Antineoplastic agents. 450. Synthesis of (+)-pancratistatin from (+)-narciclasine as relay

Article abstract of DOI:10.1021/jo000710n

(+)-Narciclasine (2) available in quantity from certain Amaryllidaceae species or by total synthesis was employed as a precursor for a 10-step synthetic conversion (3.6% overall yield) to natural (+)-pancratistatin (1a). The key procedures involved epoxidation of natural (+)-narciclasine (2) to epoxide 6, reduction to diol 8, and formation of cyclic sulfate 12 and its ring opening with cesium benzoate followed by saponification of the benzoate to afford (+)-pancratistatin (1a).

Full text of DOI:10.1021/jo000710n

J . Org. Chem. 2001, 66, 2583-2587
2583
An tin eop la stic Agen ts. 450. Syn th esis of (+)-P a n cr a tista tin fr om
(+)-Na r cicla sin e a s Rela y1a
George R. Pettit,* Noeleen Melody, and Delbert L. Herald
Cancer Research Institute and Department of Chemistry and Biochemistry, Arizona State University,
Tempe, Arizona 85287-2404
Received May 9, 2000 (Revised Manuscript Received December 14, 2000)
(+)-Narciclasine (2) available in quantity from certain Amaryllidaceae species or by total synthesis
was employed as a precursor for a 10-step synthetic conversion (3.6% overall yield) to natural (+)-
pancratistatin (1a ). The key procedures involved epoxidation of natural (+)-narciclasine (2) to
epoxide 6, reduction to diol 8, and formation of cyclic sulfate 12 and its ring opening with cesium
benzoate followed by saponification of the benzoate to afford (+)-pancratistatin (1a ).
In tr od u ction
Employing a 1980 Hawaii recollection of Pancratium
(later reassigned Hymenocallis) littorale bulbs, we located
a very potent anticancer constituent. In 1984, we
reported1b,c the isolation (0.028% yield) and structure (by
X-ray of the 7-methoxy derivative) of this important
substance designated (+)-pancratistatin (1a ). Subse-
quently, the U.S. National Cancer Institute initiated
preclinical development owing to the high level of in vitro
and in vivo cancer cell growth inhibitory (including
antiviral)2 activity. Unfortunately, the preclinical devel-
opment of this potentially important anticancer drug has
been slowed by supply constraints and the very low
aqueous solubility (53 µg/mL) properties.3 The latter
problem was finally solved by conversion of (+)-pancrat-
istatin (1a ) to a water-soluble (>230 mg/mL) and com-
parably active phosphate (1b) prodrug.4 While we have
been able to gradually increase supplies of (+)-pancrat-
istatin by cloning and growing the plant in Arizona,5 a
very efficient synthesis of this deceptively simple isocar-
bostyril would be especially useful.
enantioselective synthesis in 14 steps from bromobenzene
(2% overall yield) of (+)-pancratistatin was reported by
Hudlicky,7b in 1995. The same year, Trost7c summarized
a synthesis with an impressive 11% overall yield for 13
steps. More recently, Haseltine7d and Magnus7e (22 steps,
1.2% yield) have presented a new synthesis of (+)-
pancratistatin. In addition, a new synthesis of 7-deoxy-
pancratistatin (1c) has been completed (13 steps, 21%
overall yield),8 and other new approaches to lactam 1a
are in progress.9
From the beginning of our synthetic approaches to
pancratistatin (1a ), narciclasine10 (2) has remained an
attractive precursor, as it is available in practical quanti-
ties from the bulbs of certain Amaryllidaceae species.
Lactam 2 has been studied in some detail,11,12 leading to
its recent synthesis in 12 steps by Hudlicky, starting from
an enzymatic dihydroxylation of m-dibromobenzene.13
Earlier, we attempted to develop a practical synthesis of
pancratistatin from narciclasine (2) and easily obtained
Considerable research efforts6 have been devoted to
developing a very practical synthesis of pancratistatin
(1a ). Four of these have led to pancratistatin. The first
synthesis (Danishefsky7a) provided racemic pancratista-
tin in 26 steps with an overall yield of 0.13%. The first
(1) (a) For the preceding contribution of this series, refer to:
Mohammad, R. M.; Limvarapuss, C.; Wall, N. R.; Hamdy, N.; Beck,
F. W. J .; Pettit, G. R.; Al-Katib, A. Int. J . Oncol. 1999, 15, 367-372.
(b) Pettit, G. R.; Gaddamidi, V.; Cragg, G. M.; Herald, D. L.; Sagawa,
Y. J . Chem. Soc., Chem. Commun. 1984, 1693-1694. (c) Pettit, G. R.;
Gaddamidi, V.; Herald, D. L.; Singh, S. B.; Cragg, G. M.; Schmidt, J .
M.; Boettner, F. E.; Williams, M.; Sagawa, Y. J . Nat. Prod. 1986, 49,
995-1002.
(2) (a) Gabrielsen, B.; Monath, T. P.; Huggins, J . W.; Kevauver, D.
F.; Pettit, G. R.; Groszek, G.; Holingshead, M.; Kirsi, J . J .; Shannon,
W. M.; Schubert, E. M.; DaRe, J .; Ugarkar, B.; Ussery, M. A.; Phelan,
M. J . J . Nat. Prod. 1992, 55, 1569-1581. (b) Gabrielsen, B.; Monath,
T. P.; Huggins, J . W.; Kirsi, J . J .; Holingshead, M.; Shannon, W. M.;
Pettit. G. R. In Natural Products as Antiviral Agents; Chu, C. K.,
Cutler, H. G., Eds.; Plenum: New York, 1992; pp 121-135.
(3) Torres-Labaneira, J . J .; Davignon, P.; Pitha, J . J . Pharm. Sci.
1991, 80, 384-386.
(7) (a) Danishefsky, S.; Lee, J . Y. J . Am. Chem. Soc. 1989, 111,
4829-4837. (b) Tian, X.; Hudlicky, T.; Ko¨nigsberger, K. J . Am. Chem.
