Novel squalene-hopene cyclase inhibitors derived from hydroxycoumarins and hydroxyacetophenones

Article abstract of DOI:10.1248/cpb.52.1171

Squalene-hopene cyclase (SHC) is a useful model enzyme for predicting molecular interactions with oxidosqualene cyclase (OSC). Structure-activity relationships were investigated for numerous coumarin-derived inhibitors of SHC, and structural simplificatio

Full text of DOI:10.1248/cpb.52.1171

October 2004
Chem. Pharm. Bull. 52(10) 1171—1174 (2004)
1171
Novel Squalene-Hopene Cyclase Inhibitors Derived from
Hydroxycoumarins and Hydroxyacetophenones
Giancarlo CRAVOTTO,* Gianni BALLIANO, Silvia TAGLIAPIETRA, Simonetta OLIARO-BOSSO, and
Gian Mario NANO
Dipartimento di Scienza e Tecnologia del Farmaco, Universit a` di Torino; Via Giuria 9, 10125 Torino Italy.
Received March 8, 2004; accepted May 11, 2004
Squalene-hopene cyclase (SHC) is a useful model enzyme for predicting molecular interactions with oxi-
dosqualene cyclase (OSC). Structure–activity relationships were investigated for numerous coumarin-derived in-
hibitors of SHC, and structural simplifications are suggested. Both umbelliferone and 2,4-dihydroxyacetophe-
none provide convenient starting nuclei for the design of SHC inhibitors. Derivatives bearing an w-epoxyfarnesyl
moiety or just a plain alkyl chain showed an inhibitory effect on a recombinant SHC from Alicyclobacillus acido-
caldarius expressed in Escherichia coli.
Key words 7-alkyloxycoumarin; 4-alkyloxyacetophenone; squalene hopene cyclase; oxidosqualene cyclase; secoammoresinol;
secoferulenol
One of the most intensively studied enzymes of sterol Results and Discussion
biosynthesis, oxidosqualene cyclase (OSC), catalyzes the
We proceeded towards molecular simplification and also
conversion of an acyclic compound, 2,3-oxidosqualene, to covered a few isosters, i.e. thiocoumarin and quinolindione
the first cyclic intermediate along the pathway. This is lanos- derivatives. 2- and 4-thiocoumarin nuclei were accessed by a
2
7,28)
terol in all non-photosynthetic organisms, and cycloartenol in published procedure,
while 2,4-quinolinediol was com-
1)
plants. While several mammalian OSCs were being iso- mercially available. S-Geranylthiocoumarins 7 and 8 were
2
—9)
29)
lated, characterized, cloned and sequenced,
hundreds of prepared via a Mitsunobu reaction. 4-O-Geranyl-quinolin-
10—14)
30)
OSC inhibitors have been designed and tested
as poten- 2-one (10) was prepared according to Grundon.
15)
16)
tial anti-fungal or cholesterol-lowering drugs. Recently, a
The isosters synthesized by us (7—10) showed lower
novel series of orally active inhibitors have been tested on inhibitory activities compared to either the O-geranyl-
human liver microsomal OSC, and their effectiveness as cho- coumarins or the w-epoxy derivative 6. A shorter side chain,
3
1)
lesterol-lowering drugs has been evaluated in hyperlipidemic as in aurapten
(7-geranyloxycoumarin, 5) and 6ꢀ,7ꢀ-
17)
32)
33,34)
hamsters. OSC inhibitors are also under investigation as epoxyaurapten (6) or prenyletin
(4) was also deleterio-
18,19)
potential anti-trypanosomal agents.
The design of new OSC inhibitors received a further boost
when the complete structure of a squalene hopene cyclase results of the SHC inhibition test are summarized in Table 1.
us for the inhibitory activity.
All relevant structure modifications are shown in Fig. 2;
20)
(
SHC) was elucidated. This bacterial enzyme, isolated
from the thermophilic bacterium Alicyclobacillus acidocal-
darius, proved particularly interesting because its sequence
revealed a remarkable degree of homology and partial iden-
21,22)
tity with eukaryotic OSCs.
SHC may therefore be used
as a model to predict how newly designed inhibitors may in-
teract with OSCs. Recently, the interaction of several new in-
17)
hibitors of human OSC with the active site of SHC was in-
23)
vestigated by co-crystallization experiments. Because com-
24,25)
parative studies on the two enzyme types
have shown
close correspondence of their inhibitor activities, we recently
investigated a group of meroterpenoids that proved to be ex-
cellent inhibitors of SHC. The basic structure of bioactive
oxyprenylcoumarins is derived from umbelliprenine (7-far-
nesyloxycoumarin) by the insertion of either a tetrahydrofu-
ran unit (farnesylferol C, 1) or 1–2 oxiran rings (2, 3) (Fig.
