); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

Synthesis of 52. 2-Methyl-2-(4-trifluoromethylphenyl)aminopro-

panenitrile, 52a. A mixture of 4-trifluoromethylaniline (1.61 g, 10

mmol), acetone cyanohydrin (5 mL), and magnesium sulfate (2 g)

was heated to 80 °C and stirred for 12 h. To the medium was added

ethyl acetate (50 mL) and then washed with water (3 ꢀ 30mL). The

organic layer was dried over MgSO₄and concentrated under

vacuum to dryness to yield 52a (2.166 g, 9.5 mmol, 95%) as brown

solid.

4-(4,4-Dimethyl-5-oxo-2-thioxo-3-(4-trifluoromethylphenyl)-

imidazolidin-1-yl)-2-trifluoromethylbenzonitrile, 52. A mixture

of isothiocyanate 12a (0.114 g, 0.5 mmol) and 52a (0.092 g, 0.4

mmol) in DMF (0.3 mL) was stirred at 21 °C for 48 h. To this

mixture were added methanol (10 mL) and aqueous 2 N HCl

(3 mL). The second mixture was refluxed for 6 h. After being

cooled to 21 °C, the reaction mixture was poured into cold water

(20 mL) and extracted with ethyl acetate (20 mL). The organic

layer was dried over MgSO₄, concentrated, and chromato-

graphed (dichloromethane) to yield 52 (0.117 g, 0.256 mmol,

64%) as a white powder. ¹H NMR (CDCl₃, 400 MHz) δ 1.61 (s,

6H), 7.45-7.49 (m, 2H), 7.80-7.83 (m, 2H), 7.85 (dd, J = 8.3,

1.8 Hz, 1H), 7.97 (d, J = 1.8 Hz, 1H), 7.99 (d, J = 8.2 Hz, 1H);

¹³C NMR (CDCl₃, 100 MHz) δ 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.

4-(4-Imino-2-thioxo-1,3-diazaspiro[4.4]non-3-yl)-2-trifluoro-

methylbenzonitrile, 50b. Triethylamine (0.101 g, 0.1 mmol) was

added to a solution of isothiocyanate 12a (0.684 g, 3 mmol) and

50a (0.33 g, 3 mmol) in THF (5 mL). The reaction mixture was

stirred at 21 °C for 5 h and then concentrated to yield a brown

residue which was subjected to flash chromatography

(dichloromethane/acetone, 93:7) to afford 50b (0.741 g, 2.19

mmol, 73%).

Synthesis of 74-76. 1-(4-Hydroxymethylphenylamino)cyclobu-

tanecarbonitrile, 74a. Trimethylsilyl cyanide (0.66 mL, 5 mmol)

was added dropwise to a mixture of 4-aminobenzyl alcohol (0.492 g,

4 mmol) and cyclobutanone (0.35 g, 5 mmol) in dichloromethane

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2791

(10 mL). The reaction mixture was stirred at 21 °C for 6 h and then

concentrated under vacuum to obtain a brown liquid which was

subjected to chromatography (dichloromethane) to yield 74a

(0.677 g, 3.36 mmol, 84%) as a brown solid.

(E)-3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]oct-5-yl]phenyl}acrylic Acid, 77b. A mixture

of 77a (0.025 g, 0.05 mmol) and a solution of sodium hydroxide

(2 mL, 2 M) in methanol (2 mL) was stirred at 21 °C for 5 h.

The methanol was evaporated. The residue was adjusted to pH

5 by aqueous 2 N HCl and then extracted with ethyl acetate

(3 ꢀ 50 mL). The organic layer was dried over MgSO₄

and concentrated to dryness to obtain 77b (0.02 g, 0.042 mmol,

85%).

(E)-3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]oct-5-yl]phenyl}-2-propanamide, 77. To a sus-

pension of 77b (0.02 g, 0.042 mmol) in THF (1 mL) cooled to

-5 °C was added thionyl chloride (0.007 mL, 0.1 mmol). The

medium was stirred at -5 °C for 1 h. Then ammonia was bubbled

into the mixture. The excess of ammonia was condensed by a

reflux condenser cooled at -78 °C for 30 min and then was

allowed to evaporate. The medium was filtered and the filtrate

was concentrated and chromatographed (dichloromethane/acet-

one, 70:30) to yield 77 (0.014 g, 0.03 mmol, 71%) as an off-white

powder. ¹H NMR (DMSO-d₆, 400 MHz) δ 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), 7.78 (d, J = 8.3 Hz, 2H), 8.03

(dd, J = 8.3, 1.8Hz, 1H), 8.23 (d, J = 1.8 Hz, 1H), 8.34 (d, J= 8.3

Hz, 1H).

(E)-4-{5-[4-(3-Hydroxy-1-propenyl)phenyl]-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-7-yl}-2-trifluoromethylbenzonitrile, 78. To a

mixture of 77a (0.05 g, 0.1 mmol) in dichloromethane (2 mL)

at -78 °C was added a solution of diisobutylaluminum hydride

in THF (0.11 mL, 1 M, 0.11 mmol). The mixture was stirred at

-78 °C for 3 h. After being warmed to 21 °C, the mixture was

washed with an aqueous solution of sodium thiosulfate and

extracted with ethyl acetate. The organic layer was dried over

MgSO₄, concentrated, and chromatographed (dichloromethane/

acetone, 95:5) to yield 78 (0.040 g, 0.089 mmol, 89%) as a white

powder. ¹H NMR (CDCl₃, 400 MHz) δ 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, J₁= 16.0, 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 = 8.3, 1.8 Hz, 1H), 7.96-7.98 (m, 2H); ¹³C NMR

(CDCl₃, 100 MHz) δ13.7, 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.

