New stereocontrolled synthesis and biological evaluation of 5-(1′-hydroxyalkyl)-3-methylidenetetrahydro-2-furanones as potential cytotoxic agents

Article abstract of DOI:10.1021/jm011019v

A series of 3-methylidenetetrahydro-2-furanones 7 bearing various hydroxyalkyl substituents in position 5 were synthesized using novel diastereo- and enantioselective methodology. In vitro cytotoxicity data demonstrated that all prepared compounds were ac

Full text of DOI:10.1021/jm011019v

1142
J . Med. Chem. 2002, 45, 1142-1145
Brief Articles
New Ster eocon tr olled Syn th esis a n d Biologica l Eva lu a tion of
5-(1′-Hyd r oxya lk yl)-3-m eth ylid en etetr a h yd r o-2-fu r a n on es a s P oten tia l Cytotoxic
Agen ts
Tomasz J anecki,*,† Edyta Błaszczyk,† Kazimierz Studzian,‡ Marek Ro´z˘ alski,§ Urszula Krajewska,§ and
Anna J anecka‡
Institute of Organic Chemistry, Technical University, Z˙ eromskiego 116, 90-924 Ło´dz´, Poland, Department of Medicinal
Chemistry, Medical University, Lindleya 6, 90-131 Ło´dz´, Poland, and Institute of Biochemistry, Medical University,
Muszyn´skiego 1, Ło´dz´, Poland
Received August 13, 2001
A series of 3-methylidenetetrahydro-2-furanones 7 bearing various hydroxyalkyl substituents
in position 5 were synthesized using novel diastereo- and enantioselective methodology. In
vitro cytotoxicity data demonstrated that all prepared compounds were active against L-1210
and HL-60 tissue culture cells with 7e being the most potent (IC50 ) 6.9 µM). Also an increase
in activity with an increase in lipophilicity of the substituents in the order H < alkyl < phenyl
was observed.
In tr od u ction
groups enhance activity4 and might be involved in direct
binding to receptor sites in tumor cells.5 Neighboring
hydroxyl groups might also play an important role in
the rate and/or selectivity of the tiols addition to
furanones 1. Although no evidence was found so far to
prove this assumption,6 the influence of adjacent hy-
droxyl groups on selectivity of the Michael additions to
enones is well known7 and was also documented for tiols
as nucleophiles.8
3-Methylidenetetrahydro-2-furanones 1 (R-methylene-
γ-butyrolactones) constitute an important group of
natural products and possess wide-ranging biological
activities.1,2 The R-methylene-γ-butyrolactone ring is an
internal building block of the sesquiterpene lactones
such as vernolepin, aromaticin, or elephantopin, isolated
from plant extracts. All these compounds have cytotoxic,
antitumoral, and often bactericidal properties. Although
none of them has demonstrated enough selectivity to
be considered for clinical use, these compounds are of
interest as leads to the synthesis of analogues, which
may achieve necessary selectivity.
Despite these encouraging observations, the number
of general synthetic methods leading to 3-methylidene-
tetrahydro-2-furanones with hydroxyl functionality is
very limited. Main synthetic efforts were focused on
3-alkylidene-4-hydroxytetrahydro-2-furanones 2 which
are a class of natural products isolated from plants of
the Lauraceae family.9 Much less attention was given
to the synthesis of 5-hydroxyalkyl-3-methylidenetet-
rahydro-2-furanones 3. So far, their preparation was
realized by palladium-catalyzed carbonylation of vinyl
halides,10 the methylenation of 5-hydroxymethyltet-
rahydro-2-furanone,11 bromination of isosaccharinic
acid,12 Reformatsky reaction of R-(bromomethyl)acrylate
with trialkylsilyloxyacetone,13 or the lactonization of
R-methylidene-γ-hydroxy-δ-alkoxyesters.14 All these
methods have been shown to be of very limited ap-
plicability and often give racemic products. Also, cyto-
toxicity of the compounds prepared was not assayed.
