Topoisomerase I-mediated antiproliferative activity of enantiomerically pure fluorinated homocamptothecins

Article abstract of DOI:10.1021/jm000129j

Homocamptothecin (hCPT) is an E-ring modified camptothecin (CPT) analogue bearing a methylene spacer between the alcohol and carboxyl functions of the CPT lactone. Combining pronounced inhibitory activity of topoisomerase I (Topo I) with enhanced plasma s

Full text of DOI:10.1021/jm000129j

J . Med. Chem. 2000, 43, 2285-2289
2285
Top oisom er a se I-Med ia ted An tip r olifer a tive Activity of En a n tiom er ica lly P u r e
F lu or in a ted Hom oca m p toth ecin s
Olivier Lavergne,*,§ Danie`le Demarquay,§ Christian Bailly,¶ Christophe Lanco,§ Alain Rolland,§ Marion Huchet,§
Hele`ne Coulomb,§ Nicole Muller,§ Nicole Baroggi,§ J ose´ Camara,§ Christine Le Breton,‡ Eric Manginot,‡
J ean-Bernard Cazaux,‡ and Dennis C. H. Bigg§
Institut Henri Beaufour, 5, avenue du Canada, F-91966 Les Ulis, France, Centre Oscar Lambret - INSERM U-524, IRCL,
place de Verdun, F-59045 Lille, France, and Expansia, route d’Avignon, F-30390 Aramon, France
Received March 17, 2000
Homocamptothecin (hCPT) is an E-ring modified camptothecin (CPT) analogue bearing a
methylene spacer between the alcohol and carboxyl functions of the CPT lactone. Combining
pronounced inhibitory activity of topoisomerase I (Topo I) with enhanced plasma stablility,
hCPT constitutes an attractive template for the elaboration of new anticancer agents.
Fluorinated hCPT analogues, prepared in enantiomerically pure form, were assayed by their
stimulation of Topo I-mediated DNA cleavage. Translation into cytotoxicity against tumor cells
was evaluated on HT29 human colon adenocarcinoma and on the multidrug resistant lung
and bladder tumor cell lines, A549 and T24r. Good correlation is observed between the ability
of the drugs to stimulate Topo I-mediated DNA cleavage and the respective 50% inhibitory
concentrations (IC50 values) of the HT29, A549, and T24r cell growth. Fluorine substitution in
the A-ring of hCPT was found to have a pronounced influence on biological activity, providing
several compounds which are up to 100-fold more potent than CPT in terms of IC50. Among
these, 10,11-difluoro-hCPT has been selected for further development.
Camptothecin derivatives (CPTs) constitute a promis-
ing class of anticancer agents, exerting potent cytotox-
icity by an original mode of action which converts the
ubiquitous nuclear enzyme topoisomerase I (Topo I) into
a cellular poison via the stabilization of Topo I-DNA
covalent complexes.1 Based on the CPT template, the
current generation of compounds suffers, however, from
the intrinsic instability of the highly electrophilic R-hy-
droxylactone which, under physiological conditions,
undergoes rapid hydrolysis to an equilibrium mixture
in which the biologically inactive carboxylate form
predominates.2
We have previously reported the insertion of a meth-
ylene spacer between the alcohol and carboxyl functions
of CPT to give hCPT.3 This change to the CPT lactone
uniquely conserves Topo I-mediated activity4 and was
found to promote additional DNA breaks which are not
seen with CPT.5 Another advantage comes from the
diminished electrophilicity of the lactone, providing
hCPT with a slow hydrolysis instead of the fast inter-
conversion observed with CPT. Furthermore, even
under mildly acidic conditions, this hydrolysis is ir-
reversible, thus avoiding the possibility of haemorrhagic
cystitis, a known limiting toxicity of some CPT ana-
logues. Subsequent synthesis and screening of racemic
analogues showed fluorinated hCPTs to be worthy of
further investigation.6 Fluorine substitution of the CPT
skeleton, although reported in several instances,7-11 has
not been previously underlined as a key feature for
activity enhancement. The changes in sequence-specific-
ity of the drug-induced DNA cleavage by Topo I imply
somewhat different interactions within the cleavable
complexes5 and, hence, that the SAR of CPTs may not
be totally applicable to hCPTs. This paper describes our
approach to select an optimal fluorine substitution
pattern.
