Design and synthesis of 1,2,4-oxadiazole derivatives as non-steroidal 5α-reductase inhibitors

Article abstract of DOI:10.1002/jccs.200200014

The purpose of this study was to synthesize compounds in which the 1,2,4-oxadiazole moiety replaced the amide bond of ONO3805 and to evaluate its 5α-reductase inhibitory activity as a potential benign prostatic hyperplasia therapeutic target. Four 1,2,4-o

Full text of DOI:10.1002/jccs.200200014

JournaloftheChineseChemicalSociety, 2002, 49, 83-89
83
DesignandSynthesisof1,2,4-OxadiazoleDerivativesasNon-steroidal
5 -ReductaseInhibitors
Chih-Shiang Changa (
), Wai Ming Kanb (
), Chiu-Liang Chenb (
),
K. C. Wanga (
) and Ji-Wang Cherna* (
)
aSchoolofPharmacy,CollegeofMedicine,NationalTaiwanUniversity,Taipei,Taiwan100,R.O.C.
bDepartmentofPharmacology,CollegeofMedicine,NationalChengKungUniversity,
Tainan, Tai wan 701, R.O.C.
The pur pose of this study was to syn the size com pounds in which the 1,2,4-oxadiazole moi ety replaced
the am ide bond of ONO3805 and to eval u ate its 5 -reductaseinhibitoryactivityasapotentialbenignpros-
tatichyperplasiatherapeutictarget.Four1,2,4-oxadiazolederivatives,1, 2, 8, and20,wereevaluatedinvitro
against 5 -reductase of rat liver microsome. The pre pared 1 and 2possessedsimilarbindingaffinity(Ki)to
that of ONO3805. There fore, the use of 1,2,4-oxadiazole ring as sur ro gate of the am ide bond inONO3805 has
asuccessfulresultinthisstudy.Itleadsnotonlytoenhancechemicalstabilitybutalso tomaintainmeaningful
in hib i tory ac tiv ity. The bu tyric acid moi ety of these in hib i tors is con sid ered to playan im por tant role in
mimicing the phos pho ric acid por tion of coenzyme-NADPH in in ter act ing with the ac tive site of 5 -
reductase.
INTRODUCTION
Steroid 5 -reductase is a membrane-associated en-
of the am ide bond in ONO3805 as po ten tial 5 -reductase in-
hibitorsweredesignedandsynthesized(Fig.1).
zyme,whichcatalyzesthereductionoftestosteronetothebi-
o log i cally more ac tive dihydrotestosterone (DHT) in the
presenceofNADPH.1 ElevatedDHThasbeenimplicatedasa
causativefactorofskindisordersandbenignprostatichyper-
plasia(BPH).2 Therefore,inhibitionof5 -reductase is an at-
tractivetargetfortherapeuticinterventionindisordersasso-
ci ated with el e vated lev els of DHT such as BPH,3,4 acne,5
male pat tern bald ness,6 and hirsutism.7 A num ber of 5 -
reductase inhibitors have been reported, including both
steroidal8 andnonsteroidalinhibitors.9 Starting our re search
pro gram to find a new 5 -reductaseinhibitor,wefocusedour
at ten tion on the de vel op ment of novel nonsteroidal com-
pounds, es pe cially ONO3805. Ac cord ing to the bioiso s ter-
ism,10 some re search groups have suc cess fully re placed the
esteroramidegroupofbiologicallyactivecompoundsbyan
1,2,4-oxadiazole moi ety to im prove in vivoefficacy.11 The
intramolecular acid cat a lyzed the hy dro ly sis of the am ide
bond12,13 even though the hy dro ly sis ex hib its a half-life of 7
yearsunderneutralconditionsandatroomtemperature.14 In
ad di tion, we found that the am ide bond of ONO3805 was par-
tially cleaved in a DMSO-d6 so lu tion at 37 oC in four months.
Re place ment of the am ide bond with a 1,2,4-oxadiazole ring
for in creas ing hydrolytic re sis tance,15 metabolicstability,
andimprovingpharmacokineticperformance16 has been re-
ported. Herein, de riv a tives with 1,2,4-oxadiazole sur ro gate
RESULTS AND DISCUSSION
The syn the sis of tar get com pound 1 is il lus trated in
Fig. 1. The am ide link age of ONO3805 was re placed
with 1,2,4-oxadiazole bioisostere.

84 J. Chin. Chem. Soc., Vol. 49, No. 1, 2002
Chang et al.