The cytotoxic activity of 3-methylidenetetrahydro-2-
furanones 1 is mainly associated with the exocyclic,
conjugated double bond which acts as an alkylating
agent in a Michael-type reaction with thiol rich bionu-
cleophiles. However, it has also been postulated that
there are other factors which may enhance the biological
properties of these compounds, e.g., the presence of
conjugate side chain ester or stereochemically defined
carbinol units at strategic positions.3 The fact that many
natural furanones 1 showing in vivo antitumor activity
contain hydroxyl groups led to the conclusion that these
Continuing our investigations focused on the applica-
tion of organophosphorus reagents in the synthesis of
3-methylidenetetrahydro-2-furanones,15,16 we have re-
cently presented,17 as a preliminary communication, our
newly developed diastereo- and enantioselective tech-
niques for the preparation of 5-(1′-hydroxyalkyl)-3-
methylidenetetrahydro-2-furanones 7. In this paper we
report full experimental details of our methodology as
well as cytotoxic evaluation of 5-(1′-hydroxyalkyl)-3-
* To whom correspondence should be addressed. Phone: +(42)
6313142. Fax: +(42) 6365530. E-mail: tjanecki@ck-sg.p.lodz.pl.
† Technical University.
‡ Department of Medicinal Chemistry, Medical University.
§ Institute of Biochemistry, Medical University.
10.1021/jm011019v CCC: $22.00 © 2002 American Chemical Society
Published on Web 02/01/2002

Brief Articles
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 5 1143
Sch em e 1a
a
Reagents: (a) OsO4 cat., NMO, H2O/acetone, rt, 24 h; or AD-mix-â, 50% aqueous t-BuOH, 0 °C to rt, 48 h; (b) 36% formalin, K2CO3,
0-5 °C, 15 min; (c) MCPBA, CH2Cl2, rt, 24 h; (d) 30% HClO4, rt, 24 h.
Ta ble 1. Lipophilicity and Cytotoxic Activity of
methylidenetetrahydro-2-furanones 7, which were as-
3-Methylidenetetrahydro-2-furanones 7a -e
sayed in vitro against human leukemia HL-60 and
IC50 (µM)
mouse leukemia L-1210 cells. Some structure-activity
lipophilicity
compound
7a
(log P)
L-1210
HL-60
relationships of the new compounds are also discussed.
0.15
0.44
0.55
27.2 ( 6.1
26.7 ( 4.1
19.3 ( 3.7
17.3 ( 2.1
18.5 ( 4.4
16.9 ( 3.9
8.0 ( 2.1
7.5 ( 2.5
6.9 ( 1.9
9.7 ( 1.2
72.4 ( 16.9
51.3 ( 3.2
47.3 ( 3.7
46.8 ( 2.9
51.9 ( 5.1
52.6 ( 4.9
40.2 ( 2.4
41.2 ( 2.8
36.4 ( 3.5
2.9 ( 01
Ch em istr y
7b
(5R*,1′R*)-7c
(5R*,1′S*)-7c
(5R*,1′R*)-7d
(5R*,1′S*)-7d
(5R*,1′R*)-7e
(5R*,1′S*)-7e
(5R,1′R)-7e
carboplatin
Synthesis of the target diastereoisomeric and enan-
tiomeric 5-hydroxyalkyl-2-furanones 7 was accom-
plished applying described methodology17 and is sum-
marized in the Scheme 1. Starting (E)-2-diethoxyphos-
phoryl-4-alkenoates (E)-8a -e were prepared by alky-
lation of ethyl diethoxyphosphoryl acetate with ap-
propriate allyl bromides. Chemoselective hydrolysis of
(E)-8a -e produced other key substrates, 2-diethoxy-
phosphoryl-4-alkenoic acids (E)-4a -e.
Acids (E)-4a -e and alkenoates (E)-8c-e were next
transformed into diols (5R*,1′R*)-5a -e and (5R*,1′S*)-
5c-e, respectively, using fully diastereoselective syn-
or anti-dihydroxylation procedures (OsO4/N-methylmor-
pholine oxide or oxidation of (E)-8c-e with MCPBA and
cleavage of the epoxides (4R*,5R*)-9c-e with perchloric
acid). Spontaneous lactonization of these diols gave
3-diethoxyphosphoryltetrahydro-2-furanones (5R*,1′R*)-
6a -e and (5R*,1′S*)-6c-e. Due to the additional ste-
reogenic center at C-3, furanones 6 were obtained as
mixtures of diastereoisomers with close to 1:1 ratio.
Horner-Wadsworth-Emmons olefination of formalde-
hyde using (5R*,1′R*)-6a-e and (5R*,1′S*)-6c-e yielded
target 3-methylidenetetrahydro-2-furanones (5R*,1′R*)-
7a -e and (5R*,1′S*)-7c-e.