The biologically active enantiomers of CPT and hCPT
were presumed to have identical spatial arrangement
for their ethyl and hydroxyl groups, based on their
similarity of their stereospecific Topo I inhibition and
positive rotation of polarized sodium D emission. The
biologically active counterpart of (S)-CPT was therefore
(R)-hCPT, since homologation of the lactone changes the
substituent priority ruled by chemical nomenclature.
Our previously established synthetic strategy was ame-
nable to the preparation of enantiopure compounds via
a classical resolution. As shown in Scheme 1, racemic
â-hydroxyacid 2 was resolved with quinidine to give the
desired (R)-form in an enantiomeric excess (ee) of 70%,
and a structural analysis by X-ray diffraction of the
crystalline quinidium salt confirmed the absolute con-
figuration assignment. Removal of the benzyl protecting
group, lactonization, and demethylation gave pyridone
7, whose enantiomeric purity was brought above 99%
ee by recrystallization from acetone/water. The prepara-
tion was completed by coupling 7 with a fluorinated
quinoline, and subsequent formation of the C-ring by
intramolecular Heck reaction.12 The required quinolines
were obtained from the corresponding fluoroanilines by
the Meth-Cohn method,6 except for hCPT 8a which was
prepared from the corresponding fluoroanthranilic acid.
* To whom requests for reprints and correspondence should be
addressed. E-mail: olivier.lavergne@beaufour-ipsen.com. Fax: (33) 01
69 07 38 02.
§ Institut Henri Beaufour.
¶
Centre Oscar Lambret.
‡ Expansia.
10.1021/jm000129j CCC: $19.00 © 2000 American Chemical Society
Published on Web 05/10/2000

2286 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 11
Brief Articles
Sch em e 1a
Ta ble 1. Biological Data for Fluorinated hCPTs
cytotoxicity against
substituents
Topo I
tumor cells (IC50, nM)b
test
compd R1 R2 R3 R4 clvd DNAa HT29c A549c T24rc
8a
8b
8c
8d
8e
8f
hCPT
CPT
F
18
17
29
20
27
11
190
10
8.4
17
22
1.6
4.5
0.17
22
0.32
0.40
3.3
F
F
F
2.9
130
1.9
3.4
26
67
F
7.3
F
F
F
30
32
21
30
80
8.2
88
a
Stimulation of pKMp27 DNA cleavage by human Topo I in
the presence of 0.1 µM test compound, calculated with the formula
(N-N0)/N0 where N and N0 correspond to the fraction of nicked
DNA in the drug-treated and the drug-free runs, separated by
electrophoresis on an ethidium bromide containing agarose gel.5
Data were compiled from quantitative analysis of three gels and
a
Reagents and conditions: (a) TFA, rt, 90%; (b) quinidine,
b
i-PrOH, 40%; (c) Pd/C HCOONH4, MeOH, rt, 88%; (d) DCC, THF,
50 °C; (e) TMSCl, NaI, MeCN, reflux, 34% from 5, then recrys-
tallization from i-PrOH, 71%; (f) fluorinated hydroxymethylquino-
line, DEAD, PPh3, DMF, rt, 40-50%; (g) Pd(OAc)2, PPh3, KOAc,
TBAB, MeCN, reflux, 40-65%.
must be considered as a set of averaged values. Mean values of
50% inhibitory concentration determined by WST-1 (soluble tet-
razolium) assay in triplicate.4 c HT29 and A549 cell lines were
obtained from the ATCC (Rockville, MD), and the T24r cell line13
was a gift from Dr. Robert Kiss, Universite´ Libre de Bruxelles,
Belgium.
The activity of these fluorinated hCPTs was assayed
with human Topo I on supercoiled plasmid pKMp27,
and the products were analyzed on an agarose gel
containing ethidium bromide.4 In the presence of the
drugs, the nicked DNA fraction was significantly in-
creased in a dose-dependent manner, reflecting the
stabilization of Topo I-DNA cleavable complexes. Quan-
tification of this effect in the presence of 0.1 µM of test
compound is reported in Table 1. Stimulation of DNA
cleavage is more efficient with 8c, fluorinated at R3,
than with compounds 8a and 8b, with a fluorine at R1
or R2, respectively. At 0.1 µM, the stimulation of DNA
cleavage by 8d is about 3 times lower than with 8c, R4
being the least favorable position for fluorine substitu-
tion. The DNA cleavage values obtained for unsubti-
tuted hCPT are comparable with that of 8a or 8b, while
difluorinated compounds 8e and 8f are, like 8c, highly
efficient in stimulating DNA cleavage.