Scheme I. The cou pling of sub sti tuted amidoxime 5 and 4-
[1- (4-isobutylphenyl)ethoxy]-2,3-dimethylbezoic acid (6)
us ing 1,1 -carbonyldiimidazole in THF af forded 7, which
was fol lowed by cyclization in diglyme at 110 oC to form the
1,2,4-oxadiazolederivative8.17 Finally, treat ment of 8 with
al ka line so lu tion gave 89% yield of 1. The re quired 5 was
prepared from 2-cyanophenol by alkylation with ethyl -
bromobutyrate and subsequent treatment with hydroxyl-
amine. The other frag ment 6 waspreparedbyamodifiedpro-
cedurefromHarris.18 The oxadiazole iso mer 2 was pre pared
by ini tial treat ment of sub sti tuted amidoxime 14 and diester
16with NaH in THF to af ford ethyl bu tyr ate17and a mix ture
containing18 and 19. Finally, hydrolysisof 17 and the mix-
ture of 18 and 19 with aque ous lith ium hy drox ide af forded
1,2,4-oxadiazole 2 and 20, respectively(SchemeIII). There-
quired amidoxime 14 was pre pared as shown in Scheme II.
Firstly, the treat ment of 2,3-dimethyl-p-anisaldehyde with
hydroxylamine-o-sulfonic acid in a mixed sol vent of wa ter
and acetonitrile (1:1) to pro duce benzonitrile10,19 which was
demethylation with BBr3 to give 11. Then, 11 cou pled with
1-(4-isobutylphenyl)eth a nol by Mitsunobu re ac tion to fur-
nish benzonitrile 12.20 Sub se quent treat ment of12 with hy-
droxylamine in Et3N yielded am ide13 (48%) and amidoxime
14 (45%). The other frag ment 16 was pre pared from methyl
salicylate by alkylation with ethyl -bromobutyrate.
Scheme II
Scheme III
Theprepared1,2,4-oxadiazolederivatives,1, 2, 8, and
20,weresubjectedagainst5 -reductaseinhibitionassay(Ta-
ble 1). The 1,2,4-oxadiazole iso mers 1 and 2 pos sessed high
Scheme I
inhibitoryactivityto5 -reductase with Ki = 15.63 and 22.36
nM, respectively, whichiscomparablewiththatofONO3805
(Ki = 8.46 nM). These re sults dem on strated that the 1,2,4-
oxadiazole moi ety is as use ful as the am ide bioisostere in
ONO3805 chem i cal sta bil ity. Holt et al.21 pro posed that the
butanoic acid moi ety of ONO3805 was lo cal ized in the re-
gion of the phos phate groups of NADPH, and the lipophilic
part was ori en tated in the re gion of the steroidal rings C and D
to oc cupy the hy dro pho bic pocket of 5 -reductase. In ter est-
ingly, com pound8lostactivityagainst5 -reductase (Ki > 50
M). This re sult might in di cate that 8 could not fit into the
neg a tive-ionizable re gion be cause the carboxylic func tion
was masked by an ethyl group. Sur pris ingly, com pound 20

1,2,4-OxadiazoleDerivatives
J. Chin. Chem. Soc., Vol. 49, No. 1, 2002 85
Ethyl 4-(2-cyanophenoxy)butyrate (4)
Table 1. Binding Affinity Against 5 -Reductase of Rat Liver
Microsome
To a stirred so lu tion of 2-cyanophenol (20 g, 0.17 mol)
and K2CO3 (25.84 g, 0.19 mol) dis solved in dried acetonitril
(500 mL) was added -bromo-n-butyric acid ethyl es ter (32.8
g,0.17mol)undernitrogen.Themixturewasheatedatreflux
for11h, filtered, andconcentratedinvacuo. Theresiduewas
subjectedtochromatography(silicagel,n-hexane/ethylace-
tate = 9/1, Rf = 0.19) to af ford4(36.4 g, 92 %) as a col or less
Compounds
Binding Affinity
Ki ± SD (nM)a
1
2
8
20
15.63 ± 9.69
22.36 ± 6.30
NAb
NAb
1
ONO3805
8.46 ± 5.92
oil. H NMR (200 MHz, CDCl3) 7.42-7.51 (m, 2H, ArH),
a Values are in four experiments.
6.90-6.98 (m, 2H, ArH), 4.03-4.14 (m, 4H, CH2), 2.52 (t, 2H,
J = 7.2 Hz, COCH2), 2.04-2.17 (m, 2H, CH2), 1.20 (t, 3H, J =
7.1 Hz, CH3); 13C NMR (50 MHz, CDCl3) 172.9, 160.3,
134.2, 133.6, 120.7, 116.2, 112.1, 101.8, 69.5, 60.4, 30.2,
24.0, 14.0; MS (EI) m/z 233 (M+); Anal. Calcd. for
C13H15NO3: C, 66.94; H, 6.48; N, 6.00. Found: C, 66.88; H,
6.46; N, 5.98.
b Not active at concentration of 50 M.
didnotshowanyinhibitoryactivity. Thereasonmightbethat
the ben zoic acid moi ety is un able to func tion as the butanoic
acid moi ety in1 and2infittingintothenegative-ionizablere-
gionasproposed.