1.59
1.80
alkenoate 8c and alkenoic acid 4c were prepared as a
mixtures of diastereoisomers (E/ Z ) 80/20) and also
furanones 6c were mixtures of four diastereoisomers.
Enantioselective synthesis of (5R,1′R)-7e (ee 90%) was
accomplished using commercially available Sharpless
reagent, AD-mix-â18 in syn-dihydroxylation of the alk-
enoic acid (E)-4e. The enantiomeric excess and absolute
configuration was determined by transformation of
(5R,1′R)-7e into corresponding Mosher’s ester,19 followed
1
by the analysis of the resulting diastereoisomers by H
NMR.
Resu lts a n d Discu ssion
The obtained compounds were evaluated in vitro for
their cytotoxic activity against two cell lines: mouse
L-1210 leukemia and human HL-60 leukemia. The
activity is expressed as the concentration (µM) required
to inhibit tumor cell proliferation by 50% after 72 h
exposure of the cells to the compounds (IC50). The
results are presented in Table 1. The data for carbo-
All furanones 7 except 7c were obtained as single
diastereoisomers. Compounds 7c, obtained by employ-
ing syn- or anti-dihydroxylation procedures, were mix-
tures of diastereoisomers (5R*,1′R*)/(5R*,1′S*) ) 80/20
or (5R*,1′S*)/(5R*,1′R*) ) 20/80, respectively. This is a
consequence of using commercially available E/ Z mix-
ture of crotyl bromide (E/ Z ∼ 85/15) in the alkylation
of ethyl diethoxyphosphoryl acetate. For that reason
platin20
(cis-diamine[1,1-cyclobutanedicarboxylato]-
platinum) are included for comparison. All tested com-
pounds displayed anticancer activity and were 2-4
times more potent against the growth of L-1210 than

1144 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 5
Brief Articles
on Specord M80 spectrometer. 1H NMR (250 MHz), 13NMR
(62.9 MHz), and 31P NMR (101 MHz) spectra were recorded
on Brucker DPX250 spectrometer with TMS as an internal
standard and 85% H3PO4 as an external standard, respec-
tively. 31P NMR spectra were recorded using broadband proton
decoupling.
HL-60 cells. Matching relationships between the sub-
stituent on the carbinol carbon atom C-1′ and activity
were observed for both cell lines, with furanone 7e (R1
) Ph) being the most potent and furanone 7a (R1 ) H)
the least potent compound. Furanones 7c,d with alkyl
substituents (R1 ) Me, Pr) displayed intermediate
activity. Both diastereoisomers of compounds 7c-e were
tested, but no significant differences in activity were
found. Also, the optically active (5R,1′R) 7e (90% ee) and
racemic (5R*,1′R*) 7e were almost equipotent. The most
potent compound in the series, furanone 7e, had the
activity comparable to carboplatin against L-1210 cells.
In the search for more clear structure-activity rela-
tionships in furanones 7a -e, we decided to determine
their lipophilicity. Correlation between lipophilic char-
acter of the compound, expressed as the partition
coefficient P in 1-octanol/water system, and its biological
activity have proven meaningful.21 Moreover, structure-
cytotoxicity investigations among sesquiterpene lactones
revealed that an increase in cytotoxicity usually ac-
companies increased lipophilicity.4 Lipophilicities of
furanones 7a -e were calculated using the standard
software package Pallas (CompuDrug Chemistry, Hun-
gary) and the results, expressed as log P, are given in
Table 1. To test the accuracy of the calculations, the
lipophilicity of 7a was also determined experimentally
using a spectrophotometric method.4 Pleasingly, experi-
mental and calculated values for 7a were very similar
(log P ) 0.09 and 0.15, respectively). Examination of
the calculated log P values revealed straightforward
correlation between lipophilicity and cytotoxicity, with
the more lipophilic compound being also more cytotoxic.
However, significant difference in lipohilicity between
7c (log P ) 0.55) and 7d (log P ) 1.59) is reflected in
only a marginal increase of cytotoxicity. Obviously and
not surprisingly, other factors such as steric bulk/steric
hindrance and/or electronic effects might also influence
cytotoxicity of these compounds.