Translation of the Topo I inhibitory activity into
antitumor cytotoxicity was determined by antiprolif-
erative assays on three human cancer cell lines. HT29
colon adenocarcinoma and A549 non-small cell lung
cancer were obtained from the American Type Culture
Collection, while T24r resulted from prolonged exposure
of T24 bladder carcinoma to a cocktail of three antican-
cer drugs comprising vinorelbine, an investigational
alkylating agent PE1001, and doxorubicin.13 A549 and
T24r were selected for their multidrug resistance char-
acteristics, established by flow cytometry and RT-PCR
to be due to the overexpression of MRP for A549,14 and
Pg-P for T24r.15 Cultured cells were incubated for 72 h
with the drugs, and the viable cells were quantified by
WST-1 colorimetric assay.4 The 50% inhibitory concen-
trations (IC50s) are presented in Table 1. The antipro-
liferative activities of the compounds were found to be
in agreement with their stimulation of DNA cleavage
(Figure 1). Graph A illustrates the trend of the IC50
values on HT29, in decreasing order when going from
8d to 8f and matching the increasing capacity of the
compounds to stimulate DNA cleavage. Cell lines which
overexpress P-gp have been previously shown to have
a certain degree of cross-resistance to hydroxylated CPT
analogues such as topotecan and SN-38.6 The remark-
able levels of activity of compounds 8a -f on the A549
and T24r cell lines demonstrate, on the contrary, that
the multidrug resistant phenotypes of these cells do not
recognize fluorinated hCPTs. Here again, a good cor-
relation is observed between the relative DNA cleavage
activities of compounds 8a -f and their IC50 values on
A549 and T24r (Graphs B and C), indicating that
fluorine substitution does not introduce any bias with
respect to multidrug resistance.
In summary, fluorine substitution in the A-ring of
hCPT is found to have pronounced influence on biologi-
cal activity, and, depending on the substitution pattern,
cytotoxic activity relative to CPT may be increased
considerably on multidrug resistant tumor cell lines.
The good correlation that exists between the stimulation
of DNA cleavage and cytotoxicity is a rather critical
aspect of structure-activity studies of CPT deriva-
tives16,17 which has not always been verified,18 due to
physicochemical differences which alter cellular pen-
etration and intracellular concentration of the drugs.19
That such a good match is found for fluorinated hCPTs
confirms Topo I poisoning as their principal mechanism
of action, and may be due to the small elemental radius
of fluorine and the narrow range of lipophilicities
involved. Within the series of compounds presented
here, hCPTs 8c, 8e, and 8f, all bearing a fluorine atoms
at R3, constitute a group of highly active Topo I poisons,
which are also more resistant to hydrolysis because of

Brief Articles
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 11 2287
crystals, collected by filtration, were further purified by
crystallization from isopropyl alcohol to give the diastereo-
merically pure quinidine salt of acid 4, suitable for X-ray
diffraction to determine its absolute configuration: mp >250
°C; [R]D ) +168 (c 0.16, MeOH); IR (KBr) 837, 1067, 1230,
20
1241, 1391, 1508, 1552, 1591, 1619 cm-1; 1H NMR (DMSO-d6)
δ 0.66 (t, 3H), 1.23 (m, 1H), 1.58 (m, 2H), 1.81 (m, 2H), 1.85
(m, 1H), 2.05 (t, 1H), 2.35 (q, 1H), 2.54 (d, 1H), 2.77 (m, 1H),
2.82 (d, 1H), 2.95 (m, 2H), 3.35 (m, 4H), 3.80 (s, 3H), 3.91 (s,
3H), 4.53 (s, 2H), 4.75 (d, 1H), 5.13 (t, 2H), 5.62 (s, 1H), 6.04
(m, 2H), 7.05 (d, 1H), 7.35 (m, 7H), 7.54 (d, 1H), 7.94 (d, 1H),