Synthesis of 79 and 80. 4-[4-(1-Cyanocyclobutylamino)phe-

nyl]butanoic Acid, 79a. 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 h. After filtration to remove the sodium sulfate, the

mixture was concentrated under vacuum to obtain a brown liquid

which was subjected to chromatography (dichloromethane/acet-

one, 50:50) to yield 79a (0.665 g, 2.58 mmol, 86%) as a yellowish

solid.

4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}butanoic Acid Methyl Ester, 79b. A

mixture of isothiocyanate 12a (0.547 g, 2.4 mmol) and 79a (0.342

g, 1.5 mmol) in DMF (2 mL) was stirred at 21 °C for 15 h. To this

mixture were added methanol (10 mL) and aqueous 2 N HCl (5

mL). The second mixture was refluxed for 3 h. After being cooled

to 21 °C, the reaction mixture was poured into cold water (10 mL)

and extracted with ethyl acetate (3 ꢀ 30 mL). The organic layer

was dried over MgSO₄, concentrated, and chromatographed

(dichloromethane) to yield 79b (0.594 g, 1.18 mmol, 79%) as a

white powder. ¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.95 (d, J = 8.3 Hz, 1H), 7.98 (d,

J = 1.8 Hz, 1H); ¹³C NMR (CDCl₃, 100 MHz) δ 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

4-[8-(4-Hydroxymethylphenyl)-5-oxo-7-thioxo-6-azaspiro[3.4]oct-

6-yl]-2-trifluoromethylbenzonitrile, 74. A mixture of isothiocyanate

12a (0.342 g, 1.5 mmol) and 74a (0.21 g, 1 mmol) in dry DMF (0.5

mL) was stirred at 21 °C for 24 h. To this mixture were added

methanol (20 mL) and HCl aqueous 2 N (5 mL). The second

mixture was refluxed for 6 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (40 mL) and extracted

with ethyl acetate (60 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane/acetone,

90:10) to yield 74 (0.296 g, 0.69 mmol, 69%) as a white powder.

¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.95-7.98

(m,2H);¹³CNMR(CDCl₃,100MHz) δ13.7, 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.

4-[5-(4-Formylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-

7-yl]-2-trifluoromethylbenzonitrile, 75. To a mixture of 74 (0.303

g, 0.7 mmol) and the 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 h at 21 °C, and then ethyl

ether (10 mL) was added to precipitate the byproduct of the

reaction. After filtration and concentration under reduced pres-

sure, the mixture was chromatographed (dichloromethane/acet-

one, 95:5) to yield 75 (0.24 g, 0.56 mmol, 80%) as a white powder.

¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.97-7.99 (m, 2H), 8.11 (d,

J = 8.1 Hz, 2H), 10.12 (s, 1H); ¹³C NMR (CDCl₃, 100 MHz) δ

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.

4-{5-[4-(1-Hydroxyethyl)phenyl]-8-oxo-6-thioxo-5,7-diazaspiro-

[3.4]oct-7-yl}-2-trifluoromethylbenzonitrile, 76. A mixture of 75

(0.043 g, 0.1 mmol) and THF (1 mL) in a flamed-dried flask was

placed under argon and cooled to -78 °C. Methylmagnesium

iodide (1.1 mL, 0.1 M) was added. The mixture was stirred at

-78 °C for 30 min and warmed slowly to 21 °C. The medium was

washed with water (3 mL) and extracted with ethyl acetate (10 mL).

The organic layer was dried over MgSO₄, concentrated, and

chromatographed (dichloromethane/acetone, 95:5) to yield 76

1

(0.037 g, 0.082 mmol, 82%) as a white powder. H NMR (CD-

Cl₃, 400 MHz) δ 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 (qd, J = 6.5, 3.1 Hz, 1H), 7.29 (d, J = 8.3

Hz, 2H), 7.60 (d, J = 8.2 Hz, 2H), 7.85 (dd, J = 8.3, 1.8 Hz, 1H),

7.95-7.98 (m, 2H); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

Synthesis of 77 and 78. (E)-3-{4-[7-(4-Cyano-3-trifluoro-

methylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]oct-5-yl]phenyl}-

acrylic Acid Ethyl Ester, 77a. A mixture of 75 (0.043 g, 0.1 mmol)

and (carbethoxymethylene)triphenylphosphorane (0.039 g, 0.12

mmol) in dichloromethane (2 mL) was stirred at 21 °C for 10 h.

The mixture was concentrated and chromatographed (dichloro-

methane) to yield 77a (0.048 g, 0.096 mmol, 96%) as a white

powder. ¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.96-7.98

(m, 2H); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

2792 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

(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.

HCl (5 mL, 2 M). The second mixture was refluxed for 3 h. After

being cooled to 21 °C, the reaction mixture was poured into cold

water (10 mL) and extracted with ethyl acetate (3 ꢀ 30 mL). The

organic layer was dried over MgSO₄, concentrated, and chroma-

tographed (dichloromethane) to yield 81b (0.582 g, 1.19 mmol,

62%) as a white powder. ¹H NMR (CDCl₃, 400 MHz) δ

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 = 8.3, 1.8 Hz, 1H), 7.95 (d, J = 8.3 Hz, 1H),

7.98 (d, J = 1.8 Hz, 1H); ¹³C NMR (CDCl₃, 100 MHz) δ 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, 132.3, 133.2, 133.3 (q, J = 33.2

Hz), 135.7, 137.2, 142.5, 173.1, 174.9, 179.9.