In conclusion, we synthesized a series of 1′-hydroxy-
alkyl-3-methylidenetetrahydro-2-furanones 7a -e with
different substituents on the carbinol carbon atom C-1′
using a unique combination of syn- or anti-dihydroxy-
lation of 4-alkenoic acids (E)-4 or alkenoates (E)-8 and
the Horner-Wadsworth-Emmons olefination reaction.
All prepared compounds were evaluated for their cyto-
toxic activity against L-1210 and HL-60 leukemia cells
and invariably were more cytotoxic against the former
ones. The effect of the substitution at the carbinol
carbon atom C-1′ was also consistent for both cell lines,
with activity increasing from unsubstituted through
alkyl to phenyl substituted compounds. Calculated
lipophilicity increased in the same order. No significant
differences in cytotoxicity of stereoisomers were found.
Further studies to increase the activity of furanones 7
by introducing more lipophilic substituents (R1) and also
by changing their steric bulk and electronic character
are currently in progress in our laboratory.
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-(1′-Hy-
d r oxya lk yl)-3-m eth ylid en etetr a h yd r o-2-fu r a n on es 7. A
mixture of phosphonate 6 (2.0 mmol), K2CO3 (0.829 g; 6.0
mmol), and aqueous 36% formaldehyde solution (1.1 mL; 14.0
mmol) was stirred at 0-5 °C for 15 min. The reaction mixture
was extracted with Et2O (4 × 10 mL), and the combined
organic layers were washed with brine (10 mL), dried, and
evaporated. The crude product was purified by column chro-
matography (silica gel, CHCl3:acetone ) 9:1 as eluent).
5-(H yd r oxym et h yl)-3-m et h ylid en et et r a h yd r o-2-fu r a -
n on e (7a ): oil, 70% yield (lit.10 55%); Rf ) 0.20 (CHCl3: acetone
) 9:1); IR (film) v 3432 (O-H), 1760 (CdO), 1664 (CdC) cm-1
;
1H NMR (CDCl3) δ 2.30 (t, J ) 6.50 Hz, 1H, O-H), 2.86 (ddt,
J ) 17.26, 6.00, 3.00 Hz, 1H, HC-4), 3.00 (ddt, J ) 17.26, 8.00,
3.00 Hz, 1H, HC-4), 3.66 (ddd, J ) 12.51, 6.50, 4.75 Hz, 1H,
H-C-1′), 3.91 (ddd, J ) 12.51, 6.50, 3.00 Hz, 1H, H-C-1′), 4.66
(dddd, J ) 8.00, 6.00, 4.75, 3.00 Hz, 1H, HC-5), 5.68 (t, J )
3.00 Hz, 1H, dCH), 6.26 (t, J ) 3.00 Hz, 1H, dCH); 13C NMR
(CDCl3) δ 27.8 (C-4), 63.0 (C-1′), 76.6 (C-5), 121.4 (dCH2), 133.4
(C-3), 169.7 (C-2). Anal. (C6H8O3) C, H.
5-(Hydr oxym eth yl)-5-m eth yl-3-m eth yliden etetr ah ydr o-
2-fu r a n on e (7b): 63% yield, white solid; mp 67-68 °C (lit.13
63%, mp 66-68 °C); Rf ) 0.26 (CHCl3:acetone ) 9:1); IR (film)
ν 3480 (O-H), 1756 (CdO), 1664 (CdC) cm-1; 1H NMR (CDCl3)
δ 1.40 (s, 3H, CH3C-5), 2.18 (bs, 1H, O-H), 2.63 (dt, J ) 17.01,
2.75 Hz, 1H, HC-4), 3.08 (dt, J ) 17.01, 2.75 Hz, 1H, HC-4),
3.53 (d, J ) 12.01 Hz, 1H, HC-1′), 3.72 (d, J ) 12.01 Hz, 1H,
HC-1′), 5.65 (t, J ) 2.75 Hz, 1H, dCH), 6.24 (t, J ) 2.75 Hz,
1H, dCH); 13C NMR (CDCl3) δ 22.4 (CH3C-4), 34.7 (C-4), 66.9
(C-1′), 82.9 (C-5), 121.1 (dCH2), 134.8 (C-3), 169.4 (C-2). Anal.
(C7H10O3) C, H.