The initial result showed that the 1,2,4-oxadiazole
bioisosteres, 1 and 2,couldsuccessfullyretaininhibitoryac-
tiv ity of 5 -reductasebycomparisonwiththeamidelinkage
of ONO3805. The oxadiazole 8 lost its ac tiv ity when but-
anoic acid moi ety was masked by an ethyl group, in di cat ing
that the carboxylic acid group was im por tant to mimic the
phosphoricacidportionofNADPH.Moreover,the2-alkoxy-
benzoic acid moi ety also could not fit the ac tive site of 5 -
reductase. There fore, the neg a tive ionizable re gion of 5 -
reductase is con sid ered to play an im por tant role in its cat a-
lyticactivityofreducingtestosteronetoDHT.
Ethyl 4-[2-[amino(hydroxyimino)methyl]phenoxy]-
butyrate (5)
To a stirred so lu tion of hydroxylamine hy dro chlo ride
(2.98 g, 42.9 mmol) in ab so lute eth a nol (250 mL) was added
K2CO3 (5.92g,42.86mmol)atroomtemperatureundernitro-
gen for 10 min, and then added a so lu tion of 4 (5.0 g, 21.4
mmol) in ab so lute eth a nol (10 mL). The mix ture was heated
at re flux for 11 h and then evap o rated in vacuo. The res i due
was sub jected to chro ma tog ra phy (sil ica gel, CHCl3, Rf =
0.15) to give 5 (2.69 g, 47%) as a col or less oil. 1H NMR (200
MHz, CDCl3) 7.61 (dd, 1H, J = 7.63 Hz, ArH), 7.26-7.31
(m, 1H, ArH), 6.87-6.98 (m, 2H, ArH), 5.37 (br s, 2H, NH2),
4.01-4.16 (m, 4H, CH2), 2.48 (t, 2H, J = 7.2 Hz, CH2), 2.08-
2.14 (m, 2H, CH2), 1.21 (t, 3H, J = 7.1 Hz, CH3); 13C NMR
(50 MHz, CDCl3) 173.9, 156.3, 152.2, 130.8, 129.7, 121.1,
120.9, 112.5, 67.6, 60.5, 30.7, 24.4, 14.1; MS (EI) m/z 266
(M+); Anal. Calcd. for C13H18N2O4: C, 58.64; H, 6.81; N,
10.52. Found: C, 58.60; H, 6.58; N, 10.30.
EXPERIMENTALSECTION
All of the sol vents and chem i cals were re agent grade.
Nor mal phase sil ica gels of 9385 (230-400 mesh) for col umn
chromatography, andTLCplatesof573(Si60withF 254, 0.25
mn) were pur chased from E. Merck A. G., Darmstadt, Ger-
many. [4-14C]-Testosterone(2.10GBa/mmol) was pur chased
from NEN Life Sci ence Prod ucts, Inc., Boston, U.S.A. Melt-
ing points mea sured on a Buchi 510 melt ing point ap pa ra tus
wereuncorrected.IRspectrawererecordedonaJascoA-100
infraredspectrophotometer.1H- and 13C-NMR spec tra were
ob tained on Bruker DPX-200 or Bruker AMX-400 spec tro-
photometersusingthesolventpeaksasreferencestandards.
EIMS spec tra were re corded on Finnigan Mat GCQTM GC/
MS and HREIMS on JEOL JMS-HX-100 mass spec trom e ter.
El e men tal anal y ses were car ried out on a Perkin-Elmer 240
elementalanalyzerandtheresultswerewithin±0.4%oftheo-
reticalvalues.
4-[2-[4-[1-(4-Isobutylphenyl)ethoxy]-2,3-dimethyl-N-
benzoyloxy]carbamimidoylphenoxy]butanoic acid ethyl
ester (7)
To a stirred so lu tion of 6 (300 mg, 0.92 mmol) and
1,1 - carbonyldiimidazole (193.9 mg, 1.20 mmol) dis solved
in dried THF (30 mL) was added 5 (318.2 mg, 1.19 mmol) at
room tem per a ture. The mix ture was stirred for 18 h un der ni-
trogen. Afterthemixturewasreactedcompletely(indicated
by phosphomolybdic acid on TLC plate dis played an or ange
spot),purifiedbychromatography(silicagel,n-hexane/ethyl
acetate=7/3, Rf = 0.17) to af ford 7 which was recrystallized
from ether to give a white solid (210 mg, 40%). mp 103-104

86 J. Chin. Chem. Soc., Vol. 49, No. 1, 2002
Chang et al.
