Unnatural C-1 homologues of pancratistatin-Synthesis and promising biological activities

Article abstract of DOI:10.1139/v2012-073

Several C-1 homologues of pancratistatin and 7-deoxypancratistatin were synthesized by a phenanthrene-phenathridone oxidative recyclization strategy. The key steps involved the enzymatic dihydroxylation of bromobenzene, addition of an aryl alane to an epo

Full text of DOI:10.1139/v2012-073

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Unnatural C-1 homologues of pancratistatin —
Synthesis and promising biological activities
Sergey Vshyvenko, Jon Scattolon, Tomas Hudlicky, Anntherese E. Romero,
Alexander Kornienko, Dennis Ma, Ian Tuffley, and Siyaram Pandey
Abstract: Several C-1 homologues of pancratistatin and 7-deoxypancratistatin were synthesized by a phenanthrene–phenathri-
done oxidative recyclization strategy. The key steps involved the enzymatic dihydroxylation of bromobenzene, addition
of an aryl alane to an epoxyaziridine, an intramolecular aziridine opening on silica gel in solid phase, and the above-men-
tioned recylization strategy. Experimental and spectral data are reported for all new compounds. All synthesized C-1 ho-
mologues of pancratistatin and 7-deoxypancratistatin were evaluated for antiproliferative activity in a panel of human
cancer cell lines. As expected, the 7-hydroxy compounds were found to be more potent and the activity of the C-1
benzoxymethyl analogue exceeded that of narciclasine, which was used as a positive control.
Key words: C-1 homologues of pancratistatin, amaryllidaceae alkaloids, anticancer activities, intramolecular aziridine open-
ing, solid-phase silica catalysis.
Résumé : On a réalisé la synthèse de plusieurs homologues en C-1 de la pancratistatine et de la 7-désoxypencratistatine en
faisant appel à une stratégie de recyclisation oxydante phenantrène–phénathridone. Les étapes clés impliquent la dihydroxy-
lation du bromobenzène, l’addition d’une arylalane à une époxyaziridine, une ouverture intramoléculaire d’aziridine sur gel
de silice en phase solide et la stratégie de recyclisation mentionnée plus haut. Les données expérimentales et spectrales sont
rapportées pour tous les nouveaux produits. Tous les homologues en C-1 de la pancratistatine et de la 7-désoxypencratista-
tine ont été évalués pour leur activité à contrer la prolifération dans un éventail de lignées de cellules cancéreuses humaines.
Tel que prévu, les composés 7-hydroxy sont les plus puissants alors que l’activité de l’analogue C-1 benzoxyméthyle est su-
périeure à celle de la narciclasine qui a été utilisée comme contrôle positif.
Mots‐clés : analogues en C-1 de la pancratistatine, alcaloïdes des amaryllidacées, activité anticancéreuse, ouverture intramo-
léculaire d’aziridine, catalyse par le gel de silice en phase solide.
[Traduit par la Rédaction]
of truncated derivatives from the groups of Pettit,4 McNulty,5
Introduction
Kornienko,6 Chapleur,7 Alonso,8 Marion,9 Banwell,10 and
Gonzalez.11 Truncated derivatives of the core of these con-
stituents as well as various heteroatom analogues have been
reported and some of these compounds displayed various lev-
els of biological activity, usually of lower potency than the
natural products. The only exceptions were compounds that
contained a large lipophilic fragment in position C-1. The
first example of such an analogue was the benzoate ester of
the C-1 hydroxyl of pancratistatin, prepared and tested by
Pettit et al.,4c which displayed nanomolar activity. Later, a li-
brary of C-1 nitrogenated derivatives prepared by Marion et
al.9 showed the highest activity with substituents such as
benzamide.