(5R*,1′R*)-5-(1′-Hyd r oxyeth yl)-3-m eth ylid en etetr a h y-
d r o-2-fu r a n on e (7c): mixture of (5R*,1′R*)- and (5R*,1′S*)-
(7c) in 80/20 ratio; oil, 44% yield; Rf ) 0.30 (CHCl3:acetone )
9:1); IR (film) ν 3460 (O-H), 1754 (CdO), 1664 (CdC) cm-1
;
data for (5R*,1′R*)-7c: 1H NMR22 (CDCl3) δ 1.28 (d, J ) 6.50
Hz, 3H, H3C-2′), 2.13 (bs, 1H, O-H), 2.78 (ddt, J ) 17.26, 5.50,
2.75 Hz, 1H, HC-4), 2.99 (ddt, J ) 17.26, 8.00, 2.75 Hz, 1H,
HC-4), 3.78 (quin, J ) 6.50 Hz, 1H, HC-1′), 4.37 (ddd, J )
8.00, 6.50, 5.50 Hz, 1H, HC-5), 5.67 (t, J ) 2.75 Hz, 1H, d
CH), 6.25 (t, J ) 2.75 Hz, 1H, dCH); 13C NMR22 (CDCl3) δ
17.4 (C-2′), 28.7 (C-4), 68.6 (C-1′), 79.7 (C-5), 121.5 (dCH2),
133.0 (C-3), 169.0 (C-2). Anal. (C7H10O3) C, H.
(5R*,1′S*)-5-(1′-Hyd r oxyeth yl)-3-m eth ylid en etetr a h y-
d r o-2-fu r a n on e 7c: mixture of (5R*,1′S*)- and (5R*,1′R*)-
7c in 80/20 ratio; oil, 59% yield; Rf ) 0.30 (CHCl3:acetone )
9:1); IR (film) ν 3456 (O-H), 1752 (CdO), 1664 (CdC) cm-1
;
data for (5R*,1′S*)-7c:1H NMR22 (CDCl3) δ 1.21 (d, J ) 6.50
Hz, 3H, H3C-2′), 1.99 (bs, 1H, O-H), 2.88 (ddt, J ) 17.26, 8.00,
2.50 Hz, 1H, HC-4), 2.99 (ddt, J ) 17.26, 6.50, 2.50 Hz, 1H,
HC-4), 4.12 (qd, J ) 6.50, 3.25 Hz, 1H, HC-1′), 4.43 (ddd, J )
8.00, 6.50, 3.25 Hz, 1H, HC-5), 5.67 (t, J ) 2.50 Hz, 1H, d
CH), 6.26 (t, J ) 2.50 Hz, 1H, dCH); 13C NMR22 (CDCl3) δ
16.6 (C-2′), 26.1 (C-4), 66.5 (C-1′), 79.3 (C-5), 121.2 (dCH2),
133.4 (C-3), 169.5 (C-2). Anal. (C7H10O3) C, H.
(5R*,1′R*)-5-(1′-Hyd r oxybu tyl)-3-m eth ylid en etetr a h y-
d r o-2-fu r a n on e (5R*,1′R*)-7d : oil, 62% yield; Rf ) 0.33
(CHCl3:acetone ) 9:1); IR (film) ν 3450 (O-H), 176054 (Cd
O), 1664 (CdC) cm-1; 1H NMR (CDCl3) δ 0.96 (t, J ) 7.25 Hz,
3H, H3C-4′), 1.33-1.68 (m, 4H, H2C-2′ and H2C-3′), 1.95 (bs,
1H, O-H), 2.85 (ddt, J ) 17.26, 6.25, 3.00 Hz, 1H, HC-4), 2.98
(ddt, J ) 17.26, 8.00, 3.00 Hz, 1H, HC-4), 3.54-3.63 (m, 1H,
HC-1′), 4.44 (ddd, J ) 8.00, 6.25, 4.50 Hz, 1H, HC-5), 5.66 (t,
J ) 3.00 Hz, 1H, dCH), 6.24 (t, J ) 3.00 Hz, 1H, dCH); 13C
NMR (CDCl3) δ 13.8 (C-4′), 18.6 (C-3′), 29.5 (C-4), 34.7 (C-2′),
72.8 (C-1′), 79.5 (C-5), 121.9 (dCH2), 134.2 (C-3), 170.3 (C-2).
Anal. (C9H14O3) C, H.