oC; 1H NMR (200 MHz, CDCl3) 8.87 (dd, 1H, J = 7.2; 1.3
Hz, ArH), 6.89-7.54 (m, 8H, ArH), 6.59 (d, 1H, J = 8.7 Hz,
ArH), 5.58 (br s, 2H, NH2), 5.33 (q, 1H, J = 6.3 Hz, CH),
4.04-4.15 (m, 4H, OCH2), 2.51 (s, 3H, CH3), 2.41-2.47 (m,
4H, -COCH2; ArCH2), 2.28 (s, 3H, CH3), 2.05-2.18 (m, 2H,
CH2), 1.75-1.89 (m, 1H, CH), 1.63 (d, 3H, J = 6.5 Hz, CH3),
1.20 (t, 3H, J = 7.1 Hz, CH3), 0.87 (d, 6H, J = 6.6 Hz, CH2);
13C NMR (50 MHz, CDCl3) 173.0, 165.8, 158.2, 156.7,
156.3, 141.0, 140.1, 139.4, 131.8, 130.8, 129.3, 128.3, 127.0,
125.1, 122.7, 121.3, 119.6, 112.5, 109.9, 109.6, 76.1, 67.8,
60.6, 45.1, 30.9, 30.1, 24.5, 24.5, 22.4, 17.2, 14.2. 12.2; MS
(EI) m/z 574.5 (M+); HRMS (EI) Calcd. for C 34H42N2O6 (M+):
574.3042. Found: 574.3035.
7.26 (m, 2H, ArH), 6.99-7.11 (m, 4H, ArH), 6.68 (d, 1H, J =
8.9 Hz, ArH), 5.36 (q, 1H,J = 6.1 Hz, CH), 4.18 (t, 2H,J = 5.8
Hz, O-CH2), 2.70 (t, 2H, J = 7.1 Hz, -COCH2), 2.64 (s, 3H,
CH3), 2.42 (d, 2H, J = 7.1 Hz, ArCH2), 2.32 (s, 3H, CH3).
2.10-2.22 (m, 2H, CH2), 1.71-1.91 (m, 1H, CH), 1.64 (d, 3H,
J = 6.7 Hz, CH3), 0.86 (d, 6H,J = 6.6 Hz, CH3); 13C NMR (50
MHz, CDCl3) 176.9, 176.0, 166.7, 158.7, 157.1, 141.1,
139.9, 139.0, 132.2, 131.4, 129.4, 129.0, 127.2, 125.1, 120.9,
116.4, 116.2, 112.6, 110.7, 76.4, 66.7, 45.1, 30.6, 30.1, 24.4,
24.4, 22.4, 17.5, 12.3; MS (EI) m/z 528.3 (M+), 368.1, 161.1
(base peak); HRMS (EI) Calcd. for C H36N2O5 (M+):
32
528.2624. Found 528.2636.
2,3-Dimethyl-4-methoxybenzonitrile (10)
4-[2-[5-[4-[1-(4-Isobutylphenyl)ethoxy]-2,3-dimethylphenyl]-
[1,2,4]oxadiazol-3-yl]phenoxy]butanoic acid ethyl ester
(8)
To a stirred so lu tion of 2,3-dimethyl-p-anisaldehyde (5
g, 30.5 mmol) dis solved in acetonitrile (200 mL) was added
dropwise an aque ous so lu tion (200 mL) of hydroxylamine-
o- sulfonic acid (5.17 mg, 45.7 mmol) at room tem per a ture
dur ing 30 min pe riod, and then the mix ture was heated at 60
oC for 18 h. Af ter cool ing, the mix ture was neu tral ized with
10% NaOH and evap o rated in vacuo. The res i due was ex-
tracted with CHCl3 (5 60 mL), dried over Na2SO4,filtered,
con cen trated, chromatographed (sil ica gel, n-hexane/ethyl
acetate=96/4,Rf = 0.32), and recrystallized from acetonitrile
to fur nish 10 as a white solid (4.69 g, 96%). mp 40 oC; IR
(KBr): 2950, 2840, 2225, 1590, 1580 cm-1; 1H NMR (200
MHz, CD3OD) 7.41 (d, 1H, J = 8.6 Hz, ArH), 6.71 (d, 1H, J
= 8.6 Hz, ArH), 3.84 (s, 3H, OCH3), 2.41 (s, 3H, CH3), 2.11
(s, 3H, CH3); 13C NMR (50 MHz, CDCl3) 160.5, 141.1,
131.4, 126.3, 119.3, 107.9, 104.7, 55.6, 18.1, 11.7; MS (EI)
m/z 161 (M+), 146; Anal. Calcd. for C10H11NO: C, 74.51; H,
6.88; N, 8.69. Found: C, 74.63; H, 7.16; N, 8.40.