Fig. 1. Meroterpenoid Inhibitors for SHC
26)
1
). While farnesyferol C (IC50 7.0 mM) is a naturally occur-
ring compound, umbelliprenine-10ꢀ,11ꢀ-monoepoxyde (2)
(
IC50 2.5 mM) and diepoxyde (3) (IC 1.5 mM) have been pre-
50
26)
viously obtained by synthesis.
This encouraging start prompted us to extend the study of
structure–activity relationships by modifying both the hetero-
cyclic nucleus and the side chain, hopefully to improve mole-
cular recognition.
Fig. 2. Isosters and Chemical Modifications of Coumarin and Acetophe-
none
*
To whom correspondence should be addressed. e-mail: giancarlo.cravotto@unito.it
© 2004 Pharmaceutical Society of Japan

1
172
Vol. 52, No. 10
Table 1. SHC Inhibition Values
Compound
a)
a)
Inhibition
Compound
Inhibition
b)
b)
b)
4
5
6
7
8
No activity
12
13
14
15
16
72% resid. activity
68% resid. activity
90% resid. activity
90% resid. activity
IC50 60 mM
b)
b)
b)
No activity
b)
80% resid. activity
b)
No activity
b)
No activity
b)
9
No activity
17
18
19
IC50 30 mM
IC50 48 mM
IC50 100 mM
b)
1
1
0
1
No activity
b)
No activity
a) Values are means of three experiments. b) At 100 mM.
In compounds 11, 12 and 14, the farnesyl moiety carried Both compounds proved moderately active (IC50 48 mM and
by the corresponding natural products ferulenol (3-farnesyl- 100 mM, respectively).
4-hydroxycoumarin) and umbelliprenine (7-farnesyloxycoum-
arin) is replaced with a linear alkyl chain of similar length Conclusion
C ). Compound 11 was obtained by acylation of 4-hydroxy-
Structural modification of our original coumarin-derived
(
1
1
coumarin with 10-undecenoyl chloride in pyridine, while SHC inhibitors yielded significant clues for future improve-
ester 14 was obtained by the treatment of umbelliferone with ments in molecular design. We proved that umbelliferone and
10-undecenoic acid in the presence of dicyclohexylcarbodi- hydroxyacetophenone are useful starting structures for the
imide (DCC) and dimethylaminopyridine (DMAP). A Mitsu- design of new SHC inhibitors. Although an epoxide group
nobu reaction of umbelliferone and 10-undecenol afforded inserted at the end of the prenyl chain increased the in-
compound 12. Classical epoxydation with MCPB acid hibitory effect, it did not have the same effect when similarly
afforded derivatives 13 and 15. As shown in Table 1, the placed on a linear aliphatic chain. On comparison of several
inhibitory activity fell sharply when the farnesyl moiety was farnesylcoumarins (ferprenine, ferulenol and ammoresinol)
replaced with a similar alkyl chain, revealing a critical detail with the corresponding farnesylacetophenones (secoferu-
for molecular recognition. Although an epoxide group lenol and secoammoresinol) we found the simplified aro-
inserted at the end of the farnesyl chain increased the matic moiety to be more active. The acetophenone moiety
inhibitory activity, it did not have the same effect when probably interacts more strongly with the aromatic residues
present on an aliphatic chain. Slightly increased activity was at the active site of the enzyme.
found when the side chain was bound to the heterocyclic nu-
cleus by an ester linkage, 14–15.
The present study is to be extended to eukaryotic oxi-
dosqualene cyclases.
The coumarin skeleton itself could be usefully cut down to
o-hydroxyacetophenone, the product of coumarin hydrolysis
and decarboxylation. Our inhibition tests on these derivatives
Experimental
1
General IR: Shimadzu DR 8001 spectrophotometer; NMR: H-
13
(300 MHz) and C-NMR (75 MHz) were recorded at room temperature with
started with secoferulenol 16 and secoammoresinol 17, two a Bruker 300 Advance spectrometer. The spectra were recorded in CDCl3,
3
5,36)
naturally occurring acetophenones,
prepared as previ- and solvent signals for CHCl
3
/CDCl
Reactions were monitored by TLC (Silica gel Fluka aluminium cards,
.2 mm), spots being visualized by UV inspection and/or staining with 5%
H SO in ethanol, followed by heating. Macherey-Nagel silica gel was used
3
(7.26 ppm) were used as a reference.
37)
ously described. As shown in Table 1, both were more ac-
tive than the corresponding coumarin derivatives ferulenol
0
2
4
(
IC50 90 mM) and ammoresinol (7-farnesyloxy-4-hydroxy-
for column chromatography (CC). Melting points: Büchi SMP-20 (uncor-
coumarin, no activity at 100 mM). Keeping in mind the struc- rected). Low-resolution mass spectra (LR-MS): Finnigan-MAT TSQ70 in
tures of the original inhibitors (1—3), we also elaborated on chemical ionization with isobutane as the reactant gas. HPLC analysis:
Thermo-Quest Spectra Series P200, Detector UV/VIS Jasco 875-UV or a
Gilson 133 refractive index refractometer, integrator Millipore 740 Waters.