Pancratistatin (1), narciclasine (2), and other Amaryllida-
ceae constituents, Fig. 1, continue to attract attention on
account of their promising anticancer properties. Since the
isolation of pancratistatin in 1984 by Pettit et al.1 and its pre-
liminary evaluation as a potent antitumor agent, many crea-
tive syntheses of 1 and its congeners have been reported.2
The activity concerning the total synthesis of these constitu-
ents continues unabated to the present day.3 More recently,
however, the studies of these natural products have shifted to-
ward the investigation of unnatural derivatives that would of-
fer the potential of better bioavailability than the rather
insoluble natural products. In the last two decades, many re-
ports have appeared describing the synthesis and evaluation
Received 3 August 2012. Accepted 8 August 2012. Published at www.nrcresearchpress.com/cjc on .
S. Vshyvenko, J. Scattolon, and T. Hudlicky. Chemistry Department and Centre for Biotechnology, Brock University, 500 Glenridge
Avenue, St. Catharines, ON L2S 3A1, Canada.
A.E. Romero and A. Kornienko. Chemistry Department, New Mexico Institute of Mining and Technology, 801 Leroy Place, Socorro,
NM 87801, USA.
D. Ma, I. Tuffley, and S. Pandey. Department of Chemistry and Biochemistry, University of Windsor, 401 Sunset Avenue, Windsor, ON
N9B 3P4, Canada.
Corresponding author: Tomas Hudlicky (e-mail: thudlicky@brocku.ca).
Dedicated to Professor Derrick Clive in recognition of his outstanding contributions to the art and craft of organic synthesis.
Can. J. Chem. 90: 1–12 (2012)
doi:10.1139/V2012-073
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Can. J. Chem. Vol. 90, 2012
Fig. 1. Pancratistatin, other Amaryllidaceae congeners, and C-1 homologues.
Scheme 1.
cant over-reduction of 15. The intramolecular aziridine open-
ing was accomplished by heating the tert-butyldimethylsilyl
triflate (TBS)-protected ether 17 adsorbed on silica gel ac-
cording to a solid-phase protocol developed in our labora-
tory for an aziridine opening of this type.14–16 It is worth
mentioning that the addition of 10% w/w quinoline to the
substrate during the solid-phase cyclization helps to sup-
press the decomposition of material and improves the yield
of this reaction from 45% to 74%, probably because of the
decreased acidity of the silica gel.
The product of the silica-catalyzed cyclization, phenan-
threne 18, was subjected to several different sets of oxidative
protocols. The originally disclosed conditions,16 which in-
cluded OsO4/NaIO4-promoted oxidation–recyclization, did
not lead to the desired product. This was probably the result
of an increased steric hindrance by the methoxy group in the
environment of the double bond (this interaction was absent
in the cyclization of the precursor to 7-deoxypancratistatin).
Instead, a different protocol was developed, which included
selective ozonolysis of the double bond in the presence of azo
dye Sudan Red 7B to help prevent oxidation of the electron-
rich aromatic ring. Reductive workup of the ozonide with
sodium borohydride led to a mixture of diol 19 and hemi-
aminal 20. The former compound was subjected to selective
oxidation of the benzylic alcohol by MnO2, generating the
intermediate aldehyde, which, after in situ cyclization, pro-
duced compound 20.
In our group, we have been investigating various truncated
derivatives of pancratistatin, and 7-deoxypancratistatin,12 as
well as derivatives with variations of functionality at the aro-
matic core,13 including an indole mimic of 7-deoxypancratis-
tatin.14 More recently, and inspired by Pettit’s report of the
C-1 benzoate ester,4c we investigated carbon homologues of
7-deoxypancratistatin at position C-1 and found that the hy-
droxymethyl and acetoxymethyl derivatives (5 and 6, respec-
tively) were reasonably active.3a 7-Deoxypancratistatin is far
less active than pancratistatin as it lacks the 7-OH functional-
ity and we reasoned that the C-1 homologues of 1 might ap-
proach the activity of the natural products. Indeed, this was
shown to be the case and homologues 7 and 8 showed nano-
molar activities in preliminary screening.15 In this paper, we
report the full details of the synthesis of these compounds,
the synthesis of the C-1 benzoxymethyl analogue, as well as
the complete details of the biological evaluation in several
cell lines.