3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}propanoic Acid, 81c. A mixture of

81b (0.487 g, 1 mmol) in methanol (10 mL) and solution of

sodium hydroxide (10 mL, 2 M) was stirred at 21 °C for 5 h. The

methanol was evaporated. The residue was adjusted to pH 5 by

aqueous 2 N HCl and was then extracted with ethyl acetate (3 ꢀ

50 mL). The organic layer was dried over MgSO₄and concen-

trated to dryness to obtain 81c (0.472 g, 0.99 mmol, 99%).

3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}propanamide, 81. To a suspension

of 81c (0.094 g, 0.2 mmol) in THF (10 mL) cooled at -5 °C was

added thionyl chloride (0.019 mL, 0.26 mmol). The medium was

stirred at -5 °C for 1 h. Then ammonia was bubbled into the

mixture. The excess ammonia was condensed by a reflux con-

denser cooled to -78 °C for 30 min and then was allowed to

evaporate. The mixturewas filtered. The filtratewas concentrated

and chromatographed (dichloromethane/acetone, 70:30) to yield

81 (0.09 g, 0.19 mmol, 95%) as an off-white powder. ¹H NMR

(acetone-d₆, 400 MHz) δ 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 = 8.3 Hz, 1.8 Hz, 1H), 8.15 (d, J = 1.8

Hz, 1H), 8.22 (d, J = 8.3 Hz, 1H); ¹³C NMR (acetone-d₆, 100

MHz) δ 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.

3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}-N-methylpropanamide, 82. To a

suspension of 81c (0.094 g, 0.2 mmol) in THF (10 mL) cooled

to -5 °C was added thionyl chloride (0.019 mL, 0.26 mmol). The

mixture was stirred at -5 °C for 1 h. Then methylamine was

bubbled into the mixture at -5 °C for 30 min. The medium was

filtered. The filtrate was concentrated and chromatographed

(dichloromethane/acetone, 75:25) to yield 82 (0.092 g, 0.19 mmol,

95%) as an off-white powder. ¹H NMR (acetone-d₆, 400 MHz) δ

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 = 8.3, 1.8 Hz, 1H), 8.13 (d, J = 1.8 Hz, 1H),

8.20 (d, J = 8.3 Hz, 1H);¹³C NMR(acetone-d₆, 100 MHz) δ 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.

4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diaza-spiro[3.4]oct-5-yl]phenyl}butanoic Acid, 79c. A mixture of

79b (0.501 g, 1 mmol) and a solution of sodium hydroxide (10mL,

2 M) in methanol (10 mL) was stirred at 21 °C for 5 h. The

methanol was evaporated. The residue was adjusted to pH 5 by

aqueous 2 M HCl, and then the mixture was extracted with ethyl

acetate (3 ꢀ 50 mL). The organic layer was dried over MgSO₄and

concentrated to dryness to obtain 79c (0.482 g, 0.99 mmol, 99%).

¹H NMR (CDCl₃, 400 MHz) δ 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); ¹³C NMR

(CDCl₃, 100 MHz) δ 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.

4-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}butanamide, 79. To a suspension of

79c (0.097 g, 0.2 mmol) in THF (10 mL) cooled to -5 °C was

added thionyl chloride (0.019 mL, 0.26 mmol). The mixture

was stirred at -5 °C for 1 h. Then ammonia was bubbled into

the mixture. The excess ammonia was condensed by a reflux

condenser cooled to -78 °C for 30 min and then was allowed to

evaporate. The mixture was filtered. The filtratewasconcentrated

and chromatographed (dichloromethane/acetone, 70:30) to yield

79 (0.093 g, 0.19 mmol, 95%) as an off-white powder. ¹H NMR

(CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.95 (d, J = 8.3 Hz, 1H), 7.97 (d, J =

1.8 Hz, 1H); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

N-Methyl-4-{4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-

thioxo-5,7-diazaspiro[3.4]oct-5-yl]phenyl}butanamide, 80. To a

suspension of 79c (0.097 g, 0.2 mmol) in THF (10 mL) cooled to

-5 °C was added thionyl chloride (0.019 mL, 0.26 mmol). The

mixture was stirred at -5 °C for 1 h. Then methylamine was

bubbled into the mixture at -5 °C for 30 min. The mixture was

filtered. The filtrate was concentrated and chromatographed

(dichloromethane/acetone, 75:25) to yield 80 (0.095 g, 0.19

1

mmol, 95%) as an off-white powder. H NMR (CDCl₃, 400

MHz) δ 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 = 8.3, 1.8 Hz, 1H),

7.91 (d, J = 8.3 Hz, 1H), 7.94 (d, J = 1.8 Hz, 1H); ¹³C NMR

(CDCl₃, 100 MHz) δ 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, 133.3 (q, J = 33.2 Hz), 133.8, 135.2, 137.3,

143.7, 173.3, 174.9, 179.8.

Synthesis of 81-83. 3-[4-(1-Cyanocyclobutylamino)phenyl]-

propanoic Acid, 81a. 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 h. After filtration to remove sodium sulfate, the mixture was

concentrated under vacuum to obtain a brown liquid which was

subjected to chromatography (dichloromethane/acetone, 50:50) to

yield 81a (0.472 g, 1.93 mmol, 97%) as a yellowish solid.

3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}-N-(2-hydroxyethyl)propanamide, 83.

To a suspension of 81c (0.094 g, 0.2 mmol) in THF (10 mL) cooled

to -5 °C was added thionyl chloride (0.019 mL, 0.26 mmol). The

mixture was stirred at -5 °C for 1 h. Then 2-aminoethanol (0.0183

g, 0.03 mmol) was added into the mixture at -5 °C. After being

stirred for an additional 30 min, the mixture was filtered. The filtrate

was concentrated and chromatographed (dichloromethane/acet-

one, 50:50) to yield 83 (0.093 g, 0.18 mmol, 90%) as an off-white

3-{4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-

diazaspiro[3.4]oct-5-yl]phenyl}propanoic Acid Methyl Ester, 81b.