7.98 (d, 1H), 8.70 (d, 1H); 13C NMR (DMSO-d6) δ 8.35, 20.88,
21.76, 24.81, 27.51, 34.70, 38.41, 46.18, 47.93, 48.69, 53.58,
55.73, 59.84, 63.36, 68.97, 72.21, 75.86, 102.17, 115.45, 116.25,
117.62, 119.04, 121.38, 126.52, 127.44, 127.71, 128.24, 131.36,
138.78, 139.96, 143.96, 145.28, 147.61, 157.28, 157.30, 163.55,
174.51. Anal. (C20H24N2O2‚C19H23NO5‚H2O) C, H, N. The
filtrate was evaporated under reduced pressure, and the
residue, taken up in dichloromethane (270 mL), was washed
with 1 N HCl (270 mL), dried, and concentrated to give the
desired enantiomerically enriched acid 3 (13.5 g, 70% ee, (5µ-
Chiral-AGP 4 × 100 mm, 1.2 mL/min isopropyl alcohol/
water/10 mM (pH 6.5) phosphate buffer 30/920/50, UV 280
nm)) as an oil: IR (neat) 699, 737, 825, 1064, 1453, 1556, 1594,
1
1712, 1952 cm-1; H NMR (DMSO-d6) δ 0.67 (t, 3H), 1.88 (m,
2H), 2.76 (d, 1H), 2.98 (d, 1H), 3.85 (s, 3H), 4.53 (s, 2H), 4.76
(s, 2H), 5.75 (s, 1H), 7.04 (d, 1H), 7.31 (m, 5H), 8.03 (d, 1H),
12 (br, 1H); 13C NMR (DMSO-d6) δ 8.27, 34.66, 46.09, 53.79,
63.31, 72.22, 76.11, 116.44, 117.49, 127.63, 127.86, 128.40,
138.77, 145.58, 156.66, 163.58, 172.79. A crystal of the quini-
dine salt of 4 was analyzed on
a CAD4 (Enraf-Nonius,
Bohemia, NY) spectrometer using Cu KR emission, and the
structure was solved and refined with the SHELXS86 and
SHELXL93 (Sheldrick, G.M., University of Gottingen, Ger-
many) least-squares algorithms. The resulting three-dimen-
sional structure revealed compound 4 to be a 3S-3-hydroxy-
pentanoic acid and, by deduction, compound 3 to be a 3R-3-
hydroxy-pentanoic acid.
F igu r e 1. Correlations, by least-squares linear regression,
between the ability for the drugs to stimulate Topo I-mediated
DNA cleavage at 0.1 µM and their 50% inhibitory concentra-
tions logarithm values on HT29, A549, and T24r (r2 ) 0.90,
0.93, and 0.89, respectively). The error bars correspond to the
confidence intervals of the IC50 determinations. The symbols
used are: P for CPT, H for hCPT, and a to f for compounds
8a -f.
(5R)-5-Eth yl-1,4,5,8-tetr a h yd r o-5-h yd r oxy-oxep in o[3,4-
c]p yr id in e-3,9-d ion e (7). To a solution of acid 3 (13.5 g, 39
mmol) in methanol (90 mL) was added 10% palladium on
charcoal (50% wet, 28 g), and the resulting suspension was
degassed and treated under nitrogen with
a solution of
ammonium formate (11.5 g, 183 mmol) in methanol (135 mL).
The reaction mixture was stirred at room temperature for 30
min, then heated to 40 °C for 30 min, and allowed to cool to
room temperature. The supported catalyst was removed by
filtration, and the filtrate was evaporated under reduced
pressure. Removal of residual traces of methanol was achieved
by the addition of toluene (40 mL) and evaporation under
reduced pressure. The resulting oil was dissolved in dry THF
(50 mL) and treated with dicyclohexylcarbodiimide (7.2 g, 34.5
mmol) in THF (20 mL), and the solution was heated at 50 °C
for 1 h before cooling to room temperature. The urea byproduct
was removed by filtration, and the filtrate was concentrated
under reduced pressure. The oily residue was taken up in dry
acetonitrile (46 mL), treated with sodium iodide (6 g, 40.5
mmol) and trimethylsilyl chloride (5.2 mL, 40.5 mmol), and
stirred for 5 h at room temperature. The reaction mixture was
then diluted with acetonitrile (28 mL) and water (5.6 mL) to
give a precipitate which was collected by filtration and taken
up in water (10 mL). The resulting suspension was neutralized
to pH 7.5 with ammonia, and the precipitate was collected by
filtration and dried under reduced pressure to give the desired
product (4.2 g, 34% yield, 77% ee (Chirosebond Cl-5µ 4.6 ×