Commercially available reagents and solvents were used without further pu-
rification unless otherwise noted. Analytical samples of all new compounds
resacetophenone (2,4-dihydroxyacetophenone), producing its
-undecenyloxy- (18) and 4-(10ꢀ,11ꢀ-epoxyundecanyloxy)-
19) derivatives. Selective O-alkylation was achieved by Mi-
4
(
tsunobu reaction; a subsequent MCPBA epoxidation gave 19. have been obtained by semi-preparative HPLC.

October 2004
1173
Preparation of Compounds 4—6 and 10 According to a standard pro-
cedure, we refluxed 4-hydroxycoumarin in anhydrous acetone with 1 eq of C, 73.29; H, 7.50.
29.1, 24.5, 21.5, 19.0. Anal. Calcd for C H O : C, 73.15; H, 7.37. Found:
2
0
24
4
38)
alkyl bromide in the presence of excess K CO to obtain in good yield
prenyletin (4, yield 89%) and aurapten (5, yield 82%).
7-(10-Undecenyloxy)chromen-2-one (12) Following the Mitsunobu
protocol, umbelliferone (1.93 g, 11,9 mmol), 10-undecenol (2.03 g,
11.9 mmol), TPP (3.22 g, 11.9 mmol), diisopropylazodicarboxylate (DIAD)
2
3
When treated in the same way, 2,4-quinolindiol gave 4-geranyloxy-2-
30)
quinolinone (10, yield 39%). 5 was epoxidated at the terminal double (2.47 ml, 11.9 mmol) and anhydrous THF (20 ml) were added to a 50 ml
bond by treatment with N-bromosuccinimide in THF/H O, then conversion round-bottomed flask. The mixture was stirred under nitrogen at rt for 2 h,
2
26,39)
of the resulting bromohydrin to epoxide (6) by treatment with a base
yield 72%).
and the reaction monitored by TLC (eluent hexane/EtOAc 8 : 2. Rf
12ꢃ0.45). After evaporation under vacuum the raw material was purified by
(
Preparation of S-Geranyl Thiocoumarins 7 and 8 A solution in anhy- CC (eluent gradient: hexane/EtOAc 9 : 1Æ8 : 2), yielding 2.98 g of 12
drous THF containing triphenylphosphine (1.5 eq), geraniol (1.5 eq) and (9.5 mmol, yield 80%). White powder, mp 85 °C. IR (KBr) 1726, 1624,
thiocoumarin (1.0 eq) was magnetically stirred at 0 °C under N . Diisopropyl 1134, 837 cm . LR-MS: 315 (MH , CI-MS). H-NMR (CDCl ) d: 7.60
ꢁ1
ꢂ
1
2
3
azodicarboxylate (DIAD) (1.5 eq) was added dropwise over a period of (1H, d, Jꢃ9.5 Hz, H-4), 7.35 (1H, d, Jꢃ8.4 Hz, H-5), 6.80 (2H, m, H-6, H-
min. The orange-red color of DIAD immediately disappeared. The weakly 8), 6.25 (1H, d, Jꢃ9.5 Hz, H-3), 5.75 (1H, m, H-10ꢀ), 4.95 (2H, m, H-11ꢀ),
exothermic reaction was allowed to proceed at room temperature under stir- 3.95 (1H, t, Jꢃ6.5 Hz, H-1ꢀ), 2.00 (2H, m, H-9ꢀ), 1.80 (2H, m, H-2ꢀ),
5
1
3
ring. Its duration ranged from 1 to 24 h, depending on the nature of the enol.
1.50—1.30 (12H, m, H-3ꢀ—8ꢀ). C-NMR (CDCl ) d: 163.3, 161.2, 155.8,
3
After completion, as indicated by TLC monitoring (eluent hexane–ethyl ac- 143.3, 139.1, 127.7, 112.9, 112.8, 112.2, 101.2, 33.7, 33.3, 29.7, 29.6, 29.5,
etate), the reaction mixture was evaporated. The residue was diluted with
hexane–ether 3 : 1, filtered through a thin pad of Celite to remove the precip-
itate of triphenylphosphine oxide, and concentrated under reduced pressure.
Finally, the products were purified by flash silica gel column chromatogra-
phy. Isolated yields 7 (17%), 8 (29%).
29.3, 29.2, 29.0, 23.9, 22.6. Anal. Calcd for C H O : C, 76.40; H, 8.33.
20 26 3
Found: C, 76.40; H, 8.46.