Soc. 1995, 117, 3643-3644. (c) Trost, B. M.; Pulley, S. R. J . Am. Chem.
Soc. 1995, 117, 10143-10144. (d) Doyle, T. J .; Hendrix, M. M.; Van
Der Veer, D.; J aanmard, S.; Haseltine, J . Tetrahedron 1997, 53,
11153-11170. (e) Magnus, P.; Sebhat, I. K. Tetrahedron 1998, 54,
15509-15524.
(8) Keck, G. E.; Wagner, T. T.; McHardy, S. F.; J . Org. Chem. 1998,
63, 9164-9165.
(9) (a) Grubb, L. M.; Dowdy, A. L.; Blanchette, H. S.; Friestad, G.
K.; Branchaud, B. P. Tetrahedron Lett. 1999, 40, 2691-2694. (b) Acen˜a,
J . L.; Arjona, O.; Iradier, F.; Plumet, J . Tetrahedron Lett. 1996, 37,
105-106. (c) Gauthier, D. R.; Bender, S. L. Tetraheron Lett. 1996, 37,
13-16.
(10) (a) Ceriotti, G. Nature 1967, 213, 595-596. (b) Okamoto, T.;
Torii, Y.; Isogai, Y. Chem. Pharm. Bull. 1968, 24, 4, 1119-1131. (c)
Mondon, A.; Krohn, K. Chem. Ber. 1975, 108, 445-463.
(11) (a) Rigby, J . H.; Mateo, M. E. J . Am. Chem. Soc. 1997, 119,
12655-12656. (b) Banwell, M. G.; Cowden, C. J .; Gable, R. W. J . Chem.
Soc., Perkin Trans. 1 1994, 3515-3518.
(4) Pettit, G. R.; Freeman, S.; Simpson, M. J .; Thompson, M. A.;
Boyd, M. R.; Williams, M. D.; Pettit, G. R., III; Doubek, D. L. Anti-
Cancer Drug Design 1995, 10, 243-250.
(5) Pettit, G. R.; Pettit, G. R., III; Groszek, G.; Backhaus, R. A.;
Doubek, D. L.; Barr, R. J .; Meerow, A. W. J . Nat. Prod. 1995, 58, 756-
759.
(6) For reviews, refer to: Hoshino, O. The Alkaloids; Cordell, G. A.,
Ed.; Academic Press: San Diego, 1998; Vol. 51, p 323.
10.1021/jo000710n CCC: $20.00 © 2001 American Chemical Society
Published on Web 03/27/2001

2584 J . Org. Chem., Vol. 66, No. 8, 2001
Pettit et al.
Sch em e 1a
a
Key: (a) DMF, DMP, PTSA; (b) Ac2O, pyr; (c) m-CPBA, phosphate buffer; (d) (i) H2, 10% Pd/C; (ii) K2CO3, aq CH3OH; (e) SOCl2, Et3N;
(f) RuCl3, 3H2O/NaIO4, CH3CN, CCl4, H2O; (g) (i) PhCO2H, Cs2CO3; (ii) THF/H2O, cat. H2SO4; (h) K2CO3, CH3OH.
10b-R-hydroxypancratistatin.14 But the last step, hydro-
genolysis of the benzyl alcohol, did not lead to (+)-
pancratistatin. We now report the first synthesis of (+)-
pancratistatin (1a ) from (+)-narciclasine (2) in 3.6%
overall yield.
isolated and identified by NMR. The methyl ether 7,
trans B/C diol 8, cis B/C diol 9, and deprotected epoxide
10 were isolated in 19%, 28%, 27%, and 2% yields,
respectively. Methylation of the C-1 hydroxyl group to
provide ether 7 apparently occurred during the saponi-
fication step.
The 3,4-acetonide of narciclasine (3)10c was prepared
in 97% yield (Scheme 1), and the C-2 and C-7 hydroxyls
were protected by acetylation to afford diacetate 4 in
quantitative yield. However, an attempt at further
purification using silica gel column chromatography
caused some hydrolysis of diacetate 4 to monoacetate 5
in a ratio of 6:1, respectively. Oxidation of olefin 4 using
m-chloroperoxybenzoic acid in dichloromethane and a
phosphate buffer gave R-epoxide 6 in 55% yield. The
structure of epoxide 6 was confirmed by an X-ray
structure determination using a crystal grown from
acetone.15 Hydrogenation of the epoxide in the presence
of 10% palladium on carbon, followed by saponification,
The overall yield of C-1 R-alcohol 8 from narciclasine
1
was 15%. Detailed analysis of the NMR 2D COSY, H-
1H correlation spectrum, allowed assignment of the
hydrogen atoms. The NMR coupling constants (δa 2.93,
δb 3.58, δc 4.31, J ab ) 14.5 Hz, J ac ) 7.0 Hz) for alcohol 8
allowed assignment of the stereochemistry. These as-
signments were confirmed by an X-ray crystal structure
determination using a crystal of 8 grown from methanol
solution.15
Treatment of alcohol 8 with thionyl chloride led to a
high yield of the corresponding epimeric (sulfoxide) cyclic
sulfites 11. Oxidation7c,16a-c of the cyclic sulfite epimers
to the corresponding cyclic sulfate 12 was achieved using
0.18 equiv of ruthenium trichloride with 3.5 equiv of
sodium iodate. Such reactions are usually performed
using a catalytic amount of ruthenium trichloride and
sodium iodate, but that procedure gave incomplete oxida-
tion with only one of the epimeric sulfur atoms oxidized.
Attempts at separating the mixture were not productive.
1
gave a mixture (by H NMR) of four compounds. These
were separated by column chromatography on silica gel,
(12) (a) Krohn, K.; Mondon, A. Chem. Ber. 1976, 109, 9, 855-876.
(b) Banwell, M. G.; Cowden, C. J .; Mackey, M. F. J . Chem. Soc., Chem.