A mixture of isothiocyanate 12a (0.661 g, 2.9 mmol) and 81a

(0.472 g, 1.93 mmol) in DMF (2 mL) was stirred at 21 °C for 15 h.

To this mixture were added methanol (10 mL) and aqueous 2 N

1

powder. H NMR (acetone-d₆, 400 MHz) δ 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 = 11.2, 5.6 Hz, 2H), 3.51 (dd, J= 11.2, 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

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2793

Hz, 2H), 8.02 (dd, J = 8.3, 1.8 Hz, 1H), 8.14 (d, J = 1.8 Hz, 1H),

8.22 (d, J = 8.3 Hz, 1H); ¹³C NMR (acetone-d₆, 100 MHz) δ 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.

acetone, 90:10) to yield 86a (0.116 g, 0.44 mmol, 22%) as a

yellowish solid. 4-Methanesulfonylphenylamine (0.201 g, 1.17

mmol, 59%) was also recovered.

4-[5-(4-Methanesulfonylphenyl)-8-oxo-6-thioxo-5,7-diazaspiro-

[3.4]oct-7-yl]-2-trifluoromethylbenzonitrile, 86. A mixture of iso-

thiocyanate 12a (0.0.141 g, 0.62 mmol) and 86a (0.11 g, 0.42

mmol) in dry DMF (2 mL) was stirred at 21 °C for 3 days. To this

mixture were added methanol (10 mL) and aqueous 2 N HCl

(5 mL). The second mixture was refluxed for 3 h. After being

cooled to 21 °C, the reaction mixture was poured into cold water

(10mL) andextracted withethyl acetate(3 ꢀ 30 mL). Theorganic

layer was dried over MgSO₄, concentrated, and chromato-

graphed (dichloromethane/acetone, 97:3) to yield 86 (0.031 g,

Synthesis of 84 and 85. 4-(4-Aminophenyl)piperazine-1-car-

boxylic Acid tert-Butyl Ester, 84a. 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 in a sealed tube and heated under

argon atmosphere to 80 °C for 30 h. After being cooled to 21 °C,

the mixture was washed with water (50 mL) and extracted with

ethyl acetate (100 mL). The organic layer was dried over MgSO₄,

concentrated, and chromatographed (dichloromethane/acetone,

70:30) to yield 84a (0.36 g, 1.3 mmol, 43%) as a yellow powder.

4-[4-(1-Cyanocyclobutylamino)phenyl]piperazine-1-carboxylic

Acid tert-Butyl Ester, 84b. Trimethylsilyl cyanide (0.3 g, 3 mmol)

was added dropwise to a mixture of 84a (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 h. After

filtration to remove the sodium sulfate, the mixture was con-

centrated under vacuum to obtain a brown liquid which was

subjected to chromatography (dichloromethane/acetone, 75:25)

to yield 84b (0.448 g, 1.26 mmol, 84%) as a yellow solid.

4-[8-Oxo-5-(4-piperazin-1-ylphenyl)-6-thioxo-5,7-diazaspiro-

[3.4]oct-7-yl]-2-trifluoromethylbenzonitrile, 84. A mixture of iso-

thiocyanate 12a (0.228 g, 1 mmol) and 84b (0.472 g, 0.63 mmol)

in DMF (1 mL) was stirred at 21 °C for 20 h. The mixture was

concentrated and chromatographed (dichloromethane/acetone,

90:10) to yield the crude thiohydantoin imine 84c (0.173 g, 0.296

mmol, 47%) as an off-white powder. A mixture of this crude

imine 84c (0.117 g, 0.2 mmol), methanol (5 mL), and aqueous 2

N HCl (2 mL) was refluxed for 2 h. After being cooled to 21 °C, the

reaction mixture was poured into cold water (10 mL) and extracted

with ethyl acetate (3 ꢀ 30 mL). The organic layer was dried over

MgSO₄, concentrated, and chromatographed (dichloromethane/

acetone, 50:50, and then methanol/acetone, 50:50) to yield 84

(0.089 g, 0.184 mmol, 92%) as a white powder. ¹H NMR

(CD₃OD, 400 MHz) δ 1.51-1.61 (m, 1H), 2.01-2.11 (m, 1H),

2.48-2.59 (m, 4H), 2.95 (t, J = 4.6 Hz, 4H), 3.19 (t, J = 4.7 Hz,

4H), 7.03 (d, J = 8.9 Hz, 2H), 7.16 (d, J = 8.9 Hz, 2H), 7.86 (dd,

J = 8.3, 1.8 Hz, 1H), 8.02 (d, J = 8.3 Hz, 1H), 8.07 (d, J = 1.8 Hz,

1H); ¹³C NMR (CD₃OD, 100 MHz) δ 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.

4-{5-[4-(4-Methanesulfonylpiperazin-1-yl)phenyl]-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]oct-7-yl}-2-trifluoromethylbenzonitrile, 85. A mix-

ture of 84 (0.049 g, 0.1 mmol), methanesulfonyl chloride (0.012 mL,

0.15 mmol), and triethylamine (0.15 mL) in dichloromethane was

stirred at 21 °C for 5 h. The mixture was filtered and the filtrate was

concentrated and chromatographed (dichloromethane/acetone,

95:5) to yield 85 (0.042 g, 0.074 mmol, 74%) as a white powder.