Results and discussion
The strategy for the synthesis of C-1 homologues followed
the previously disclosed approach in which a C-1 aldehyde
possessing the full pancratistatin core is generated by the ox-
idative cleavage–recyclization in a phenanthrene–phenanthridol
transformation.16 To adjust this protocol to the synthesis of
the pancratistatin core, the hydroxy aldehyde 10, prepared
from piperonal by a known method,17 was chosen as the
starting material.
The primary alcohol in 20 was selectively acetylated by
stirring with acetic anhydride in pyridine–dichloromethane.
Several different oxidation conditions were attempted to
transform the hemiaminal moiety to the phenathridone amide,
including pyridinium dichromate (PDC), pyridinium chloro-
chromate (PCC), o-iodoxybenzoic acid (IBX), Dess–Martin,
and tetrapropylammonium perruthenate / N-methylmorpho-
line-N-oxide (TPAP/NMO). Of these, only the latter condi-
tions led to the desired product; all other conditions led to
the decomposition of the starting material. Detosylation with
Na/naphthalene in 1,2-dimethoxyethane (DME) provided
compound 23 with traces of alcohol 24 (Scheme 2).
As shown in Scheme 1, compound 10 was methylated to
1118 and converted via the Corey–Fuchs reaction to acetylene
13.
The alane generated from 13 was reacted with epoxyaziri-
dine 14,19 which was prepared in five steps from bromoben-
zene via enzymatic dihydroxylation, the Yamada aziridination
protocol, dehalogenation, and epoxidation. Addition of the ep-
oxyaziridine 14 to the alane gave acetylene 15, which was
subjected to reduction conditions. The best results for selec-
tive alkyne–alkene reduction were obtained using a short time
for the hydrogenation (1–1.5 h) under Lindlar’s conditions
with 1 atm (1 atm = 101.325 kPa) of hydrogen to provide the
cis-olefin 16. Different catalysts and conditions led to signifi-
The O-demethylation of 23 was performed by freshly
fused LiCl in dry dimethylformamide (DMF).20 Alternative
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Scheme 2.
21
protocols of demethylation such as BBr3 or sodium dodeca-
thiolate22 did not lead to product. In the first case, overdepro-
tection of the acetonide and the methylenedioxo groups was
observed. In the second case, the reaction did not proceed at
all. Desilylation led to compound 25, which can be selec-
tively deprotected to provide either acetoxymethyl derivative
8 or hydroxymethyl analogue 7 (Scheme 3).
ent. For example, both 7 and 8 were about 10 times more
potent than their 7-deoxy analogues (5 and 6) against the
prostate DU-145 cells, and this 10-fold difference in activity
is similar to that between 1 and 3.2a,13b In addition, the dou-
ble-digit nanomolar potency of the acetoxymethyl analogue
(8) is noteworthy and it approaches that of narciclasine,
which was used as a reference compound. The most encour-
aging activity was found, however, with the benzoxymethyl
compound (9). It is three to five times more potent than nar-
ciclasine against the pancreatic BXPC-3 and prostate DU-145
cancer cells, while showing a similar magnitude of effect to-
ward the other two cell lines used. This finding is consistent
with the previous observations of the beneficial effects im-
parted by large hydrophobic C-1 substituents as was reported
by Pettit et al.4c for the C1-benzoate derivative of pancratis-
tatin. The benzoate moiety could be part of the cytotoxic
pharmacophore or it could merely assist the parent hydroxyl
compound in cell penetration and then undergo intracellular
hydrolytic removal to its hydroxy analogue (7). Further
studies are required to provide a satisfactory answer to this
important question.