7-(9-Oxiranylnonyloxy)chromen-2-one (13) In a 100 ml round-bot-
tomed flask, 12 (1.30 g, 4.1 mmol) was dissolved in anhydrous CH Cl2
2
(10 ml). The solution was stirred, then cooled to 0 °C. Anhydrous sodium ac-
2
-Geranylsulfanylchromen-4-one (7): Yellow oil. IR (liquid film) 1643, etate (0.712 g, 4.1 mmol) was added all at once, and 2.76 g of MCPBA 70%
ꢁ1 ꢂ
1
1
587, 1356, 1130, 922, 759 cm . LR-MS: 315 (MH , CI-MS). H-NMR (12.3 mmol) was added portionwise under stirring. After 4 h of stirring, the
(
CDCl ) d: 8.12 (1H, d, Jꢃ7.5 Hz, H-5), 7.59 (1H, m, H-7), 7.37—7.24 (2H, reaction was monitored by TLC (eluent hexane/EtOAc 8 : 2. Rf 13ꢃ0.21).
3
m, H-6, H-8), (1H, t, Jꢃ7.3 Hz,), 6.23 (1H, br s, H-3), 5.28 (1H, br t,
Work-up: the reacted mixture was washed with 5% Na S O solution and
2 2 3
Jꢃ6.4 Hz, H-2ꢀ), 5.10 (1H, m, H-6ꢀ), 3.67 (2H, d, Jꢃ7.6 Hz, H-1ꢀ), 1.77 5% NaOH and brine. Silica gel CC eluent gradient: hexane/EtOAc
1
3
(
3H, s, H-10ꢀ*), 1.68 (3H, s, H-9ꢀ*), 1.61 (3H, s, H-8ꢀ). C-NMR (CDCl3) 9 : 1Æ7 : 3 obtaining 0.954 g of 13 (2.88 mmol, yield 70%). White powder,
ꢁ1 ꢂ
d: 159.1, 156.7, 151.9, 143.2, 131.9, 131.4, 128.4, 124.0, 123.3, 115.4, mp 85 °C. IR (KBr) 1726, 1624, 1134, 915, 837 cm . LR-MS: 331 (MH ,
06.9, 38.9, 29.2, 26.1, 25.5, 17.6, 16.3. Anal. Calcd for C H O S: C,
1
1
7
CI-MS). H-NMR (CDCl ) d: 7.60 (1H, d, Jꢃ9.5 Hz, H-4), 7.35 (1H, d,
1
9
22
2
3
2.57; H, 7.05. Found: C, 73.00; H, 7.16.
Jꢃ8.4 Hz, H-5), 6.80 (2H, m, H-6, H-8), 6.25 (1H, d, Jꢃ9.5 Hz, H-3), 3.95
4
-Geranylsulfanylchromen-2-one (8): Yellow oil. IR (liquid film) 1721, (2H, t, Jꢃ6.5 Hz, H-1ꢀ), 2.75 (2H, t, Jꢃ4.9 Hz, H-11ꢀ), 2.45 (1H, m, H-10ꢀ),
ꢁ1 ꢂ
1
1
(
595, 1192, 929, 756, 745 cm . LR-MS: 315 (MH , CI-MS). H-NMR
CDCl ) d: 7.83 (1H, d, Jꢃ7.9 Hz, H-5), 7.55 (1H, m, H-7), 7.32 (1H, d,
Jꢃ8.1 Hz, H-8), 7.24 (1H, t, Jꢃ7.3 Hz, H-6), 6.15 (1H, s, H-3), 5.34 (1H, t,
1.80 (2H, m, H-2ꢀ), 1.65 (2H, m, H-9ꢀ), 1.50—1.20 (12H, m, H-3ꢀ—8ꢀ).
1
3
C-NMR (CDCl ) d: 159.1, 156.7, 151.9, 143.2, 131.9, 131.4, 128.4, 124.0,
3
3
123.3, 115.4, 106.9, 38.9, 29.2, 26.1, 25.5, 17.6, 16.3. Anal. Calcd for
Jꢃ7.4 Hz, H-2ꢀ), 5.07 (1H, m, H-6ꢀ), 3.67 (2H, d, Jꢃ8.4 Hz, H-1ꢀ), 1.77 C H O : C, 72.70; H, 7.93. Found: C, 72.59; H, 7.86.
2
0
26
4
1
3
(
3H, s, H-10ꢀ*), 1.68 (3H, s, H-9ꢀ*), 1.61 (3H, s, H-8ꢀ). C-NMR (CDCl3)
10-Undecenoic Acid 2-Oxo-2H-chromen-7-yl Ester (14) In a 100 ml
d: 159.1, 156.7, 151.9, 143.2, 131.9, 131.4, 128.4, 124.0, 123.3, 115.4, round-bottomed flask, umbelliferone (4.0 g, 24.7 mmol), 10-undecenoic acid
1
06.9, 38.9, 29.2, 26.1, 25.5, 17.6, 16.3. Anal. Calcd for C H O S: C,
(5.0 ml, 24.7 mmol), DCC (5.1 g, 24.7 mmol) and DMAP (0.3 g, 2.4 mmol)
were dissolved in anhydrous toluene (40 ml). After 2 h magnetic stirring at
1
9
22
2
7
2.57; H, 7.05. Found: C, 72.80; H, 7.21.