¹H NMR (CDCl₃, 400 MHz) δ 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 = 8.3, 1.8 Hz, 1H), 7.95 (d, J = 8.3 Hz, 1H), 7.97 (d, J = 1.8 Hz,

1H); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

Synthesis of 86. 1-(4-Methanesulfonylphenylamino)cyclobutane-

carbonitrile, 86a. Trimethylsilyl cyanide (0.4 g, 4 mmol) was added

dropwise to a mixture of 4-methanesulfonylphenylamine hydro-

chloride (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

h at 120 °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/

1

0.065 mmol, 15%) as a white powder. H NMR (CDCl₃, 400

MHz) δ 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); ¹³C NMR (CDCl₃, 100

MHz) δ 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.

Synthesis of 87 and 88. 4-Aminobenzoic Acid Methyl Ester,

87a. Concentrated sulfuric acid was slowly added to a mixture of

4-aminobenzoic acid (4 g, 29.2 mmol) in methanol cooled to

0 °C. After the addition, the mixture was stirred at 21 °C for 5 h.

The mixture was washed with a saturated solution of sodium

bicarbonate and extracted with ethyl acetate. The organic layer

was dried over MgSO₄and concentrated under vacuum to

obtain 87a (4.22 g, 27.9 mmol, 96%) as an off-white solid.

4-[(Cyanodimethylmethyl)amino]benzoic Acid Methyl Ester,

87b. A mixture of 4-aminobenzoic acid methyl ester (0.32 g,

2.12 mmol), acetone cyanohydrin (3 mL), and sodium sulfate (1

g) was refluxed for 15 h. After filtration to remove the sodium

sulfate, the filtrate was washed with brine and extracted with

ethyl acetate. The organic layer was concentrated and chroma-

tographed (dichloromethane/acetone, 60:40) to yield 87b (0.398

g, 1.95 mmol, 92%) as a white solid.

4-[3-(4-Cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-

2-thioxoimidazolidin-1-yl]benzoic Acid Methyl Ester, 87. A mix-

ture of isothiocyanate 12a (0.228 g, 1 mmol) and 87b (0.14 g, 0.64

mmol) in DMF (2 mL) was heated under microwave irradiation

at 60 °C for 12 h. To this mixture were added methanol (6 mL)

and aqueous 2 N HCl (2 mL). The second mixture was refluxed

for 4 h. After being cooled to 21 °C, the reaction mixture was

poured into cold water (10 mL) and extracted with ethyl acetate

(3 ꢀ 30 mL). The organic layer was dried over MgSO₄, con-

centrated, and chromatographed (dichloromethane; dichloro-

methane/acetone, 75:25) to yield 87 (0.18 g, 0.4 mmol, 63%) as a

white powder. ¹H NMR (CDCl₃, 400 MHz) δ 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); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

4-[3-(4-Cyano-3-trifluoromethylphenyl)-5,5-dimethyl-4-oxo-

2-thioxoimidazolidin-1-yl]-N-methylbenzamide, 88. A mixture of

87 (0.02 g, 0.0435 mmol) and methylamine (2 mL distilled from

its 40% aqueous solution) was kept at -20 °C for 15 h. After

evaporation of the methylamine, the mixture was chromato-

graphed (dichloromethane/acetone, 80:20) to yield 88 (0.01 g,

0.0224 mmol, 51%). The ester 87 (0.08 g, 0.0179 mmol, 41%)

was also recovered. ¹H NMR (acetone-d₆, 400 MHz) δ 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); ¹³C

NMR (acetone-d₆, 100 MHz) δ 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.

2794 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

Synthesis of 89. 4-(1-Cyanocyclobutylamino)benzoic Acid,

89a. 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 21 °C for 15 h. The mixture was washed

with aqueous HCl (pH 2) and extracted with ethyl acetate. The

organic layer was dried over magnesium sulfate and concen-

trated to dryness under vacuum to yield 89a (0.426 g, 1.97 mmol,

99%) as a white solid.

4-[7-(4-Cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-5,7-diaza-

spiro[3.4]oct-5-yl]benzoic Acid Methyl Ester, 89b. A mixture of iso-

thiocyanate 12a (0.51 g, 2.22 mmol) and 89a (0.343 g, 1.59 mmol) in

DMF (2 mL) was heated under microwave irradiation at 60 °C and

stirred for 16 h. To this mixture were added methanol (10 mL) and

aqueous 2 M HCl (5 mL). The second mixture was refluxed for 12 h.

After being cooled to 21 °C, the reaction mixture was poured into

cold water (20 mL) and extracted with ethyl acetate (3 ꢀ 30 mL).

The organic layer was dried over MgSO₄, concentrated, and

chromatographed (dichloromethane/acetone, 95:5) to yield 89b

(0.09 g, 0.196 mmol, 12%) as a white powder. ¹H NMR (CDCl₃,

400 MHz) δ 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); ¹³C NMR (CDCl₃, 100 MHz) δ

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.

N-Methyl-4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]oct-5-yl]benzamide, 89. A mixture of 89b (0.046 g,

0.1 mmol) and methylamine (1 mL distilled from its 40% aqueous

solution) was kept at -20 °C for 15 h. After evaporation of the

methylamine, the mixture was chromatographed (dichloromethane/

acetone, 80:20) to yield 89 (0.041 g, 0.085, 84%). ¹H NMR (CDCl₃,

400 MHz) δ 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); ¹³C NMR (CDCl₃, 100 MHz) δ 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.

24 h. 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 91c (2.19 g,

8.87 mmol, 89%). ¹H NMR (CDCl₃, 400 MHz) δ 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); ¹³C NMR (CDCl₃, 100 MHz) δ 15.7,

26.7, 33.9, 49.4, 100.2 (d, J = 29.5 Hz), 110.6, 111.0 (d, J = 11.8

Hz), 120.8, 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).