Commun. 1994, 61-62. (c) Khaldi, M.; Chretien, F.; Chapleur, Y.
Tetrahedron Lett. 1995, 36, 3003-3006. (d) Park, T. K.; Danishefsky,
S. J . Tetrahedron Lett. 1995, 36, 195-196. (e) Angle, S. R.; Wada, T.
Tetrahedron Lett. 1997, 38 7955-7958.
(13) Gonzalez, D.; Martinot, T.; Hudlicky, T. Tetrahedron Lett. 1999,
40, 3077-3080.
(14) Pettit, G. R.; Melody, N.; O’Sullivan, M.; Thompson, M.; Herald,
D. L.; Coates, B. J . Chem. Soc., Chem. Commun. 1994, 2725-2726.
(15) Supporting Information available.
(16) (a) Gao, Y.; Sharpless, K. B. J . Am. Chem. Soc. 1988, 110, 7538-
7539. (b) Kim, B. M.; Sharpless, K. B. Tetrahedron Lett. 1989, 30, 655-
658. (c) J eong, L.; Marquez, V. Tetrahedron Lett. 1996, 2353-2356.

Antineoplastic Agents
J . Org. Chem., Vol. 66, No. 8, 2001 2585
Ta ble 1. Hu m a n Ca n cer a n d Mu r in e Lym p h ocytic Leu k em ia Cell Lin e In h ibition Va lu es for P a n cr a tista tin (1a ) a n d
Na r cicla sin e (2) a n d Syn th etic In ter m ed ia tes (3-13)
isocarbostyril
cell type
cell line
1a
2
3
4
5
6
7
8
9
10
12
13
GI50 (µg/mL) pancreas-a
BXPC-3
Ovcar-3
SF-295
2.8 × 10-2 1.6 × 10-2 >10 >10 >10 >10 >10
3.2 × 10-2 1.6 × 10-2 >10 >10 >10 >10 >10
1.7 × 10-2 1.2 × 10-2 >10 >10 >10 >10 >10
2.6 × 10-2 1.8 × 10-2 >10 >10 >10 >10 >10
1.6 × 10-2 7.1 × 10-3 >10 >10 >10 >10 >10
1.4 × 10-1 4.5 × 10-2 >10 >10 >10 >10 >10 >10
1.1 × 10-1 4.4 × 10-2 >10 >10 >10 >10 >10 >10
9.1 × 10-2 4.9 × 10-2 >10 >10 >10 >10 >10 >10
8.8 >10 >10 >10
9.4 × 10-4
1.3 × 10-3
ovarian
5.0 >10 >10 >10 <1.0 × 10-3
CNS
lung-NSC
colon
7.2 >10 >10 >10
NCI-H460 4.8 × 10-2 2.2 × 10-2 >10 >10 >10 >10 >10 >10
>10 >10 >10 <1.0 × 10-3
KM20L2
DU-145
BXPC-3
Ovcar-3
SF-295
6.9 >10 >10 >10 <1.0 × 10-3
prostate
3.8 >10 >10 >10 <1.0 × 10-3
TGI (µg/mL) pancreas-a
>10 >10 >10
>10 >10 >10
>10 >10 >10
>10 >10 >10
>10 >10 >10
>10 >10 >10
5.3 × 10-3
1.9 × 10-3
4.9 × 10-3
1.1 × 10-3
3.4 × 10-3
2.6 × 10-3
0.0017
ovarian
CNS
lung-NSC
colon
prostate
NCI-H460 4.3 × 10-1 1.0 × 10-1 >10 >10 >10 >10 >10 >10
KM20L2
DU-145
2.3 × 10-1 1.2 × 10-2 >10 >10 >10 >10 >10 >10
10.0 × 10-2 3.6 × 10-2 >10 >10 >10 >10 >10 >10
ED50 (µg/mL) murine leukemia P388
0.032
0.0044
>10 >10 >10 >10 >10
2.6 >10 >10 >10
the monoacetate (5, 0.15 g, 4.2%): Rf 0.79 (98:2, CH2Cl2-CH3-
Using the excess oxidant method, cyclic sulfate 12 was
obtained in 47% yield from alcohol 8.
OH); [R]27 +84 (c 1.01, CHCl3); mp 250-253 °C; 1H NMR
D
(CDCl3, 300 MHz) δ 12.85 (s, 1H), 6.65 (s, 1H), 6.4 (s, 1H),
6.09 (m, 2H), 5.36 (m, 1H), 4.3 (dd, J ) 5.4, 6.3 Hz, 1H), 4.12
(m, 2H), 2.18 (s, 3H), 1.51 (s, 3H), 1.37 (s, 3H); 13C NMR (CDCl3
300 MHz) δ 170.3, 167.6, 153, 146, 134.7, 128, 127.9, 121.5,
111.6, 104.9, 102.4, 94.5, 78.5, 75.3, 74, 55.2, 27.1, 24.9, 21.1;
EIMS m/z 389 (M+, 37), 331 (33), 289 (74), 271 (29), 260 (42),
247 (100), 242 (43), 218 (10), 85 (7), 44 (59). Anal. Calcd for
C19H19O9N: C, 58.6; H, 4.9; N, 3.59. Found: C, 58.2, H, 5.0,
N, 3.4.
Nucleophilic attack by cesium benzoate7c,16a on the
cyclic sulfate 12 in dimethylforamide occurred over 4 h.
Hydrolysis of the alkyl sulfate in tetrahydrofuran using
sulfuric acid allowed simultaneous cleavage of the ac-
etonide to afford, following column chromatography,
benzoate 13 in 74% yield. The benzoyl group was easily
removed to give (+)-pancratistatin (1a ) identical with the
natural product.1 (+)-Pancratistatin (1a ) was obtained
in 10 steps with an overall yield of 3.6% from narciclasine
(2).