-Geranyl-2-mercaptochromen-4-one (9) In
round-bottomed flask equipped with a magnetic stirrer and a nitrogen inlet,
3
a 50 ml two-necked rt, the reaction was monitored by TLC (eluent hexane/EtOAc 7 : 3 Rf
14ꢃ0.45). CC (eluent gradient: hexane/EtOAc 95 : 5Æ9 : 1Æ7 : 3) yielded
5.51 g of ester 14 (16.8 mmol, yield 68%). White powder, mp 57 °C. IR
37)
excess t-BuOK (0.600 g) anhydrous toluene (5 ml), deprenylsecoferulenol
ꢁ1
ꢂ
(1-(2-hydroxyphenyl)-5,9-dimethyldeca-4,8-dien-1-one) (0.500 g, 1.8 mmol) (KBr) 1732, 1626, 1468, 1269, 1165, 991, 927 cm . LR-MS: 329 (MH ,
1
and CS (130 ml) were added. Progressively, the yellow solution turned to CI-MS). H-NMR (CDCl ) d: 7.70 (1H, d, Jꢃ9.5 Hz, H-4), 7.48 (1H, d,
red. After 14 h the reaction was complete, as shown by TLC (eluent
2
3
Jꢃ8.4 Hz, H-5), 7.05 (2H, m, H-6, H-8), 6.40 (1H, d, Jꢃ9.5 Hz, H-3), 5.85
hexane/EtOAc 4 : 1. Rf 9ꢃ0.35). Work-up: the reacted mixture was diluted (1H, m, H-10ꢀ), 4.95 (2H, d, Jꢃ13.3 Hz, H-11ꢀ), 2.60 (1H, t, Jꢃ7.4 Hz, H-
with EtOAc, washed with sat. NH Cl and brine, dried over Na SO , filtered, 2ꢀ), 2.05 (2H, m, H-9ꢀ), 1.75 (2H, m, H-3ꢀ), 1.50—1.30 (10H, m, H-4ꢀ—8ꢀ).
4
2
4
1
3
and evaporated to dryness. CC (eluent hexane/EtOAc 9 : 1) yielded 115 mg
C-NMR (CDCl ) d: 171.5, 160.3, 154.5, 153.2, 142.8, 139.0, 128.9, 118.3,
3
of 9 (0.36 mmol, 20%). Yellow oil. IR (liquid film) 1742, 1608, 1550, 1460,
116.4, 115.9, 114.1, 110.3, 34.2, 33.6, 29.6, 29.1, 29.0, 28.9, 28.7, 24.6.
ꢁ1
ꢂ
1
1
8
375, 1120, 1025 cm . LR-MS: 315 (MH , CI-MS). H-NMR (CDCl ) d: Anal. Calcd for C H O : C, 73.15; H, 7.37. Found: C, 73.22; H, 7.31.
3
20 24
4
.21 (1H, d, Jꢃ7.9 Hz, H-5), 7.63 (1H, m, H-7), 7.40 (2H, m, H-6), 7.31
9-Oxyranyl-nonanoic Acid 2-Oxo-2H-chromen-7-yl Ester (15) In a
(1H, d, Jꢃ7.9 Hz, H-8), 5.17 (1H, t, Jꢃ7.3 Hz, H-2ꢀ), 5.06 (1H, t, Jꢃ7.1 Hz,
100 ml round-bottomed flask, 7-(10-undecenoyloxy)coumarin (700 mg,
H-6ꢀ), 3.48 (1H, d, Jꢃ7.1 Hz, H-1ꢀ), 2.07 (2H, m, H-5ꢀ), 2.00 (2H, m, H-4ꢀ), 2.13 mmol) was dissolved in anhydrous CH Cl (10 ml). To the ice-cold
2
2
1
3
1
.82 (3H, s, CH ), 1.66 (3H, s, CH ), 1.58 (3H, s, CH ). C-NMR (CDCl3) magnetically stirred solution, anhydrous sodium acetate (700 mg,
3
3
3
d: 184.2, 157.2, 151.9, 143.2, 131.9, 131.4, 128.4, 124.0, 123.3, 115.4, 8.52 mmol) was added all at once, and MCPBA 70% (1.435 g, 6.39 mmol)
1
06.9, 38.9, 29.2, 26.1, 25.5, 17.6, 16.3. Anal. Calcd for C H O S: C,
was added portionwise. After 4 h of stirring, the reaction was monitored by
1
9
22
2
7
2.57; H, 7.05. Found: C, 72.40; H, 9.92.