Asolutionof7(220 mg, 0.38 mmol) dis solved in dried
2-methoxyethyl ether (diglyme, 10 mL) was heated at 110oC
un der ni tro gen for 10 h. Af ter cool ing, the so lu tion was con-
centratedinvacuo, purifiedbychromatography(silicagel,
n-hexane/ethyl ac e tate = 9/1) to give 8 (Rf = 0.27), and fol-
lowed by recrystallization from eth a nol to fur nish a white
o
solid (200 mg, 95%). mp 81.5-82.0 C; 1H NMR (200 MHz,
CDCl3) 8.05 (dd, 1H,J = 7.7; 1.8 Hz, ArH), 7.76 (d, 1H, J =
8.8 Hz, ArH), 7.0-7.39 (m, 7H, ArH), 6.69 (d, 1H,J = 8.8 Hz,
ArH), 5.37 (q, 1H, J = 6.4 Hz, CH), 4.04-4.20 (m, 4H, CH2),
2.67 (s, 3H, CH3), 2.64 (t, 2H, J = 7.0 Hz, -COCH2), 2.43 (d,
2H, J = 7.2 Hz, ArCH2), 2.33 (s, 3H, CH3), 2.10-2.27 (m, 2H,
CH2), 1.76-1.89 (m, 1H, CH), 1.65 (d, 3H, J = 6.4 Hz, CH3),
1.20 (t, 3H, J = 7.1 Hz, CH3), 0.87 (d, 6H, J = 6.6 Hz, CH3);
13C NMR (50 MHz, CDCl3) 175.7, 173.4, 166.9, 158.5,
157.3, 141.0, 139.9, 138.9, 131.9, 131.3, 129.3, 128.9, 127.1,
125.1, 120.6, , 116.7, 116.4, 112.7, 110.6, 76.1, 67.5, 60.3,
45.0, 30.5, 30.1, 24.4, 24.4, 22.3, 17.5, 14.2, 12.3; MS (EI)
m/z 556.5 (M+), 511, 396 (base peak); HRMS (EI) Calcd. for
C34H40N2O5 (M+): 556.2937. Found 556.2928.
2,3-Dimethyl-4-hydroxybenzonitrile (11)
To a stirred so lu tion of 10 (4.69 g, 29.1 mmol) dis-
solved in dry di chloro methane (200 mL) was added BBr3
(21.9 g, 87.3 mmol) at 0oCundernitrogen.Thereactionmix-
o
ture was warmed up to 50 C for 6 h, and then poured into
4-[2-[5-[4-[1-(4-Isobutylphenyl)ethoxy]-2,3-dimethylphenyl]-
[1,2,4]oxadiazol-3-yl]phenoxy]butanoic acid (1)
crushed-ice while stir ring vig or ously. The re sulted mix ture
was ex tracted with CHCl3 (4 150 mL), dried over Na2SO4,
filtered, decoloured by active char coal, and fol lowed by
recrystallization from CHCl3 to give 11 as a white solid (4.21
g, 98%). mp 134 oC; IR (KBr): 3285, 2227 cm-1; 1H NMR
(400 MHz, CDCl3) 7.32 (d, 1H, J = 8.4 Hz, ArH), 6.73 (d,
1H, J = 8.4 Hz, ArH). 6.19 (s, 1 H, OH). 2.44 (s, 3H, CH3),
2.17 (s, 3H, CH3); 13C NMR (50 MHz, CDCl3) 157.8, 142.5,
131.3, 124.4, 119.3, 113.3, 104.4, 18.3, 11.7; MS (EI) m/z 147
(M+); Anal. Calcd. for C9H9NO: C, 73.45; H, 6.16; N, 9.52.
Found: C, 73.10; H, 6.34; N, 9.31.
To a stirred sus pen sion of8 (500 mg, 0.89 mmol) in
meth a nol (150 mL) was added 10% NaOH (15 mL) at room
tem per a ture. And the mix ture was stirred for 5 days. Af ter
cool ing in an ice-bath, the mix ture was acid i fied with 6 N
HCl to pH 3-4 un der vig or ous stir ring, fil tered, and washed
with wa ter. The solid was recrystallized from eth a nol to af-
ford 1 as a white solid (408 mg, 89%). mp 143-144 oC; 1H
NMR (200 MHz, CDCl3) 8.03 (dd, 1H, J = 7.7 Hz, ArH),
7.73 (d, 1H, J = 8.7 Hz, ArH), 7.39-7.47 (m, 1H, ArH), 7.22-

1,2,4-OxadiazoleDerivatives
J. Chin. Chem. Soc., Vol. 49, No. 1, 2002 87
2,3-Dimethyl-4-[1-(4-isobutylphenyl)ethoxy]benzonitrile
(12)