Synthetic conversion of (+)-narciclasine (2) to (+)-
pancratistatin (1a ) provided a useful opportunity to
further study structure/activity relationships, and the
result (Table 1) was a clear demonstration that even
minor structural modifications of pancratistatin (1a ) led
to decreased cancer cell growth inhibitory activity in all
but the case of 13, where the activity was greatly
enhanced by the presence of a C-1 benzoyl group.
The diacetate (4, 3.2 g, 81%) exhibited: Rf 0.6 (98:2 CH2-
Cl2-CH3OH); [R]24 ) +60° (c 0.91, MeOH); mp 130-132 °C;
D
1H NMR (CDCl3 300 MHz) δ 6.97 (s, 1H), 6.12 (s, 1H), 6.08 (s,
2H), 6.01 (s, 1H), 5.37 (m, 1H), 4.40 (dd, J ) 5.7, 6.6 Hz, 1H),
4.12 (m, 2H), 2.38 (s, 3H), 2.21 (s, 3H), 1.51 (s, 3H), 1.38 (s,
3H); 13C NMR (CDCl3 300 MHz) 170.3, 169.2, 160.9, 152.4,
141.4, 134.0, 129.5, 128.5, 121.9, 113.4, 111.4, 103.0, 100.1,
78.7, 75.3, 73.8, 55.1, 27.11, 24.9, 21.1, 20.9; EIMS m/z 431
(M+, 14), 389 (60), 373 (28), 331 (75), 314 (9), 289 (100), 271
(35), 260 (42), 247 (99), 242 (39), 85 (9), 44 (95). Anal. Calcd
for C21H21O9N‚CH3OH; C, 57.02; H, 5.43; N, 3.25. Found: C,
57.16, H, 4.99, N, 3.11.
1,10b-R-Ep oxy-2,7-d ia cetoxyn a r cicla sin e 3,4-a ceton id e
(6). Diacetate 4 (3.0 g, 6.96 mmol) was dissolved in CH2Cl2
(180 mL), and phosphate buffer (pH 8) (180 mL) was added
followed by m-chloroperoxybenzoic acid (4.5 g, 26 mmol, 3.7
equiv). The biphasic mixture was stirred for 16 h, whereby all
the starting material had reacted (TLC). Dichloromethane (300
mL) was added, and the organic layer was separated, thor-
oughly washed with 5% Na2S2O3 (3 × 300 mL), followed by
5% Na2CO3 (3 × 300 mL) and water (3 × 300 mL), dried
(MgSO4), and concentrated. The crude product was washed
with acetone and the white precipitate collected to give an
amorphous solid (1.6 g, 52% yield). Recrystallization from hot
acetone gave crystals which were examined (see below) by
X-ray crystallography: mp 224-225 °C; [R]24D ) +138° (c 1.06,
CH2Cl2); 1H NMR (300 MHz CDCl3) δ 6.46 (s, 1H), 6.1 (m, 2H),
5.86 (bs, 1H), 5.3 (d, J ) 6 Hz, 1H), 4.4 (dd, J ) 6.3, 6.9 Hz,
1H), 4.3 (t, J ) 8.7 Hz, 1H), 4.06 (s, 1H), 4.03 (d, J ) 8.4 Hz),
2.38 (s, 3H), 2.24 (s, 3H), 1.47 (s, 3H), 1.34 (s, 3H); 13C NMR
(300 MHz, CDCl3) 170.5, 168.8, 161.3, 152.4, 142, 134, 128.4,
118.6, 109.38, 103.2, 101.1, 75.7, 75.4, 74.0, 58.1, 55.1, 53.4,
26.8, 24.2, 21.0, 20.8; EIMS M+ 447 (M+, 10), 405 (84), 345
(10), 287 (33), 258 (16), 234 (54), 44 (100). Anal. Calcd for
Exp er im en ta l Section
Na r cicla sin e 3,4-Aceton id e (3).10c A solution prepared
from natural (+)-narciclasine (2, 5 g, 16.3 mmol), 2,2 dimethox-
ypropane (25 mL), and p-toluenesulfonic acid (0.83 g) in DMF
(25 mL) was stirred at rt for 16 h. Pyridine (8.5 mL) followed
by water (150 mL) was added to the suspension and the
mixture stirred for 1 h at rt. The precipitated product was
collected, washed with water, and dried (at 64 °C over P2O5
under high vacuum) to give narciclasine 3,4-acetonide (5.5 g,
97%) as an amorphous powder: mp 270-273 °C (lit.10c mp 275
°C); IR (KBr) 3497, 3184, 1676, 1599, 1467, 1373, 1294, 1111,
1039 cm-1; HNMR δ (DMSO, 300 MHz) 13.7 (s, 1H), 8.8 (s,
1
1H), 7.01 (s, 1H), 6.47 (bs, 1H), 6.06 (m, 2H), 5.73 (bs, 1H),
4.2-3.9 (m, 4H), 1.45 (s, 3H), 1.31 (s, 1H); 13CNMR (DMSO,
300 MHz) δ 167.6, 152.5, 145.2, 133.3, 128.8, 128.3, 125.9,
109.8, 104.3, 102.0, 94.2, 79.0; EIMS m/z 347 (M+, 28), 332
(1), 289 (2), 273 (6), 260 (8), 247 (100), 242 (22), 218 (10), 85
(2), 73 (10).
2,7-Dia cetoxyn a r cicla sin e 3,4-Aceton id e (4) a n d 2-Ac-
etoxyn a r cicla sin e 3,4-Aceton id e (5). Narciclasine 3,4-ac-
etonide (3, 3.18 g, 9.15 mmol) was dissolved in acetic anhydride
(20 mL)-pyridine (20 mL). The mixture was heated and
stirred at 60 °C (under argon) for 24 h. The suspension slowly
dissolved. Ethyl acetate (300 mL) was added, and the organic
layer was washed with water (2 × 40 mL), dried (Na2SO4),
and concentrated. The product was dried under high vacuum
for 24 h to remove traces of pyridine. The amorphous powder
was examined by 1H NMR (CDCl3) and found to be the
diacetate (4) with a trace of pyridine still present. Column
chromatographic purification was conducted on silica gel using
a gradient elution system 1:99 f 1:9 (CH2Cl2-CH3OH) to give
C
21H21O10N: C, 56.37; H, 4.73; N, 3.13. Found: C, 56.72; H,
4.83; N, 3.09.