Exp er im en ta l Section
All reactions requiring anhydrous and oxygen-free condi-
tions were conducted in an argon atmosphere. THF was freshly
distilled from NaH before use. Pyridine was distilled and
stored over KOH. All other solvents were of pure grade and
used as received. Column chromatography was performed on
Fluka silica gel 60 (230-400 mesh). IR spectra were recorded

Brief Articles
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 5 1145
(5R*,1′S*)-5-(1′-Hyd r oxybu tyl)-3-m eth ylid en etetr a h y-
d r o-2-fu r a n on e (5R*,1′S*)-7d : oil, 68% yield; Rf ) 0.33
(CHCl3:acetone ) 9:1); IR (film) ν 3440 (O-H), 1760 (CdO),
1664 (CdC) cm-1; 1H NMR (CDCl3) δ 0.97 (t, J ) 6.75 Hz, 3H,
H3C-4′), 1.25-1.65 (m, 4H, H2C-2′ and H2C-3′), 1.83 (bs, 1H,
O-H), 2.86 (ddt, J ) 17.26, 8.00, 2.50 Hz, 1H, HC-4), 3.01
(ddt, J ) 17.26, 6.00, 2.50 Hz, 1H, HC-4), 3.96 (ddd, J ) 11.59,
6.50, 3.50 Hz, 1H, HC-1′), 4.46 (ddd, J ) 8.00, 6.25, 3.50 Hz,
1H, HC-1′), 5.66 (t, J ) 3.00 Hz, 1H, dCH), 6.24 (t, J ) 3.00
Hz, 1H, dCH); 13C NMR (CDCl3) δ 13.9 (C-4′), 18.7 (C-3′), 27.1
(C-4), 33.8 (C-2′), 71.1 (C-1′), 79.6 (C-5), 121.9 (dCH2), 134.6
(C-3), 170.4 (C-2). Anal. (C9H14O3) C, H.
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(5R*,1′R*)-5-(Hyd r oxy-p h en yl-m eth yl)-3-m eth ylid en e-
tetr a h yd r o-2-fu r a n on e (5R*,1′R*)-7e: oil, 63% yield; IR
(film) ν 3432 (O-H), 1764 (CdO), 1664 (CdC) cm-1; 1H NMR
(CDCl3) δ 2.62 (bs, 1H, OH), 2.65-2.86 (m, 2H, H2C-4), 4.63-
4
4.73 (m, 2H, C-5H, H C1′), 5.59 (t, J ) 2.50 Hz, 1H, dCH),
6.21 (t, 4J ) 2.5 Hz, 1H, dCH), 7.34-7.42 (m, 5H, Ph); 13C
NMR (CDCl3) δ 28.56 (C-4), 75.13 (C-5), 79.11 (C-1′), 121.35
(dCH2), 126.14, 127.59, 127.65, and 137.28 (Ph), 132.91 (C-
3), 169.16 (C-2). Anal. (C12H12O3) C, H.
(5R*,1′S*)-5-(Hyd r oxy-p h en yl-m eth yl)-3-m eth ylid en e-
tetr a h yd r o-2-fu r a n on e (5R*,1′S*)-7e: oil, 41% yield; IR
1
(film) ν 3425 (OH), 1766 (CdO), 1665 (CdC) cm-1; H NMR
(CDCl3) δ 2.47 (bs, 1H, OH), 2.63 (ddt, 2J ) 17.50 Hz, 3J )
8.25 Hz, 4J ) 2.50 Hz, 1H, C-4H), 3.02 (ddt, 2J ) 17.50 Hz, 3J
(9) Harcken, Ch.; Bru¨ckner, R. Stereopure 1,3-butadiene-2-carboxy-
lates and their conversion into nonracemic R-alkylidenebutyro-
lactone natural products by asymmetric dihydroxylation. Tet-
rahedron Lett. 2001, 42, 3967-3971 and references therein.
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4
3
) 5.75 Hz, J ) 2.50 Hz, 1H, C-4H), 4.73 (ddd, J ) 8.25 Hz,
3J ) 5.75 Hz, J ) 3.25 Hz, 1H, C-5H), 5.14 (d, J ) 3.25 Hz,
3
3
1H, C-1′H), 5.60 (t, J ) 2.50 Hz, 1H, dCH), 6.21 (t,4J ) 2.50
4
Hz, 1H, dCH), 7.30-7.40 (m, 5H, Ph); 13C NMR (CDCl3) δ
26.52 (C-4), 72.91 (C-5), 80.11 (C-1′), 121.94 (dCH2), 125.96,
128.03, 128.56 and 138.21 (Ph), 134.46 (C-3), 170.58 (C-2).