cm-1; 1H NMR (200 MHz, CD3OD) 7.23 (d, 2H, J = 8.0 Hz,
ArH), 7.07 (d, 2H, J = 8.0 Hz, ArH), 6.90 (d, 1H, J = 8.5 Hz,
ArH), 6.55 (d, 1H, J = 8.5 Hz, ArH), 5.25 (q, 1H, J = 6.4 Hz,
OCH), 4.69 (br, 2H, NH2), 2.42 (d, 2H,J = 7.1 Hz, CH2), 2.28
(s, 3H, CH3), 1.88 (s, 3H, CH3), 1.75-1.88 (m, 1H, CH), 1.55
(d, 3H, J = 6.4 Hz, CH3), 0.86 (d, 3H, J = 6.6 Hz, CH3); 13C
NMR (50 MHz, CDCl3) 156.5, 153.8, 140.8, 140.4, 136.7,
129.3, 126.8, 126.4, 125.3, 125.2, 110.5, 76.0, 45.1, 30.2,
24.4, 22.4, 16.9, 12.2; MS (ESI, acetic acid) m/z 341.1
(MH+); Anal. Calcd. for C21H28N2O2: C, 74.08; H, 8.29; N,
8.23. Found: C, 74.12; H, 8.10; N, 7.98.
To a stirred soution of 11 (300 mg, 2.04 mmol), 1-(4-
isobutylphenyl)eth a nol (472.8 mg, 2.65 mmol) and tri phen-
yl phosphine (802.6 mg, 3.06 mmol) dis solved in THF (30
mL) was added diethylazodicarboxylate (532.9 mg, 3.06
mmol)atroomtemperatureundernitrogen.After8h,thecol-
ourofsolutionwaschangedfromcolorlesstoblackishgreen,
con cen trated, triturated with ether, fil tered, and then the fil-
tratewaspurifiedbychromatography(silicagel, n-hexane,
Rf = 0.1), fol lowed by recrystallization from n-hexane to give
12 as a white solid (305 mg, 49%). mp 66 oC; IR (KBr): 2960,
2930, 2875, 2225, 1590 cm-1. 1H NMR (200 MHz, CD3OD)
7.09-7.27 (m, 5H, ArH), 6.60 (d, 1H,J = 8.7 Hz, CH), 5.33 (q,
1H, J = 6.4 Hz, CH), 2.44 (d, 2H, J = 7.1 Hz, CH2), 2.44 (s,
3H, CH3), 2.26 (s, 3H, CH3), 1.77-1.91 (m, 1H, CH), 1.65 (d,
3H, J = 6.4 Hz, CH3), 0.88 (d, 6H,J = 6.6 Hz, CH3); 13C NMR
(50 MHz, CDCl3) 158.9, 141.2, 141.1, 139.5, 131.0, 129.3,
126.8, 125.0, 119.2, 110.8, 104.5, 76.3, 45.0, 30.0, 24.3, 22.2,
18.2, 12.0; MS (EI) m/z 307 (M+); Anal. Calcd. for C21H25NO:
C, 82.04; H, 8.20; N, 4.56. Found: C, 81.71; H, 8.21; N, 4.51.
2-(3-Ethoxycarbonylpropoxy)benzoic acid methyl ester
(16)
To a stirred sus pen sion of15 (10 g, 65.72 mmol) and
K2CO3 (18.2 g, 131.44 mmol) in dried acetonitrile (300 mL)
was added -bromobutyric acid ethyl es ter (12.8 g, 65.72
mmol) at re flux for 24 h un der ni tro gen. The hot mix ture was
fil tered, washed with acetonitrile, and con cen trated. The re-
sultingresiduewassubjectedtochromatography(silicagel,
n-hexane/ethyl ac e tate = 95/5) to ob tain16 (17.1 g, 98%) as a
yellowishoil.1H NMR (400 MHz, CDCl3) 7.74 (dd, 1H,J =
1.8, 7.62 Hz, ArH), 7.37-7.41 (m, 1H, ArH), 6.90-6.94 (m,
2H, ArH), 4.09 (q, 2H, J = 7.2 Hz, CH2), 4.05 (t, 2H, J = 6.1
Hz, CH2). 3.84 (s, 3H, CH3), 2.54 (t, 2H, J = 7.3 Hz, CH2),
4-[1-(4-Isobutylphenyl)ethoxy]-2,3-dimethylbenzamide
(13) and 4-[1-(4-Isobutylphenyl)ethoxy]-2,3- dimethyl-
benzamidoxime (14)
To a stirred so lu tion of hydroxylamine hy dro chlo ride
(2.03 g, 29.28 mmol) and triethylamine (2.96 g, 29.28 mmol)
dissolvedinabsoluteethanol(200mL)wasadded12 (0.90 g,
2.93 mmol). The mix ture was heated at re flux for 8 days un-
der ni tro gen. The mix ture was then evap o rated in vacuo and
the res i due was sub jected to chro ma tog ra phy (sil ica gel,
n-hexane/ethyl ac e tate = 7/3). The fast-moving frac tion was
collectedtogive13 (Rf = 0.33) which was recrystallized from
CHCl3 to af ford a white solid (0.32 g, 48%). mp 125-126oC;
IR (KBr): 3383, 3193, 2920, 1706, 1646, 1596, 1464 cm-1; 1H
NMR (200 MHz, CD3OD) 7.23 (d, 2H, J = 8.1 Hz, ArH),
7.07 (d, 2H, J = 8.1 Hz, ArH), 7.04 (d, 1H, J = 8.5 Hz, ArH),
6.64 (d, 1H, J = 8.5 Hz, ArH), 5.39 (q, 1H, J = 6.4 Hz, CH),
4.63 (br s, 2H, NH2), 2.41 (d, 2H, J = 7.2 Hz, CH2), 2.31 (s,
3H, CH3), 2.25 (s, 3H, CH3), 1.74-1.87 (m, 1H, CH), 1.60 (d,
3H, J = 6.4 Hz, CH2), 0.86 (d, 6H,J = 4.7 Hz, CH3); 13C NMR
(50 MHz, CD3OD) 175.7, 157.1, 141.1, 140.9, 135.6,
129.3, 126.7, 125.5, 125.2, 110.6, 76.1, 45.1, 39.5, 23.8, 21.7,
16.1, 11.2; LCMS (ESI, ace tic acid) m/z 326.2 (MH+); Anal.