X-r a y Cr ysta l Str u ctu r e Deter m in a tion . Ep oxid e 6. A
thick, plate-shaped X-ray sample (∼0.70 × 0.60 × 0.40 mm)
was obtained by crystallization from acetone. Data collection
was performed at 293 ( 1 K. Accurate cell dimensions were
determined by least-squares fitting of 25 carefully centered
reflections in the range of 35° < θ < 40° using Cu KR radiation.
Cr ysta l Da ta : C21H21O10N1, FW ) 447.39, monoclinic, C2,
a ) 18.200(4) Å, b ) 9.2435(18) Å, c ) 12.864(3) Å, â ) 105.73

2586 J . Org. Chem., Vol. 66, No. 8, 2001
Pettit et al.
(3)°, V ) 2083.2(7) Å3, Z ) 4, Fc ) 1.426 Mg/m3, µ(Cu KR) )
0.982 mm-1, λ ) 1.541 78 Å, F(000) 936.
g, 18.7%), diol 8 (0.68 g, 27.7%), diol 9 (0.31 g), and a mixture
of acetate 9 and epoxide 10 (0.78 g). The mixture of 9 and 10
was separated by dissolution in hot methanol and collection
of the insoluble material (10, 0.051 g, 2%). Recrystallization
of the mother liquor gave alcohol 9 (0.38 g). The total amount
of alcohol 9 recovered from the column was 0.68 g (27%).
Recrystallization of diol 8 from hot methanol gave crystals that
were used for X-ray crystal structure elucidation (see below).
Methyl ether 7 corresponded to: Rf 0.53 (CH2Cl2/CH3OH, 4%);
[R]24D ) 15° (c 1.1, CH3OH); mp 239-240 °C; 1H NMR (DMSO,
500 MHz) δ 12.8 (s, 1H), 9.39 (s, 1H), 6.77 (s, 1H), 6.1 (dd, J
) 3.5, 9.0 Hz, 2H), 5.17 (d, J ) 7.5 Hz, 1H), 4.68 (d, J ) 3 Hz,
1H), 4.63 (t, J ) 7 Hz, 1H), 3.86 (m, 1H), 3.42 (s, 3H), 2.58 (d,
J ) 8 Hz, 1H), 1.48 (s, 3H), 1.46 (s, 3H); 13C NMR (DMSO,
500 MHz) 165.6, 154.4, 144.8, 134.3, 132.3, 111.5, 110.5, 108.5,
102.5, 92.9, 77.7, 74.8, 73.2, 71.5, 58.0, 27.2, 25.2; EIMS m/z
377 (M+, 100), 362 (3), 349 (5), 319 (8), 288 (14), 270 (14), 258
(35), 247 (17), 292 (8), 218 (13), 101 (24), 56 (14), 44 (24), 28
(34). Anal. Calcd for C18H19O8N‚CH3OH: C, 55.89; H, 5.39; N,
3.43. Found: C, 56.06; H, 5.39; N, 3.46.
Diol 8 was characterized with Rf 0.39 (96:4 CH2Cl2-CH3-
OH); [R]24D ) -26° (c 0.48, CH3OH); mp 231-232 °C; 1H NMR
(CDCl3 500 MHz) δ 12.45 (s, 1H), 6.9 (s, 1H), 6.18 (s, 1H), 6.03
(narrow m, 1H), 4.38 (t, J ) 8 Hz, 1H), 4.31 (m, 2H), 3.92 (dd,
J ) 5, 7.25 Hz, 1H), 3.57 (dd, J ) 14.5, 8.5 Hz, 1H), 3.05 (bs,
1H), 2.93 (dd, J ) 14.5, 7.5 Hz), 2.68 (bs, 1H), 1.48 (s, 3H),
1.38 (s, 3H); 13C NMR (CDCl3, 500 MHz) 169.8, 153.2, 146.2,
135.4, 133.1, 110.9, 106.3, 102.3, 98.8, 77.6, 75.5, 70.8, 70.6,
52.9, 41.0, 27.3, 24.9; EIMS m/z 365 (M+, 100), 350 (7), 290
(3), 272 (4), 260 (4), 234 (9), 205 (35), 190 (4), 176 (5), 147 (5),
85 (5), 73 (7), 60 (10), 44 (9), 28 (1). Anal. Calcd for
All reflections corresponding to a complete quadrant (0 e h
e 13, 0 e k e 9, -19 e l e 19) were collected over the range
of 0 < 2θ < 110° using the ω/2θ scan technique. Friedel
reflections were also collected (whenever possible) immediately
after each reflection. Three intensity control reflections were
also measured for every 60 min of X-ray exposure time and
showed a maximum variation of 0.9% over the course of the
collection. A total of 6050 reflections were collected. Subse-
quent statistical analysis of the complete reflection data set
using the XPREP17 program verified that the space group as
C2. After Lorentz and polarization corrections, merging of
equivalent reflections and rejection of systematic absences,
2605 unique reflections (R(int) ) 0.0524) remained, of which
2583 were considered observed (Io > 2σ(Io)) and were used in
the subsequent structure determination and refinement. Lin-
ear and anisotropic decay corrections were applied to the
intensity data as well as an empirical absorption correction
(based on a series of psi-scans).18 Structure determination
was readily accomplished with the direct-methods program
SHELXS.19 All non-hydrogen atom coordinates were located
in a routine run using default values in that program. The
remaining H atom coordinates for the parent molecule were
calculated at optimum positions. All non-hydrogen atoms were
refined anisotropically in a full-matrix least-squares refine-
ment using SHELXL.19 The H atoms were included, their Uiso
thermal parameters fixed at either 1.2 or 1.5 (depending upon
their atomic environment) of the value of the Uiso of the atom
to which they were attached and forced to ride that atom. The
final standard residual R1 value for 6 was 0.066 for observed
data and 0.0663 for all data. The corresponding Sheldrick R
values were wR2 of 0.1551 and 0.1555, respectively. The
goodness-of-fit on F2 was 1.101. The structure of epoxide 6 is
shown in Figure S1 (Supporting Information). The absolute
stereochemistry of the epoxide15 could be assigned with
certainty on the basis of the value of the Flack absolute
structure parameter,20 i.e., -0.1(3). A final difference Fourier
map showed minimal residual electron density; the largest
difference peak and hole being +0.474 and -0.417 e/Å3. Final
bond distances and angles were all within expected and
acceptable limits.