250mm, 1 mL/min dichloromethane/isopropyl alcohol 88/12,
UV 300 nm)) as an off-white solid. Recrystallization from
acetone (40 mL) and water (150 mL) provided enantiomerically
pure pyridone 7 (3g, 99.7% ee) as white crystals: mp >250
their modified lactone. After 2 h of incubation in human
plasma at 37 °C, more than 50% of difluorinated hCPT
8f remained in the biologically active lactone form,
whereas CPT decayed exponentially to reach an equi-
librium of less than 10% lactone within 30 min. Further
work has shown 8f, also known as BN 80915, to be
orally active at very low doses in a variety of human
tumor xenografts.20
Exp er im en ta l Section
Resolu tion of 3-(3-Ben zyloxym eth yl-2-m eth oxy-4-p y-
r id yl)-3-h yd r oxy-p en ta n oic Acid (2). Racemic tert-butyl
ester 16 (40 g, 100 mmol) was treated with trifluoroacetic acid
(150 mL) at room temperature for 18 h. The reaction mixture
was concentrated under reduced pressure, and the residue was
taken up in dichloromethane (200 mL) and extracted with
saturated sodium bicarbonate (2 × 100 mL). The combined
aqueous extracts were washed with dichloromethane (100 mL)
and acidified to pH 1 with 6 N HCl. Extraction with dichlo-
romethane (2 × 200 mL), drying of the combined extracts over
MgSO4, and concentration under reduced pressure gave race-
mic acid 2 which was engaged in the subsequent resolution
without further purification. A mixture of acid 2 (31.1 g, 90
mmol) and quinidine (29.2 g, 90 mmol) was stirred in isopropyl
alcohol (60 mL) at 50 °C. The clear solution obtained was
allowed to cool to room temperature and then stored at 4 °C
until crystal seeds appeared (16 h). Crystallization was then
completed with stirring at room temperature for 2 h. The
°C; [R]D ) +125 (c 0.154, DMSO); IR (KBr) 812, 1041, 1181,
20
1284, 1530, 1563, 1626, 1746 cm-1; 1H NMR (DMSO-d6) δ 0.77
(t, 3H), 1.66 (m, 2H), 3.12 (dd, 2H), 5.24 (dd, 2H), 5.65 (br,
1H), 6.30 (d, 1H), 7.29 (d, 1H), 11.59 (br, 1H); 13C NMR

2288 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 11
Brief Articles
(DMSO-d6) δ 8.23, 35.89, 42.44, 61.15, 72.84, 105.02, 122.73,
133.86, 155.78, 161.31, 172.04. Anal. (C11H13NO4) C, H, N.
151.81 (dd, 253, J 16 Hz), 153.32, 155.77, 159.05, 171.80. Anal.
(C21H16F2N2O4‚0.75H2O) C, H, N.
F lu or in a ted h CP Ts. The following compounds were pre-
pared by coupling under Mitsunobu conditions of 7 and the
appropriate fluorinated hydroxymethylquinoline, followed by
an intramolecular Heck reaction, according to the general
method previously reported for racemic compounds.6 It was
checked by HPLC on chiral stationary phase that the tertiary
alcohol did not epimerize.
Ack n ow led gm en t. We thank Edith Bresson-Navet
and Gilles Mario for their technical assistance at IHB,
as well as Daniel Mancier and colleagues, and the
chemical process development group at Expansia, for
analytical measurements and supply of enantiopure DE-
synthon.
(5R)-5-Eth yl-11-flu or o-5-h yd r oxy-4,5,13,15-tetr a h yd r o-
1H,3H- oxep in o[3′,4′:6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8a ): mp >250 °C; IR (KBr) 815, 1205, 1612, 1629, 1660,
Refer en ces
1
1734 cm-1; H NMR (DMSO-d6) δ 0.87 (t, 3H), 1.86 (q, 2H),
(1) Lerchen, H.-G. Camptothecin Antitumor Agents. IDrugs 1999,
2, 896-906.
(2) Burke, T. G. Chemistry of the Camptothecins in the Blood-
stream. Drug Stabilization and Optimization of Activity. Ann.
N.Y. Acad. Sci. 1996, 803, 29-31.
(3) Lavergne, O.; Lesueur-Ginot, L.; Pla Rodas, F.; Bigg, D. C. H.
BN 80245: An E-Ring Modified Camptothecin With Potent
Antiproliferative and Topoisomerase I Inhibitory Activities.
Bioorg. Med. Chem. Lett. 1997, 7, 2235-2238.