N-Methyl-4-[7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]oct-5-yl]-2-fluorobenzamide, 91. A mixture of iso-

thiocyanate 12a (2.16 g, 9.47 mmol) and 91c (1.303 g, 5.27 mmol) in

DMF (20 mL) was heated under microwave irradiation at 80 °C for

16 h. To this mixture was added methanol (50 mL) and aqueous 2 N

HCl (20 mL). The second mixture was refluxed for 3 h. After being

cooled to 21 °C, the reaction mixture was poured into cold water

(100 mL) and extracted with ethyl acetate (150 mL). The organic

layer was dried over MgSO₄, concentrated, and chromatographed

(dichloromethane/acetone, 95:5) to yield 91 (1.43 g, 3.0 mmol, 57%)

as a yellow powder. ¹H NMR (CDCl₃, 400 MHz) δ 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); ¹³CNMR(CDCl₃,100MHz) δ13.6, 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; ¹⁹F NMR (CDCl₃, 100 MHz) δ -111.13,

-62.58.

Synthesis of 92. 2-Fluoro-4-nitrobenzoic Acid, 92a. 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 exother-

mic 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

dichloromethane (2 ꢀ 30 mL) and water (2 ꢀ 30 mL). The

organic layer was dried over MgSO₄and concentrated to give

Synthesis of 91. N-Methyl-2-fluoro-4-nitrobenzamide, 91a.

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 °C. The mixture was stirred for an

additional 1 h at -5 °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 addi-

tional 1 h. Ethyl acetate (300 mL) was added to the mixture,

which was washed with brine (3 ꢀ 150 mL). The organic layer

was dried over MgSO₄and concentrated to yield 91a (2.89 g,

1

92a (0.32 mg, 81%) as a white solid. H NMR (CDCl₃, 400

MHz) δ 8.06 (ddd, 1H, J = 9.9, 2.2, 0.3 Hz), 8.13 (ddd, 1H, J =

8.6, 2.2, 0.9 Hz), 8.25 (ddd, 1H, J = 8.6, 7.0, 0.3 Hz).

N-Methyl-2-fluoro-4-(1,1-dimethylcyanomethyl)aminobenza-

mide, 92b. A mixture of 91b (96 mg, 0.57 mmol), acetone

cyanohydrin (0.3 mL, 3.14 mmol), and magnesium sulfate

(50 mg) was heated to 80 °C and stirred for 12 h. To the medium

was added ethyl acetate (25 mL), and then the sample was

washed with water (2 ꢀ 25 mL). The organic layer was dried

over MgSO₄and concentrated and the residue was purified with

SiO₂column chromatography (dichloromethane/acetone,

95:5) to give 92b (101 mg, 75%) as a white solid. ¹H NMR

(CDCl₃, 400 MHz) δ 1.74 (s, 6H), 2.98 (dd, 3H, J = 4.8, 1.1 Hz),

6.58 (dd, 1H, J = 14.6, 2.3 Hz), 6.63 (dd, 1H, J = 8.7, 2.3 Hz),

6.66 (br s, 1H), 7.94 (dd, 1H, J = 8.7, 8.7 Hz).

1

14.6 mmol, 91%) as a yellow solid. H NMR (acetone-d₆, 400

MHz) δ 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).

N-Methyl-2-fluoro-4-aminobenzamide, 91b. A mixture of 91a

(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 h. The solid

particles were filtered off. The filtrate was washed with water and

extracted with ethyl acetate. The organic layer was dried over

MgSO₄, concentrated, and chromatographed (dichloromethane/

acetone, 95:5) to yield 91b (2.3 g, 13.7 mmol, 94%) as an off-white

solid. ¹H NMR (acetone-d₆, 400 MHz) δ 2.86 (d, J = 4.3 Hz, 3H),

5.50 (bs, 2H), 6.37 (dd, J₁= 14.7, 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); ¹³C

NMR (acetone-d₆, 100 MHz) δ 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).

N-Methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide, 91c.

Sodium cyanide (1.47 g, 30 mmol) was added to a mixture of

91b (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 °C for

N-Methyl-4-[3-(4-cyano-3-trifluoromethylphenyl)-5,5-dimeth-

yl-4-oxo-2-thioxoimidazolidin-1-yl]-2-fluorobenzamide, 92.

A

mixture of 92b (30 mg, 0.13 mmol) and 12a (58 mg, 0.26 mmol)

in DMF (1 mL) was heated under microwave irradiation at

100 °C for 11 h. To this mixture was added methanol (20 mL)

and aqueous 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 MgSO₄

and concentrated and the residue was purified with SiO₂column

Article

Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7 2795

chromatography (dichloromethane/acetone, 95:5) to give 92 (30

mg, 51%) as colorless crystals. ¹H NMR (CDCl₃, 400 MHz) δ

1.61 (s, 6H), 3.07 (d, 3H, J = 4.1 Hz), 6.71 (m, 1 H), 7.15 (dd, 1H,

J = 11.7, 2.0 Hz), 7.24 (dd, 1H, J = 8.4, 2.0 Hz), 7.83 (dd, 1H,

J = 8.2, 2.1 Hz), 7.95 (d, 1H, J = 2.1 Hz), 7.99 (d, 1H, J = 8.2

Hz), 8.28 (dd, 1H, J = 8.4, 8.4 Hz). ¹³C NMR (CDCl₃, 125

MHz) δ 23.8, 26.9, 66.5, 110.3, 114.6, 117.7, 117.9, 121.7 (q, J =

272.3 Hz), 126.1, 126.9 (q, J = 4.6 Hz), 132.0, 133.3, 133.6 (q,

J = 33.4 Hz), 135.2, 136.7, 138.9 (d, J = 10.8 Hz), 160.3 (d, J =

248.6 Hz), 162.6 (d, J = 3.3 Hz), 174.3, 179.6. ¹⁹F NMR (CDCl₃,

100 MHz) δ -111.13, -62.58. HRMS: found 465.1023 [M þ H]^þ,

calculated for [C₂₁H₁₆F₄N₄O₂S þ H]^þ465.1003.