1-Meth oxyison a r cicla sin e 3,4-Aceton id e (7), 1-Isop a n -
cr a tista tin 3,4-Aceton id e (8), B/C cis-1-Isop a n cr a tista tin
3,4-Aceton id e (9), a n d 1,10b-R-Ep oxyn a r cicla sin e 3,4-
Aceton id e (10). The epoxide 6 was hydrogenated and the
resulting mixture saponified in 3 × 1 g batches as follows. To
a solution of epoxide 6 (1 g) in ethyl acetate (150 mL) was
added 10% palladium-on-carbon (1 g). The flask was evacu-
ated, flushed with hydrogen (5×), and hydrogenated at rt for
2.5 h using a hydrogen-filled balloon. The palladium-on-carbon
was collected and the filtrate concentrated to dryness to give
a white solid (0.94 g). The 1H NMR (CDCl3) indicated a mixture
of products. The solid was dissolved in 10% aq CH2OH (20 mL)
and DCM (10 mL). Potassium carbonate (0.6 g, 4.4 mmol) was
added, and the reaction mixture stirred overnight at rt. A
precipitate slowly developed. The reaction mixture was neu-
tralized using IR-120 H+ Amberlite resin. The resin was
collected and the filtrate concentrated to a yellow solid (0.91
g). The results of three such reactions were combined to give
a yellow solid (2.65 g) which was purified by silica gel flash
chromatography using 98:2 DCM/MeOH to give alcohol 7 (0.46
C
17H19O8N: 55.89; H, 5.24; N, 3.83. Found: C, 55.5; H, 5.42;
N, 3.85.
B/C-cis-d iol 9 showed: Rf 0.13 (96:4 CH2Cl2-CH3OH); [R]D
) +27° (c 0.86, CH3OH); mp 245 °C; 1H NMR (DMSO, 500
MHz) δ 13.06 (s, 1H), 8.08 (s, 1H), 6.55 (s, 1H), 6.01 (s, 2H),
4.94 (d, J ) 6.5 Hz, 1H), 4.53 (d, J ) 4 Hz, 1H), 4.38 (d, J )
4.5 Hz, 1H), 4.1 (d, J ) 5.5 Hz, 1H), 4.05 (dd, J ) 8.5, 5.5 Hz,
1H), 3.58 (m, 1H), 3.52 (m, 1H), 3.01 (m, 1H), 1.42 (s, 3H),
1.29 (s, 3H); 13C NMR (DMSO, 500 MHz) 169.5, 151.5, 144.4,
135.7, 131.9, 108.6, 107.1, 101.6, 99.5, 76.1, 75.5, 74.9, 71.9,
49.6, 38.7, 28.3, 26.5; EIMS m/z 365 (M+, 100), 350 (6), 305
(3.5), 258 (7), 234 (31.6), 205 (77), 176 (7), 147 (10), 119 (5),
100 (9), 85 (12), 73 (17.5), 60 (24), 44 (22), 28 (22). Anal. Calcd
for C17H19O8N‚H2O: C, 53.3; H, 5.5; N, 3.6. Found: C, 53.6;
H, 5.5; N, 3.6.
Ep oxid e 10: Rf 0.15 (96:4 CH2Cl2-CH3OH); [R]22 ) -4.2°
D
1
(c 0.57, THF); mp 242-244 °C; H NMR (DMSO 300 MHz) δ
11.9 (s, 1H), 6.83 (s, 1H), 6.12 (s, 1H), 5.09 (dd, J ) 9, 11 Hz,
1H), 4.96 (d, J ) 10 Hz, 1H), 4.66 (dd, J ) 3.6, 6 Hz, 1H), 4.33
(dd, J ) 6.6, 3 Hz, 1H), 3.53 (m, 1H), 1.38 (s, 3H), 1.36 (s, 3H);
13C NMR (DMSO, 300 MHz) δ 166.4, 154.2, 144.3, 132.0, 131.3,
114.9, 109.9, 108.9, 102.9, 100.2, 94.5, 76.2, 72.4, 71.2, 66.3,
28.7, 26.7; EIMS m/z 363 (M+, 100), 345 (5), 305 (22), 288 (10),
276 (13), 258 (39), 233 (21), 205 (11), 60 (14), 44 (33), 28 (34).
X-r a y Cr ysta l Str u ctu r e Deter m in a tion . Diol a ce-
ton id e 8: A plate-shaped crystal (∼0.35 × 0.20 × 0.10 mm)
was obtained by crystallization from methanol and mounted
in a sealed capillary with the specimen immersed in mother
liquor. Data collection was performed at 293 ( 1 K. Accurate
cell dimensions were determined by least-squares fitting of 25
carefully centered reflections in the range of 35° < θ < 40°
using Cu KR radiation.
Cr ysta l Da ta : C17H19O8N1‚CH3OH, FW ) 397.37, triclinic,
P1, a ) 7.9890(12) Å, b ) 9.072(2) Å, c ) 13.564(2) Å, R )
87.805(16)°, â ) 82.317(13)°, γ ) 71.734(16)°, V ) 925.1(3) Å3,
Z ) 2, Fc ) 1.426 Mg/m3, µ(Cu KR) ) 0.982 mm-1, λ ) 1.541 78
Å, F(000) 420.