(4) Lesueur-Ginot, L.; Demarquay, D.; Kiss, R.; Kasprzyk, P. G.;
Dassonneville, L.; Bailly, C.; Camara, J .; Lavergne, O.; Bigg, D.
C. H. Homocamptothecin, an E-Ring Modified Camptothecin
With Enhanced Lactone Stability, Retains Topoisomerase I-
Targeted Activity and Antitumor Properties. Cancer Res. 1999,
59, 2939-2943.
(5) Bailly, C.; Lansiaux, A.; Dassonneville, L.; Demarquay, D.;
Coulomb, H.; Huchet, M.; Lavergne, O.; Bigg, D. C. H. Ho-
mocamptothecin, an E-Ring Modified Camptothecin Analogue,
Generates New Topoisomerase I-Mediated DNA Breaks. Bio-
chemistry 1999, 38, 15556-15556.
(6) Lavergne, O.; Lesueur-Ginot, L.; Pla Rodas, F.; Kasprzyk, P. G.;
Pommier, J .; Demarquay, D.; Pre´vost, G.; Ulibarri, G.; Rolland,
A.; Schiano-Liberatore, A. M.; Harnett, J .; Pons, D.; Camara,
J .; Bigg, D. C. H. Homocamptothecins: Synthesis and Antitumor
Activity of Novel E-Ring-Modified Camptothecin Analogues. J .
Med. Chem. 1998, 41, 5410-5419.
(7) Lackey, K.; Besterman, J . M.; Fletcher, W.; Leitner, P.; Morton,
B.; Sternbach, D. D. Rigid Analogs of Camptothecin As DNA
Topoisomerase I Inhibitors. J . Med. Chem. 1995, 38, 906-911.
(8) J osien, H.; Bom, D.; Curran, D. P.; Zheng, Y. H.; Chou, T. C.
7-Silylcamptothecins (Silatecans): A New Familly of Camptoth-
ecin Antitumor Agents. Bioorg. Med. Chem. Lett. 1997, 7, 3189-
3194.
3.07 (d, 1H), 3.46 (d, 1H), 5.28 (s, 2H), 5.40 (d, 1H), 5.53 (d,
1H), 6.03 (s, 1H), 7.43 (s, 1H), 7.55 (t, 1H), 7.85 (q, 1H), 8.01
(d, 1H), 8.82 (s, 1H); 13C NMR (DMSO-d6) δ 8.35, 36.40, 42.49,
50.86, 61.37, 73.25, 100.13, 111.50 (d), 118.30 (d), 123.12,
124.80 (d), 125.41 (d), 130.28 (d), 130.50, 144.19, 148.79,
153.89, 155.83, 157.55 (d, J 253 Hz), 159.15, 171.93. Anal.
(C21H17FN2O4‚0.25H2O) C, H, N.
(5R)-5-Eth yl-10-flu or o-1,4,5,13-tetr a h yd r o-5-h yd r oxy-
3H,15H-oxep in o[3′,4′:6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8b): mp > 250 °C; IR (KBr) 828, 1201, 1508, 1585,
1
1651, 1754 cm-1; H NMR (DMSO-d6) δ 0.86 (t, 3H), 1.83 (q,
2H), 3.05 (d, 1H), 3.47 (d, 1H), 5.26 (s, 2H), 5.39 (d, 1H), 5.52
(d, 1H), 6.05 (s, 1H), 7.38 (s, 1H), 7.78 (m, 1H), 7.96 (d, 1H),
8.21 (m, 1H), 8.65 (s, 1H); 13C NMR (DMSO-d6) δ 8.47, 36.50,
42.53, 50.70, 61.44, 73.35, 99.70, 111.90 (d), 120.72 (d), 122.77,
129.00 (d), 130.95, 131.24 (d), 131.96 (d), 144.49, 145.28, 152.61
(d), 155.92, 159.23, 160.46 (d, J 247 Hz), 172.11. Anal. (C21H17
FN2O4‚0.5H2O) C, H, N.