(CDCl₃, 400 MHz) δ 1.89 (p, 2H, J = 7.7 Hz), 2.27 (t, 2H, J =

7.7 Hz), 2.60 (t, 2H, J = 7.7 Hz), 2.89 (s, 3H), 2.91 (s, 3H), 7.11

(d, 2H, J = 8.6 Hz), 7.55 (d, 2H, J = 8.6 Hz), 9.70 (br s, 1H).

N,N-Dimethyl-4-(4-aminophenyl)butyramide, 94c. A 1 N NaOH

solution (3 mL) was added to a solution of 94b (0.17 g, 0.55 mmol)

in MeOH (2 mL) at room temperature. The mixture was stirred for

14 h. The mixture was partitioned with chloroform (25 mL) and

water (25 mL). The organic layer was dried over MgSO₄and

concentrated and the residue was purified with SiO₂column

chromatography (dichloromethane/acetone, 9:1) to give 94c (74

mg, 66%) as a white solid. ¹H NMR (CDCl₃, 400 MHz) δ 1.91 (p,

2H, J = 7.7 Hz), 2.28 (t, 2H, J = 7.7 Hz), 2.56 (t, 2H, J = 7.7 Hz),

2.92 (s, 6H), 3.56 (br s, 2H), 6.61 (d, 2H, J = 8.3 Hz), 6.97 (d, 2H,

J = 8.3 Hz).

Synthesis of 93. N-Methyl-4-(1-cyanocyclopentylamino)-2-

fluorobenzamide, 93a. A mixture of 91b (62 mg, 0.37 mmol),

cyclopentanone (0.07 mL, 0.74 mmol), and TMSCN (0.1 mL,

0.74 mmol) was heated to 80 °C and stirred for 13 h. To the

medium was added ethyl acetate (2 ꢀ 20 mL), and then the

sample was washed with water (2 ꢀ 20 mL). The organic layer

was dried over MgSO₄and concentrated and the residue was

purified with SiO₂column chromatography (dichloromethane/

N,N-Dimethyl-4-[4-(1-cyanocyclobutylamino)phenyl]butyramide,

94d. A mixture of 94c (74 mg, 0.36 mmol), cyclobutanone (54 mg,

0.78 mmol), and TMSCN (77 mg, 0.78 mmol) was heated to 80 °C

and stirred for 15 h. To the medium was added ethyl acetate (2 ꢀ

20 mL), and then the sample was washed with water (2 ꢀ 20 mL).

The organic layer was dried over MgSO₄and concentrated and

the residue was purified with SiO₂column chromatography

(dichloromethane/acetone, 9:1) to give 94d (58 mg, 57%) as a white

1

acetone, 95:5) to give 93a (61 mg, 63%) as a white solid. H

NMR (CDCl₃, 400 MHz) δ 1.82-1.95 (m, 4H), 2.10-2.18 (m,

2H), 2.36-2.45 (m, 2H), 2.99 (dd, 3H, J = 4.8, 1.1 Hz), 4.60 (br

s, 1H), 6.50 (dd, 1H, J = 14.6, 2.3 Hz), 6.59 (dd, 1H, J = 8.8, 2.3

Hz), 6.65 (br s, 1H), 7.95 (dd, 1H, J = 8.8, 8.8 Hz).

1

solid. H NMR (CDCl₃, 400 MHz) δ 1.93 (p, 2H, J = 7.6 Hz),

2.11-2.28 (m, 2H), 2.30 (t, 2H, J = 7.6 Hz), 2.33-2.42 (m, 2H),

2.60 (t, 2H, J = 7.6 Hz), 2.75-2.83 (m, 2H), 2.93 (s, 3H), 2.94

(s, 3H), 3.94 (br s, 1H), 6.59 (d, 2H, J = 8.5 Hz), 7.07 (d, 2H, J =

8.5 Hz).

N-Methyl-4-[3-(4-cyano-3-trifluoromethylphenyl)-4-oxo-2-thioxo-

1,3-diazaspiro[4.4]nonan-1-yl]-2-fluorobenzamide, 93. A mixture of

93a (57 mg, 0.22 mmol) and 12a (0.15 g, 0.65 mmol) in DMF (3

mL) was heated under microwave irradiation at 130 °Cfor12h. To

this mixture was added methanol (20 mL) and aqueous 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 MgSO₄and concentrated and the

residue was purified with SiO₂column chromatography

(dichloromethane/acetone, 95:5) to give 93 (56 mg, 48%) as a

pale-yellowish solid. ¹H NMR (CDCl₃, 400 MHz) δ 1.49-1.59 (m,

2H), 1.85-1.96 (m, 2H), 2.13-2.21 (m, 2H), 2.32-2.41 (m, 2H),

3.07 (d, 3H, J = 4.3 Hz), 6.67-6.77 (m, 1H), 7.17 (dd, 1H, J =

11.7, 1.8 Hz), 7.27 (dd, 1H, J = 8.4, 1.8 Hz), 7.84 (dd, 1 H, J = 8.3,

1.8 Hz), 7.96 (d, 1H, J = 1.8 Hz), 7.98 (d, 1H, J = 8.3 Hz), 8.28

(dd, 1H, J = 8.4, 8.4 Hz). ¹³C NMR (CDCl₃, 125 MHz) δ 25.1,

26.9, 36.3, 75.2, 110.2, 114.6, 118.1, 118.3, 121.7 (q, J = 275.9 Hz),

126.4, 127.0 (q, J = 3.8 Hz), 132.0, 133.3, 133.5 (q, J = 33.4 Hz),

135.1, 136.8, 139.6 (d, J = 10.1 Hz), 160.3 (d, J = 245.4 Hz), 162.5

(d, J = 3.3 Hz), 175.5, 179.9. ¹⁹F NMR (CDCl₃, 100 MHz) δ

-111.23, -62.57. HRMS: found 491.1156 [M þ H]^þ, calculated

for [C₂₃H₁₈F₄N₄O₂S þ H]^þ491.1159.