Calcd. for C21H27NO2: C, 77.50; H, 8.36; N, 4.30. Found: C,
77.55; H, 8.56; N, 4.24. The low-moving frac tion was col-
lected to give 14 (Rf = 0.18) which was recrystallized from
acetonitrile to give a white solid (0.31 g, 45%). mp 140-141
oC; IR (KBr): 3490, 3381, 3250, 2927, 1640, 1595, 1494
2.07-2.13 (m, 2H, CH2), 1.21 (t, 3H, J = 7.3 Hz, CH3); 13
C
NMR (50 MHz, CDCl3) 173.2, 166.7, 158.2, 133.3, 131.6,
120.2, 120.2, 113.1, 67.4, 60.3, 51.8, 30.4, 24.4, 14.1. Anal.
Calcd. for C14H18O5: C, 63.15; H, 6.81. Found: C, 63.30; H,
6.78.
4-[2-[3-[4-[1-(Isobutylphenyl)ethoxy]-2,3-dimethylphenyl]-
[1,2,4]oxadiazol-5-yl]phenoxy]bu tyric acid (2) and
2-[3-[3-[4-[1-(Isobutylphenyl)ethoxy]-2,3-dimethylphenyl]-
[1,2,4]oxadiazol-5-yl]propoxy]benzoic acid (20)
Toamixturesolutionof14 (1 g, 2.94 mmol) dis solved
inanhydrousTHF(20mL)containing4Åpowderedmolecu-
lar sieves (3 g) was stirred at room tem per a ture for 30 min un-
der ni tro gen, and then so dium hy dride was added (17.54 mg
of 80% dis per sion in oil, 3.23 mmol). The mix ture was then
heated at 60 oC for 1 h. Af ter cool ing, to the mix ture was
added a so lu tion of16 (2 g, 7.51 mmol) in THF (10 mL) and
the re sult ing mix ture was then heated at 60oC for 3 h. The so-
lutionwasevaporatedinvacuoandtheresiduewassubjected
tochromatography(silicagel,n-hexane/CHCl3 = 6/4). The
fast-moving band was col lected to af ford17 (Rf = 0.28) as a
yel low ish oil (145 mg, 9%). IR (neat): 2955, 1734, 1595,
1
1546, 1464 cm-1; H NMR (200 MHz, CD3OD) 8.11 (dd,
1H, J = 1.8; 8.0 Hz, ArH), 7.62 (d, 1H, J = 8.6 Hz, ArH),

88 J. Chin. Chem. Soc., Vol. 49, No. 1, 2002
Chang et al.
7.44-7.53 (m, 1H, ArH), 7.26 (d, 2H, J = 6.2 Hz, ArH),
7.01-7.11 (m, 4H, ArH), 6.68 (d, 1H, J = 8.7 Hz, ArH), 5.35
(q, 1H, J = 6.3 Hz, OCH), 4.04-4.20 (m, 4H, CH2), 2.64 (t,
2H, J = 7.2 Hz, CH2), 2.55 (s, 3H, CH3), 2.43 (d, 2H, J = 7.1
Hz, CH2), 2.32 (s, 3H, CH3), 2.12-2.26 (m, 2H, CH2),
1.74-1.89 (m, 1H, CH), 1.65 (d, 3H, J = 6.3 Hz, CH3), 1.20 (t,
3H, J = 7.1 Hz, CH3), 0.88 (d, 6H,J = 6.6 Hz, CH3); 13C NMR
(50 MHz, CDCl3) 173.8, 173.2, 169.4, 157.7, 157.3, 140.9,
140.3, 137.9, 133.8, 131.6, 129.3, 128.5, 126.8, 125.2, 120.7,
119.2, 113.9, 113.0, 110.7, 76.0, 67.6, 60.3, 45.1, 30.5, 30.1,
24.5, 24.3, 22.4, 17.7, 14.2, 12.3. Com pound17 was used for
thenextreactionwithoutfurtherpurification.Subsequently,
to the so lu tion of17(60 mg, 0.11 mmol) dis solved in dioxane
(8 mL) was added 3 mL of 1 N LiOH at 80oC for 30 min un der
vigorousstirring. Aftercooling, themixturewasacidified
with4NHCltopH3-4,evaporated,andsubjectedtochroma-
tog ra phy (sil ica gel, CHCl3/CH3OH = 97/3) to give 2 which
was recrystallized from acetonitrile to af ford a white soild
(35 mg, 60%). mp 103-104 oC; IR (KBr): 2955, 1709, 1595,
1547, 1463, 1339, 1265 cm-1; 1H NMR (200 MHz, CD3OD)
8.10 (dd, 1H, J = 1.61; 6.91 Hz, ArH), 7.61 (d, 1H, J = 8.66
Hz, ArH), 7.44-7.52 (m, 1H, ArH), 7.26 (d, 2H, J = 7.4 Hz,
ArH), 6.99-7.10 (m, 4H, ArH), 6.69 (d, 1H, J = 8.7 Hz, ArH),
5.35 (q, 1H,J = 6.3 Hz, OCH), 4.17 (t, 2H,J = 5.9 Hz, OCH2),
2.71 (t, 2H, J = 7.1 Hz, CH2), 2.53 (s, 3H, CH3), 2.42 (d, 2H, J
= 7.2 Hz, CH2), 2.31 (s, 3H, CH3), 2.12-2.27 (m, 2H, CH2),
1.76-1.89 (m, 1H, CH), 1.63 (d, 3H,J = 6.3 Hz, CH3), 0.87 (d,
6H, J = 6.6 Hz, CH3); 13C NMR (50 MHz, CDCl3) 178.1,
173.8, 169.4, 157.6, 157.4, 140.9, 140.3, 137.9, 133.8, 131.6,
129.3, 128.5, 126.8, 125.3, 120.9, 119.2, 113.8, 113.0, 110.8,
76.0, 67.3, 45.1, 30.9, 30.1, 24.3, 24.3, 22.4, 17.7, 12.3;
LCMS (ESI, Et3N) m/z 527.3 (M-1), 441.3 (base peak);
HRMS (ESI) Calcd. for C32H35N2O5 (M+): 528.2624. Found:
528.2618.