All reflections corresponding to a complete hemisphere (0
e h e 9, -10 e k e 10, -15 e l e 15) were collected over the
range of 0 < 2θ < 120° using the ω/2θ scan technique. Friedel
reflections were also collected (whenever possible) immediately
after each reflection. Three intensity control reflections were
also measured for every 60 min of X-ray exposure time and
showed a maximum variation of -1.5% over the course of the
(17) XPREP: The automatic space group determination program
in the SHELXTL. (see ref 19).
(18) North, A. C.; Phillips, D. C.; Mathews, F. S. Acta Crystallogr.
1968 A2, 351.
(19) SHELXTL-PC Version 5.101, 1997, an integrated software
system for the determination of crystal structures from diffraction data,
Bruker Analytical X-ray Systems, Inc., Madison, WI 53719. This
package includes, among others: XPREP, an automatic space group
determination program; XS, the Bruker SHELXS module for the
solution of X-ray crystal structures from diffraction data; XL, the
Bruker SHELXL module for structure refinement; XP, the Bruker
interactive graphics display module for crystal structures.
(20) Flack, H. D. Acta Crystallogr. 1983, 876-881.

Antineoplastic Agents
J . Org. Chem., Vol. 66, No. 8, 2001 2587
collection. A total of 6235 reflections were collected. Subse-
quent statistical analysis of the complete reflection data set
using the XPREP17 program verified that the space group as
P1. After Lorentz and polarization corrections, merging of
equivalent reflections and rejection of systematic reflections,
a total of 5550 were considered observed (Io > 2σ(Io)) and were
used in the subsequent structure determination and refine-
ment. Linear and anisotropic decay corrections were applied
to the intensity data as well as an empirical absorption
correction (based on a series of ψ-scans).18 The structure was
solved with the direct-methods program SHELXS.19 All non-
hydrogen atom coordinates were located in a routine run using
default values in that program. The asymmetric unit (i.e., unit
cell) was found to contain two independent molecules of the
parent compound, as well as two molecules of disordered
methanol solvent. The remaining H atom coordinates for the
parent molecules were calculated at optimum positions. All
non-hydrogen atoms were refined anisotropically in a full-
matrix least-squares refinement using SHELXL.19 The H
atoms were included, their Uiso thermal parameters fixed at
either 1.2 or 1.5 (depending upon their atomic environment)
of the value of the Uiso of the atom to which they were
attached and forced to ride that atom. The final standard
residual R1 value for diol acetonide 8 was 0.0661 for observed
data and 0.0766 for all data. The corresponding Sheldrick R
values were wR2 of 0.1863 and 0.2048, respectively. The
goodness-of-fit on F2 was 1.065. The structure of one of the
molecules of the diol acetonide 8, present in the unit cell, is
shown in Figure S2 (Supporting Information). The Flack
absolute structure parameter20 was -0.4(3). A final difference
Fourier map showed minimal residual electron density; the
largest difference peak and hole being +0.585 and -0.398 e/Å3.
Final bond distances and angles were all within expected and
acceptable limits.
vacuum), and the residue was suspended in THF (2 mL).
Water (three drops from a pipet) followed by H2SO4 (concen-
trated two drops from a pipet)7c was added, and the suspension
became a solution. The solution was stirred at 70 °C for 24 h.
Additional THF (2 mL), water (two drops), and H2SO4 (two
drops) were added, and the reaction was allowed to continue
for 2 h. The crude reaction mixture was separated by passing
through a silica gel flash column (9:1 CH2Cl2-CH3OH). The
product was collected and recrystallized by dissolving in 9:1
CH2Cl2-CH3OH, concentrating to a viscous oil, followed by
solution in chloroform and addition of hexane to turbidity. The
benzoate (13) recrystallized at rt as a colorless amorphous solid
weighing 0.066 g, 74%: [R]24 ) -21° (c 0.19 CH3OH); mp
D
1
180-185 °C; H NMR (DMSO, 500 MHz) δ 13.2 (s, 1H), 7.98
(m, 3H), 767 (t, J ) 9 Hz, 1H), 7.52 (t, J ) 10 Hz, 2H), 6.25 (s,
1H), 6.07 (d, J ) 1.5 Hz, 1H), 6.02 (d, J ) 1.5 Hz, 1H), 5.8 (bs,
1H), 5.68 (s, 1H), 4.96 (bs, 1H), 4.14-4.09 (m, 2H), 3.93-3.86
(m, 2H), 3.37 (m, 1H); 13C NMR (DMSO, 500 MHz) δ 169.3,
165.4, 152.1, 145.7, 134.0, 133.4, 132.2, 129.6, 129.5, 129.1,
128.6, 128.3, 107.3, 101.9, 95.7, 71.7, 70.0, 69.9, 68.8, 50.6, 38.0;