-
(5R)-5-E t h yl-9-flu or o-1,4,5,13-t et r a h yd r o-5-h yd r oxy-
3H,15H-oxep in o[3′,4′ :6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8c): mp > 250 °C; IR (KBr) 837, 1226, 1505, 1587, 1653
cm-1; 1H NMR (DMSO-d6) δ 0.86 (t, 3H), 1.84 (q, 2H), 3.04 (d,
1H), 3.47 (d, 1H), 5.24 (s, 2H), 5.39 (d, 1H), 5.52 (d, 1H), 6.06
(s, 1H), 7.39 (s, 1H), 7.65 (t, 1H), 7.89 (d, 1H), 8.23 (dd, 1H),
8.72 (s, 1H); 13C NMR (DMSO-d6) δ 8.35, 36.41, 42.51, 50.56,
61.39, 73.24, 99.97, 112.47 (d), 118.03 (d), 122.91, 125.35,
129.62 (d), 131.29 (d), 131.94, 144.33, 148.99 (d), 153.78,
155.83, 159.16, 162.93 (d, J 249 Hz), 171.95. Anal. (C21H17
-
(9) J oto, N.; Ishii, M.; Minami, M.; Kuga, H.; Mitsui, I.; Tohgo, A.
FN2O4) C, H, N.
DX-8951f,
a Water-Soluble Camptothecin Analog, Exhibits
(5R)-5-E t h yl-8-flu or o-1,4,5,13-t et r a h yd r o-5-h yd r oxy-
3H,15H-oxep in o[3′,4′:6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8d ): mp > 250 °C; IR (KBr) 763, 1211, 1508, 1613,
Potent Antitumor Activity Against a Human Lung Cancer Cell
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1
1659, 1731 cm-1; H NMR (DMSO-d6) δ 0.88 (t, 3H), 1.87 (q,
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2H), 3.08 (d, 1H), 3.47 (d, 1H), 5.29 (s, 2H), 5.40 (d, 1H), 5.53
(d, 1H), 6.06 (s, 1H), 7.44 (s, 1H), 7.69 (m, 2H), 7.96 (m, 1H),
8.75 (s, 1H); 13C NMR (DMSO-d6) δ 8.34, 36.37, 42.44, 50.62,
61.38, 73.29, 100.03, 114.52 (d), 123.00, 124.61 (d), 127.72 (d),
129.70 (d), 131.16, 131.70, 138.06 (d), 144.27, 153.08, 155.93,
157.45 (d, J 255 Hz), 159.15, 171.95. Anal. (C21H17FN2O4‚
0.25H2O) C, H, N.
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(5R)-5-Eth yl-9,11-diflu or o-1,4,5,13-tetr ah ydr o-5-h ydr oxy-
3H,15H-oxep in o[3′,4′:6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8e): mp 227 °C (dec); IR (KBr) 854, 1245, 1505, 1638,
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1748 cm-l 1H NMR (DMSO-d6) δ 0.87 (t, 3H), 1.85 (q, 2H),
;
3.07 (d, 1H), 3.46 (d, 1H), 5.26 (s, 2H), 5.40 (d, 1H), 5.52 (d,
1H), 6.03 (s, 1H), 7.42 (s, 1H), 7.73 (t, 1H), 7.81 (d, 1H) 8.82
(s, 1H); 13C NMR (DMSO-d6) δ 8.32, 36.07, 42.49, 50.79, 61.34,
73.21, 100.43, 103.33 (dd), 109.29 (dd), 115.81 (d), 123.38,
125.28 (d), 130.19, 143.87, 148.79 (dd), 155.03, 155.80, 158.23
(dd, J 255, 15 Hz), 159.09, 162.20 (dd, J 248, 14 Hz), 171.90.
Anal. (C21H16F2N2O4‚0.25H2O) C, H, N.
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(5R)-5-Eth yl-9,10-diflu or o-1,4,5,13-tetr ah ydr o-5-h ydr oxy-
3H,15H-oxep in o[3′,4′:6,7]in d olizin o[1,2-b]qu in olin e-3,15-
d ion e (8f): mp > 250 °C; IR (KBr) 871, 1261, 1512, 1579, 1654,
1
1746 cm-l; H NMR (DMSO-d6) δ 0.87 (t, 3H), 1.85 (m, 2H),
3.08 (d, 1H), 3.44 (d, 1H), 5.26 (s, 2H), 5.39 (d, 1H), 5.52 (d,
1H), 5.99 (s, 1H), 7.38 (s, 1H), 8.14 (dd, 1H), 8.23 (dd, 1H),
8.67 (s, 1H); 13C NMR (DMSO-d6) δ 8.23, 36.33, 42.50, 50.54,
61.33, 73.14, 99.80, 114.32 (d), 115.30 (d), 122.87, 125.38 (d),
130.42, 131.32, 144.09, 145.24 (d), 149.58 (dd, J 250, 16 Hz),

Brief Articles
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