-(10-Undecenoyl)chroman-2,4-dione (11) In
round-bottomed flask equipped with a magnetic stirrer and a nitrogen inlet,
-hydroxycoumarin (5.0 g, 30.8 mmol) and a catalytic amount of piperidine 60%). White powder, mp 75 °C. IR (KBr) 1763, 1618, 1228, 1120, 916,
TLC (eluent hexane/EtOAc 7 : 3. Rf 15ꢃ0.27). Work-up: The reacted mix-
3
a 50 ml two-necked ture was washed with 5% Na S O , 5% NaOH and brine. CC (eluent gradi-
2 2 3
ent: hexane/EtOAc 9 : 1Æ6 : 4) yielded 436 mg of 15 (1.27 mmol, yield
4
ꢁ1
ꢂ
1
were dissolved in anhydrous pyridine (35 ml). The solution was placed under 756 cm . LR-MS: 345 (MH , CI-MS). H-NMR (CDCl ) d: 7.70 (1H, d,
3
stirring, cooled down to 0 °C, and 10-undecenoyl chloride was added drop-
wise. The mixture was stirred for 8 h under nitrogen at 40 °C, and the reac-
tion monitored by TLC (eluent hexane/EtOAc 4 : 1. Rf 11ꢃ0.4). Work-up:
the reacted mixture was diluted with EtOAc and washed with 5% HCl and
with NaHCO , dried over Na SO , filtered and evaporated to dryness. CC
Jꢃ9.5 Hz, H-4), 7.48 (1H, d, Jꢃ8.4 Hz, H-5), 7.05 (2H, m, H-6, H-8), 6.40
(1H, d, Jꢃ9.5 Hz, H-3), 2.70 (1H, m, H-10ꢀ), 2.60 (2H, t, Jꢃ7.5 Hz, H-11ꢀ),
2.30 (2H, t, Jꢃ7.4 Hz, H-2ꢀ), 1.75 (2H, m, H-3ꢀ), 1.50 (2H, m, H-9ꢀ),
1
3
1.40—1.20 (10H, m, H-4ꢀ—10ꢀ). C-NMR (CDCl ) d: 170.4, 160.9, 155.1,
3
153.6, 139.5, 129.1, 118.3, 116.6, 116.0, 110.8, 52.3, 47.1, 34.2, 33.8, 29.6,
3
2
4
(
eluent hexane/EtOAc 19 : 1) yielded 7.0 g of 11 (21.3 mmol, 70%). White
29.4, 29.2, 28.9, 28.7, 24.7. Anal. Calcd for C H O : C, 69.75; H, 7.02.
2
0
24
5
ꢁ1
powder, mp 98 °C. IR (KBr) 1717, 1607, 1547, 1225, 980, 901 cm . LR- Found: C, 69.63; H, 7.20.
ꢂ
1
MS: 329 (MH , CI-MS). H-NMR (CDCl ) d: 8.08 (1H, d, Jꢃ7.90 Hz, H-
5
8
Jꢃ7.45 Hz, H-2ꢀa,b). C-NMR (CDCl ) d: 209.3, 207.5, 179.2, 160.3,
1
1-(2-Hydroxy-4-undec-10-enyloxy-phenyl)ethanone (18) In a 50 ml
two-necked round-bottomed flask equipped with a magnetic stirrer and a
3
), 7.73 (1H, td, Jꢃ7.14 Hz, Jꢀꢃ1.39 Hz, H-7), 7.38—7.28 (2H, m, H-6, H-
), 5.90—5.76 (1H, m, H-10ꢀ), 5.03—4.93 (2H, m, H-11ꢀa,b), 3.21 (2H, t, nitrogen inlet, resacetophenone (1.0 g, 6.57 mmol), 10-undecenol (1.31 ml,
13
6.57 mmol), TPP (2.07 g, 6.57 mmol), diisopropylazodicarboxylate (DIAD)
(1.56 ml, 6.57 mmol) and anhydrous THF (20 ml) were added. The mixture
3
36.5, 126.0, 124.8, 117.4, 115.8, 101.5, 85.2, 68.5, 42.1, 31.4, 29.6, 29.4,

1
174
Vol. 52, No. 10
was stirred for 1 h under nitrogen at rt, and the reaction monitored by TLC
eluent hexane/EtOAc 19 : 1. Rf 18ꢃ0.42). After evaporation under vacuum,
the raw material was purified by CC (eluent hexane), yielding 1.28 g of 18
Singh V., Sarshar S., J. Am. Chem. Soc., 117, 11819—11820 (1995).
12) Ceruti M., Balliano G., Viola F., Grosa G., Rocco F., Cattel L., J. Med.
Chem., 35, 3050—3058 (1992).
(
(
4.2 mmol, yield 64%). Yellow powder, mp 41 °C. IR (KBr) 3350, 1634, 13) Ceruti M., Rocco F., Viola F., Balliano G., Milla P., Arpicco S., Cattel
ꢁ1 ꢂ
1
1
1
6
372, 1256, 1069 cm . LR-MS: 305 (MH , CI-MS). H-NMR (CDCl ) d:
L., J. Med. Chem., 41, 540—554 (1998).