N,N-Dimethyl-4-[4-(7-(4-cyano-3-trifluoromethylphenyl)-8-oxo-

6-thioxo-5,7-diazaspiro[3.4]octan-5-yl)phenyl]butanamide, 94. A

mixture of 94d (58 mg, 0.20 mmol) and 12a (74 mg, 0.32 mmol)

in DMF (3 mL) was heated under reflux for 2 h. To this mixture

was added methanol (20 mL) and aqueous 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 MgSO₄and concentrated and the residue was

purified with SiO₂column chromatography (dichloromethane/

acetone, 95:5) to give 94 (44 mg, 42%) as a pale-yellowish solid. ¹H

NMR (CDCl₃, 400 MHz) δ 1.62-1.73 (m, 1H), 2.04 (p, 2H, J =

7.5 Hz), 2.15-2.30 (m, 1H), 2.40 (t, 2H, J = 7.5 Hz), 2.52-2.63

(m, 2 H), 2.62-2.70 (m, 2H), 2.78 (t, 2H, J = 7.5 Hz), 2.96 (s, 3H),

2.99 (s, 3H), 7.22 (d, 2H, J=8.3Hz), 7.42(d, 2H, J= 8.3 Hz), 7.86

(d, 1H, J = 8.2 Hz), 7.97 (d, 1H, J = 8.2 Hz), 7.98 (s, 1H).

Synthesis of 95. 4-[5-(4-(3-Cyanopropyl)phenyl)-8-oxo-6-thioxo-

5,7-diazaspiro[3.4]octan-7-yl]-2-(trifluoromethyl)benzonitrile, 95. 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 °C. After 15 min,

a dichloromethane solution of 79 (35 mg, 0.07 mmol) was added to

the reaction mixture. Stirring was continued for 20 min at -78 °C,

and then triethylamine (0.03 mL, 0.22 mmol) was added. After

30 min at -78 °C, the reaction mixture was warmed to room

temperature, and then reaction was quenched with saturated

aqueous NH₄Cl solution. The reaction mixture was diluted with

dichloromethane and extracted with dichloromethane. The organic

layer was dried over MgSO₄, concentrated, and chromatographed

(dichloromethane/acetone, 95:5) to yield 95 (29 mg, 87%) as a

viscous oil. ¹H NMR (CDCl₃, 400 MHz) δ1.63-1.73 (m, 1H), 2.07

(p, 2H, J = 7.3 Hz), 2.18-2.30 (m, 1H), 2.42 (t, 2H, J = 7.3

Hz), 2.52-2.62 (m, 2H), 2.63-2.73 (m, 2H), 2.90 (t, 2H, J =

7.3 Hz), 7.27 (d, 2H, J = 8.4), 7.43 (d, 2H, J = 8.4 Hz), 7.86

(dd, 1H, J = 8.3, 1.8 Hz), 7.98 (d, 1H, J = 8.3 Hz), 7.98 (d, 1 H,

J = 1.8).

Synthesis of 94. 4-[4-(2,2,2-Trifluoroacetylamino)phenyl]butyric

Acid, 94a. 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 °C. The mixture was warmed to

room temperature and stirred for 3 h. The mixture was partitioned

with chloroform (20 mL) and water (20 mL). The organic layer

was dried over MgSO₄and concentrated and the residue was

purified with SiO₂column chromatography (dichloromethane/

1

acetone, 9:1) to give 94a (0.53 g, 69%). H NMR (CDCl₃, 400

MHz) δ 1.96 (p, 2H, J = 7.5 Hz), 2.38 (t, 2H, J = 7.5 Hz), 2.68 (t,

2H, J= 7.5 Hz), 7.22 (d, 2H, J =8.5Hz), 7.48(d, 2H, J=8.5Hz),

7.81 (br s, 1H).

N,N-Dimethyl-4-[4-(2,2,2-trifluoroacetylamino)phenyl]butyra-

mide, 94b. Thionylchloride (71 mg, 0.60 mmol) was added slowly

to a solution of 94a (0.15 g, 0.55 mmol) in DMF (5 mL) cooled at

-5 °C. The mixture was stirred for an additional 1 h at -5 °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 h. Ethyl acetate (50 mL) was

added to the mixture, which was washed with brine (2 ꢀ 50 mL).

The organic layer was dried over MgSO₄and concentrated to

Acknowledgment. We thank CaPCURE and the Prostate

Cancer Foundation for generous financial support. S.O. also

acknowledges support from NIH SPORE Grant 5P50CA-

92131. C.L.S. is an Investigator of the Howard Hughes

Medical Institute and a Doris Duke Distinguished Clinical

Scientist.

1

yield 94b (0.17 g, quantitative) as a yellowish solid. H NMR

2796 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 7

Jung et al.

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has six animals. After they reach this tumor volume, they are given

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(15) LNCaP in 10% FBS, split on day 1, add drugs on day 5, harvest on

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Article Doi

Structure-activity relationship for thiohydantoin androgen receptor antagonists for castration-resistant prostate cancer (CRPC)

DOI: 10.1021/jm901488g

Source and publish data:

Authors:

Article abstract of DOI:10.1021/jm901488g

Full text of DOI:10.1021/jm901488g

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