158.0, 157.6, 141.4, 140.7, 138, 135.4, 134.2, 129.7, 128.9,
127.4, 125.6, 122.8, 119.0, 118.6, 113.1, 111.1, 110.0, 76.5,
68.8, 45.5, 30.6, 26.2, 24.8, 23.2, 22.8, 18.2, 12.8; LCMS
(ESI, Et3N) m/z 527.4 (M-1); HRMS (FAB) Calcd. for
C32H37O5N2 (MH+): 529.2701. Found: 529.2702.
Rat 5 -Reductase Inhibition Assay
The in hi bi tion as say was per formed ac cord ing to that
re ported by Rus sell et. al22 withsomeminormodifications.
Briefly, in 200 L of 20 mM so dium phos phate buffer (pH
6.5),containing5 M [4-14C]-testosterone(2.10GBa/mmol),
200 M of NADPH and 0.05% Tween-80, var i ous amounts of
the tested com pound was added. The re ac tion was ini ti ated
with the ad di tion of 20 g of rat liver microsome and in cu-
o
bated at 37 Cfor30minutes.Afterincubation,thereaction
was ter mi nated with the ad di tion of 1 mL of ac e tone. The
supernatant was re moved and dried. The res i due was spot ted
on a TLC plate (sil ica gel) and de vel oped with di chloro-
methane/ether 1:1. Radioactivities were recorded with a
Bioscan -ray de tec tor. The amount of prod uct formed was
calculatedcorrespondingly.
ACKNOWLEDGMENTS
The re search grants from the Na tional Sci ence Coun cil
(NSC 86-2314-B-002-089-M38 and NSC 86-2314-B-002-
155-M38) and the National Health Research Institutes
(NHRI-GT-EX89S919C)arehighlyappreciated.
Received October 4, 2001.
Key Words
Testosterone; Dihydrotestosterone; 5 -Reductase;
Benign prostatic hyperplasia; Oxadiazole.
The slow-moving band was col lected to give a mix ture
of 18 (Rf = 0.15) and 19 (Rf = 0.17) as a yel low ish oil (0.7 g)
and then it was hy dro lyzed with 1 N LiOH for 1 h at 60 oC.
Afteritwasacidifiedandevaporated,thesolutionwaseluted
by a mix ture of CHCl3/CH3OH (97/3) to af ford 20 (Rf = 0.14)
as a yel low ish oil (132 mg, two steps gave 9% yield). IR
(neat): 2954, 1694, 1600, 1460 cm-1; 1H NMR (200 MHz,
CD3OD) 8.13 (dd, 1H, J = 1.7; 7.8 Hz, ArH), 7.47-7.55 (m,
2H, ArH), 7.23 (d, 2H, J = 8.0 Hz, ArH), 7.0-7.14 (m, 4H,
ArH), 6.65 (d, 1H, J = 6.4 Hz, ArH), 5.32 (q, 1H, J = 6.4 Hz,
OCH), 4.37 (t, 2H, J = 6.4 Hz, OCH2), 3.14 (t, 2H, J = 7.1 Hz,
CH2), 2.46 (s, 3H, CH3), 2.42 (d, 2H, J = 7.1 Hz, CH2),
2.36-2.53 (m, 2H, CH2), 2.29 (s, 3H, CH3), 1.75-1.88 (m, 1H,
CH), 1.63 (d, 3H, J = 6.4 Hz, CH3), 0.86 (d, 6H, J = 6.6 Hz,
CH3); 13C NMR (50 MHz, CDCl3) 177.2, 169.9, 165.8,
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