14) Abe I., Liu W., Oelhschlager A. C., Prestwich G. D., J. Am. Chem.
Soc., 118, 9180—9181 (1996).
3
2.7 (1H, s, OH), 7.54 (1H, d, Jꢃ8.7 Hz, H-6), 6.37 (1H, d, overlap., H-5),
.33 (1H, s, H-3), 5.79 (1H, m, H-10ꢀ), 4.92 (2H, m, H-11ꢀa, 11ꢀb), 3.91
(
2H, t, Jꢃ6.4 Hz, H-1ꢀ), 2.48 (3H, s, CH ), 2.00 (2H, m, H-9ꢀ), 1.73 (2H, m, 15) Jolidon S., Polak-Wyss A., Hartman P. G., Guerry P. G., “Recent Ad-
3
1
3
H-2ꢀ), 1.28 (12H, m, H-3ꢀ—8ꢀ). C-NMR (CDCl ) d: 202.1, 165.4, 165.0,
vances in the Chemistry of Anti-infective Agents,” Royal Soc. Chem.,
Cambridge, U.K., 1993, pp. 223—233.
3
1
2
7
38.8, 131.9, 113.9, 113.4, 107.6, 101.0, 68.1, 33.5, 29.5, 29.2, 29.1, 28.8,
8.7, 28.6, 25.8. Anal. Calcd for C H O : C, 74.96; H, 9.27. Found: C, 16) Morand O. H., Aebi J. D., Dehlmlow H., Ji Y. H., Gains N., Lengsfeld
1
9
28
3
5.00; H, 9.27.
H., Himber J. J., Lipid Res., 38, 373—390 (1997).
1-[2-Hydroxy-4-(9-oxyranyl-nonyloxy)phenyl]ethanone (19) As de- 17) Dehlmlow H., Aebi J. D., Jolidon S., Ji Y. H., von der Mark E. M.,
scribed for the preparation of 15, employing 18 (231 mg, 0.763 mmol), anhy-
drous sodium acetate (311 mg, 2.29 mmol), MCPBA 70% (512 mg,
Himber J., Morand O. H., J. Med. Chem., 46, 3354—3370 (2003).
18) Buckner F. S., Griffin J. H., Wilson A. J., Van Voorhis W. C., Antimi-
crob. Agents Chemother., 45, 1210—1215 (2001).
2
.29 mmol) and anhydrous CH Cl (10 ml) we recovered 176 mg of 19 (yield
2 2
7
2%) after CC purification (eluent hexane/EtOAc 19 : 1). TLC (eluent
19) Oliaro-Bosso S., Ceruti M., Balliano G., Matsuda S. P. T., Milla P.,
Rocco F., Viola F., 93rd AOCS Annual Meeting & Expo, Montreal,
Quebec, (Canada), on May 5—8, 2002.
hexane/EtOAc 8 : 2): Rf 19ꢃ0.47. An analytical sample was obtained by
HPLC (eluent hexane/EtOAc 9 : 1). Yellow oil. IR (liquid film) 1638, 1508,
1
ꢁ1
ꢂ
1
427, 1370, 1257, 1134, 802 cm . LR-MS: 321 (MH , CI-MS). H-NMR 20) Wendt K. U., Poralla K., Schulz G. E., Science, 277, 1811—1815
(
CDCl ) d: 12.7 (1H, s, OH), 7.62 (1H, dd, Jꢃ11.3, Jꢀꢃ2.3 Hz, H-6), 6.43
(1997).
3
(1H, dd overlap., Jꢃ6.5 Hz, H-5), 6.40 (1H, s, H-3), 3.99 (2H, t, Jꢃ6.5 Hz,
21) Wendt K. U., Lenhart A., Schulz G. E., J. Mol. Biol., 286, 175—197
(1999).
H-1ꢀ), 2.91 (1H, br s, H-10ꢀ), 2.75 (1H, t, Jꢀꢃ4.2 Hz, H-11ꢀa), 2.55 (3H, s,
CH ), 2.47 (1H, t, Jꢀꢃ4.2 Hz, H-11ꢀb), 1.79 (2H, m, H-2ꢀ), 1.53 (2H, m, H-
22) Wendt K. U., Schulz G. E., Corey E. J., Liu D. R., Angew. Chem. Int.
Ed., 39, 2821—2833 (2000).
3
1
3
9
ꢀ), 1.51—1.26 (12H, m, H-3ꢀ—8ꢀ). C-NMR (CDCl ) d: 202.4, 183.7,
3
1
65.6, 165.1, 132.1, 113.6, 107.9, 101.1, 68.3, 52.3, 47.1, 38.6, 32.4, 29.3, 23) Lenhart A., Reinert D. J., Aebi J. D., Dehmlow H., Morand O. H.,
2
9.2, 29.1, 29.0, 28.8, 26.1. Anal. Calcd for C H O : C, 71.22; H, 8.81.