Anal. (C12H12O3) C, H.
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a Highly Controlled
Cells a n d Cytotoxicity Assa ys. Mouse leukemia L-1210
cells were cultured in RPMI 1640 medium (Sigma, USA)
supplemented with 10% fetal calf serum (Gibco, USA), gen-
tamycin (50 µg/mL), and 0.02 M HEPES buffer (Gibco, USA)
in a 5% CO2-95% air atmosphere. Cytotoxic effects were
assayed by measuring the inhibitory effects on L-1210 cell
proliferation. In this assay, cells were seeded in 2 mL aliquots
in 6 mL tissue culture tubes (Corning, USA) at a concentration
of 5 × 103 cells/mL and exposed to drugs for 72 h at 37 °C.
The cell number relative to control was then determined by
the colorimetric tetrazolium dye method.23
Human promyelocytic leukemia HL-60 cells were cultured
in RPMI 1640 medium supplemented with 10% foetal calf
serum and gentamycin (50 µg/mL) in a 5% CO2-95% air
atmosphere. Expotentially growing HL-60 cells were seeded
at 6 × 105 per each well of 6-well plate (Numc), and cells were
then exposed to the compounds. Stock solutions were prepared
freshly in RPMI 1640, then dilutions in complete culture
medium were made. HL-60 cells were exposed to compounds
for 72 h. The number of viable cells was counted in a Burker
hemocytometer using trypan-blue exclusion assay. All the data
were expressed as means ( SD.
(12) Bock, K.; Castilla I. M.; Lundt, I.; Pedersen, Ch. Synthesis of
an Optically Active R-Methylene Lactone Starting from Isosac-
charinic Acid. Acta Chem. Scand. 1987, B41, 13-17.
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Approach to Substituted 3-Diethoxyphosphoryl-2,5-dihydro-2-
furanones. Synlett. 2000, 611-614.
(16) J anecki, T.; Błaszczyk, E. A Convenient Synthesis of 3-Alkyli-
denetetrahydro-2-furanones from 3-Diethoxyphosphoryl-2,5-di-
hydro-2-furanones. Synthesis 2001, 403-408.
(17) J anecki, T.; Błaszczyk, E. Stereocontrolled Synthesis of 5-(1′-
Hydroxyalkyl)-3-methylidenetetrahydro-2-furanones. Tetrahe-
dron Lett. 2001, 42, 2919-2922.
(18) Kolb, H. C.; VanNieuwenhze, M. S.; Sharpless, K. B. Catalytic
Asymmetric Dihydroxylation. Chem. Rev. 1994, 94, 2483-2547.
(19) Yamaguchi, S. In Asymmetric Synthesis; Morrison, J . D., Ed.;
Academic Press: New York, 1983; Vol. 1, pp 128-134.
(20) Canetta, R.; Goodlow, J .; Sinaldone, L. Pharamcologic Charac-
teristic of Carboplatin: Clinical Experience, Current Perspec-
tives and Future Directions. Saunders 1990, 19-38.
(21) Leo, A.; Hansch, C.; Church, C. Comparison of Parameters
Currently Used in the Study of Structure-Activity Relation-
ships. J . Med. Chem. 1969, 12, 766-771.
(22) Values for specific isomers were taken from the spectrum of the
mixture of isomers.
(23) Carmichael, J .; DeGraff, W. G.; Gazdar, A. F.; Minna, J . D.;
Mitchell, J . B. Evaluation of Tetrazolium-based Semiautomatic
Colorimetric Assay: Assessment of Chemosensitivity Testing.
Cancer Res. 1987, 47, 936-942.
Ack n ow led gm en t. This work has been partially
supported by the Polish State Committee for Scientific
Research (KBN, Project No. 3 T09A 060 14).
Su p p or tin g In for m a tion Ava ila ble: General procedures
and detailed 1H, 13C, and 31P NMR assignments for all
nontarget compounds and (5R,1′R)-7e, description of the
software package Pallas, some statistics to the log P calcula-
tions, and experimental determination of the log P for 7a . This
material is available free of charge via the Internet at http://
pubs.acs.org.
J M011019V