EIMS m/z 429 (M+, 12.5), 325 (3.5), 307 (9.8), 271 (15.2), 247
(50), 218 (9.8), 205 (7), 122 (69.6), 105 (100), 77 (80), 52 (46),
28 (53). Anal. Calcd for C21H19O9N‚H2O: C, 56.38; H, 4.73; N,
3.13. Found: C,56.82; H,4.61; N, 2.95.
(+)-P a n cr a tista tin (1a ). A methanol (0.5 mL) solution of
benzoate 13 (0.025 g, 0.058 mmol) was treated with K2CO3
for 16 h at rt. TLC (5:1 CH2Cl2-CH3OH) showed product
development with starting material still present. The reaction
mixture was heated to 55-60 °C for 3 h, whereby TLC showed
no starting material present. The mixture was concentrated
to a brown solid, and the residue was dissolved using a mixture
of DMF (0.4 mL) and 9:1 CH2Cl2-CH3OH. The crude material
was purified by passage through a column of silica gel with
4:1 CH2Cl2-CH3OH as the eluent, to afford (+)-pancratistatin
(1a ) as an off-white solid (0.014 g, 75%): Rf 0.38 (4:1 CH2Cl2-
1-Isop a n cr a t ist a t in 1,2-Cyclic Su lfa t e-3,4-a cet on id e
(12). To a solution of diol 8 (0.25 g, 0.67 mmol) in THF (10
mL)-triethylamine (0.4 mL, 2.9 mmol) was added thionyl
chloride (0.1 mL, 1.37 mmol, 2 equiv). The reaction was carried
out at rt under argon for 40 min. Ethyl acetate (50 mL) was
added, and the organic layer was washed with water (2 × 10
mL) followed by brine (10 mL). After drying and concentration,
an off-white solid (0.26 g, 93%) was obtained. The 1H NMR
(CDCl3, 300 MHz) indicated the sulfite product (11) was
obtained as a mixture of epimers. The solid (0.26 g, 0.63 mmol)
was dissolved in CH3CN (15 mL)-CCl4 (12 mL), and RuCl3‚
H2O (0.024 g, 0.115 mmol, 0.18 equiv) and NaIO4 (0.48 g, 2.24
mmol, 3.5 equiv) were added together as a solid, followed by
water (12 mL). The mixture was stirred at rt for 19 h. Ethyl
acetate (80 mL) was added and the organic layer separated,
washed with water (2 × 15 mL), dried (MgSO4), filtered
through a pad of silica gel, and concentrated to a brown solid
that was dried under high vacuum for 16 h (0.13 g, 45%).
Recrystallization from THF/hexane provided 12 as a white
amorphous solid: Rf 0.7 (96:4 CH2Cl2-CH3OH); [R]24D ) +7.3
1
CH3OH); mp 265-270 °C dec; H NMR (DMSO, 500 MHz) δ
13.05 (s, 1H), 7.49 (s, 1H), 6.48 (s, 1H), 6.05 (s, 1H), 6.02 (s,
1H), 5.36 (bs, 1H), 5.07 (m, 2H), 4.82 (d, J ) 7 Hz, 1H), 4.27
(s, 1H), 3.95 (bs, 1H), 3.84 (bs, 1H), 3.72 (m, 2H), 2.96 (d, J )
12.5 Hz, 1H); 13C NMR (DMSO, 500 MHz) d 169.5, 152.0,
145.4, 135.6, 131.7, 107.5, 101.7, 97.7, 73.3, 70.2, 69.9, 68.5,
50.5. Comparison (NMR, IR, TLC) with an authentic speci-
men1b,c of natural (+)-pancratistatin confirmed the mutual
identity of the synthetic and natural samples.
Ack n ow led gm en t. The very necessary financial
support for this investigation was provided by Out-
standing Investigator Grant CA-44344-01A1-01-12 from
the Division of Cancer Treatment and Diagnosis, Na-
tional Cancer Institute, DHHS; the Arizona Disease
Control Research Commission; the Fannie E. Rippel
Foundation; Robert B. Dalton Endowment Fund; Gary
L. and Diane R. Tooker; Sally Schloegel; Billie J ane
Baguley; the Fraternal Order of Eagles Art Ehrmann
Cancer Fund; and the Ladies Auxiliary to the Veterans
of Foreign Wars. We are also very pleased to thank for
other assistance Professor Barry M. Trost (Stanford
University), Drs. Gordon M. Cragg, Ralph A. Backhaus,
and Dennis L. Doubek, Professors Cherry L. Herald and
Fiona Hogan, Dr. Sheo Bux Singh, George R. Pettit III,
and the U. S. National Science Foundation (Grant No.
CHE-9808678).
1
(c 0.15, THF); mp 223 °C; H NMR (DMSO, 500 MHz) δ 13.1
(s, 1H) 9.06 (s, 1H), 6.39 (s, 1H), 6.11 (d, J ) 1 Hz, 1H), 6.09
(d, J ) 1 Hz, 1H), 5.58 (t, J ) 7.5 Hz, 1H), 5.26 (dd, J ) 7.0,
9.0 Hz, 1H), 4.71 (dd, J ) 7.8, 9.3 Hz, 1H), 4.6 (t, J ) 8 Hz,
1H), 3.74 (dd, J ) 7.8, 14.8 Hz, 1H), 3.66 (dd, J ) 8, 14 Hz,
1H), 1.47 (s, 3H), 1.35 (s, 3H); 13C NMR (DMSO, 500 MHz) δ
168.3, 152.5, 145.5, 133.0, 132.5, 110.8, 102.3, 97.5, 84.4, 82.5,
77.0, 73.2, 56.0, 50.2, 37.2, 27.1, 25.0; EIMS m/z 427 (M+, 100),
412 (10), 396 (1.7), 370 (2.6), 352 (3.5), 327 (10.5), 272 (21),
242 (36), 218 (10.5), 205 (29), 147 (17), 119 (9.6), 85 (8.7), 60
(1.7), 44 (42). Anal. Calcd for C17H17O16NS: C, 47.8; H, 4.01;
N, 3.27. Found: C, 48.08; H, 4.16; N, 3.13.
Su p p or tin g In for m a tion Ava ila ble: Tables of crystal-
lographic data, bond lengths and angles, atomic coordinates,
and anisotropic thermal parameters are available for struc-
tures 6 and 8. This material is available free of charge via the
Internet at http://pubs.acs.org.
P a n cr a tista tin 1-Ben zoa te (13). To a solution of cyclic
sulfate 12 (0.09 g, 0.21 mmol) in DMF (2 mL) was added
benzoic acid (0.042 g, 0.36 mmol, 1.7 equiv) followed by Cs2-
CO3 (0.1 g, 0.31 mmol, 1.5 equiv). The mixture was stirred
under argon at 60 °C for 4 h. The DMF was removed (high
J O000710N