Schulz G. E., J. Med. Chem., 46, 2063—2092 (2003).
24) Viola F., Ceruti M., Cattel L., Milla P., Poralla, K., Balliano G., Lipids,
35, 297—303 (2000).
1
9
28
4
Found: C, 71.08; H, 8.90.
General Procedure for SHC Inhibition Tests IC50 values (inhibitor
concentrations that reduced by 50% the rate of enzymatic conversion of 25) Ceruti M., Balliano G., Rocco F., Milla P., Arpicco S., Cattel L., Viola
squalene to hopene) were determined at 10 mM substrate concentration. F., Lipids, 36, 629—636 (2001).
C]squalene was obtained by incubating 1 mCi of [ C]mevalonolactone 26) Cravotto G., Balliano G., Robaldo B., Oliaro-Bosso S., Chimichi S.,
with the S10 supernatant of a pig liver homogenate (25 mg of protein), fol- Boccalini M., Bioorg. Med. Chem. Lett., 14, 1931—1934 (2004).
lowing the method of Popják, in the presence of the oxidosqualene cyclase 27) Di Braccio M., Roma G., Leoncini G., Poggi M., Farmaco, 50, 703—
inhibitor U-18666A. Under these conditions, the bulk of radioactivity of 711 (1995).
nonsaponifiable extract was shared between squalene and oxidosqualene, 28) Majumdar K. C., Biswas A., Mukhopadhyay P. P., Synthesis, 15,
14
14
[
40)
41)
which can be easily separated by TLC.
2385—2389 (2003).
29) Cravotto G., Tagliapietra S., Palmisano G., Nano G. M., Heterocycles,
Acknowledgements The financial support of this work from Medestea
60, 1351—1358 (2003).
Research and Production Srl (Italy) is gratefully acknowledged.
30) Grundon M. F., Ramachandran V. N., Donnelly M. E., Perkin Trans 1,
, 633—635 (1981).
2
References
31) Angioni A., Cabras P., D’hallewin G., Pirisi F., M. Reniero F., Schirra
1
2
)
)
Nes W. D., Recent Adv. Phytochem., 24, 283—327 (1990).
Abe I., Sankawa U., Ebizuka Y., Chem. Pharm. Bull., 40, 1755—1760 32) Aziz M., Rouessac F., Tetrahedron Lett., 28, 2579—2582 (1987).
M., Phytochemistry, 47, 1521—1525 (1998).
(
1992).
Moore W. R., Schatzman G. L. J., Biol. Chem., 267, 22003—22006
1992).
Corey E. J., Matzuda S. P. T., J. Am. Chem. Soc., 113, 8172—8174
1991).
Kelly R., Miller S. M., Lai M. H., Kirsch D. R., Gene, 87, 177—183
1990).
33) Pistelli L., Bertoli A., Bilia A. R., Morelli I., Phytochemistry, 41,
1579—1582 (1996).
34) Ganguly N. C., Sukai A. K., Dutta S., De P., J. Ind. Chem. Soc., 78,
380—382 (2001).
35) Al-Yahya M. A., Muhammad I., Mirza H. H., El-Feraly F. S., Phy-
tother. Res., 12, 335—339 (1998).
36) Mizukami H., Ogihara Y., Chem. Pharm. Bull., 46, 1781—1784
(1998).
3)
4)
5)
6)
7)
8)
9)
(
(
(
Roessner C. A., Min C., Hardin S. H., Harris-Haller L. W., McCollum
J. C., Scott A. I., Gene, 127, 149—150 (1993).
Lees N. D., Skaggs B., Kirsch D. R., Bard M., Lipids, 30, 221—226
37) Appendino G., Tagliapietra S., Cravotto G., Nano G. M., Gazz. Chim.
Ital., 119, 385—388 (1989).
(
1995).
38) Gonzales A. G., Jorge D., Anales de Quimica, 79, 265—269 (1983).
Baker C. H., Matsuda S. P. T., Liu D. R., Corey E. J., Biochem. Bio- 39) Van Tamelen E. E., Nadeau R. G., J. Am. Chem. Soc., 89, 176—177
phys. Res. Commun., 213, 154—160 (1995). (1967).
Abe I., Prestwich G. D., Proc. Natl. Acad. Sci. U.S.A., 92, 9274—9278 40) Popják, G., “Methods in Enzymology, Steroids and Terpenoids,” Vol.
(
1995).
0) Abe I., Rohmer M., Prestwich G. D., Chem. Rev., 93, 2189—2206
1993).
1) Corey E. J., Virgil S. C., Cheng H., Hunter-Baker C., Matsuda S. P. T.,
XV, ed. by Clayton R. B., Academic Press, New York, 1969, pp.
438—443.
41) Balliano G., Viola F., Ceruti M., Cattel L., Biochim. Biophys. Acta,
959, 9—19 (1988).
1
1
(