To provide access to the benzoate derivative 9, a point of
diversion in synthesis was chosen to be compound 23, which
possesses all the required functionality and is prone to basic
hydrolysis. Benzoylation of alcohol 24 provided access to
compound 27, which was subjected to the deprotection proto-
col to furnish the C-1 benzoxymethyl derivative 9 (Scheme 4).
Evaluation of biological activities
The hydroxymethyl, acetoxymethyl, and benzoxymethyl
analogues (7, 8, and 9, respectively), together with their 7-
deoxy counterparts (5 and 6), were evaluated in a small
panel of cancer cell lines diversely representing several
types of human malignancy (Table 1). As expected, the 7-
hydroxy compounds were found to be more potent, again
underscoring the beneficial effect of the 7-hydroxy substitu-
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Can. J. Chem. Vol. 90, 2012
Scheme 3.
Scheme 4.
As pancratistatin was previously found to be effective in
(Me2SO). Values are expressed as the mean SD from quad-
ruplicates of three independent experiments.
selectively producing cytotoxicity in an array of cancers, we
additionally evaluated the activity of 5–9 in the colorectal
cancer cell line HCT 116 and in the osteosarcoma cell line
Saos-2 with the water soluble tetrazolium salt (WST-1)-based
colorimetric assay for cell viability. After 48 h of treatment,
all of these pancratistatin analogues decreased cell viability
in a dose-dependent manner. The benzoate ester analogue (9)
demonstrated the greatest anticancer activity at lower concen-
trations and had half-maximal inhibitory concentration (IC50)
values below 0.05 µmol/L after 48 h in both HCT 116 and
Saos-2 cells (Figs. 2A and 2B, respectively); however, its ef-
fectiveness plateaus at 0.1 µmol/L, after which 7 becomes
more effective against HCT 116 cells and 8 becomes more
effective against Saos-2 cells.
The effect of pancratistatin analogues on cellular viability
of colorectal cancer and osteosarcoma cells was determined
by the WST-1-based colorimetric assay. HCT 116 and Saos-
2 cells (Figs. 2A and 2B, respectively) were treated with pan-
cratistatin analogues for 48 h and the WST-1 reagent was
used to quantify cell viability. Absorbance was read at
450 nm and expressed as a percentage of the control
Conclusions
Three new C-1 homologues of pancratistatin have been
synthesized and tested in various cancer cell lines. The re-
sults were compared with the two previously tested C-1 ho-
mologues of 7-deoxypancratistatin. The newly synthesized
C-1 homologues manifested nanomolar antiproliferative ac-
tivities against a panel of human cancer cell lines similar to
the parent natural products. The benzoxymethyl compound
(9), however, exhibited the most encouraging activity, which
was three to five times more potent than that of narciclasine
against the pancreatic BXPC-3 and prostate DU-145 cancer
cells. The various C-1 analogues screened in this study have
very high cytoxicity at low concentrations in a variety of ag-
gressive cancer cells. We observed remarkable enhancement
of cytotoxicity in compounds containing the hydroxyl group
at C-7 and the benzoate ester at C-1 in human colorectal can-
cer and osteosarcoma. These novel analogues may hold the
potential to be very effective anticancer therapeutic agents.
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Table 1. Activity of C-1 analogues with narciclasine as a standard (IC50, µmol/L).
Structure
Cancer
type
Cell line
Pancreatic
Prostate
Lung
BxPC-3
DU-145
NCI-H460
MCF-7
0.05 0.01
0.03 0.01
0.05 0.03
0.06 0.03
0.77 0.01
1.10 0.20
0.40 0.01
0.86 0.06
0.34 0.05
0.72 0.27
0.53 0.01
1.81 1.20
0.22 0.01
0.09 0.01
0.09 0.01
0.24 0.10
0.07 0.01
0.06 0.01
0.07 0.01
0.52 0.47
0.01 0.00
0.01 0.00
0.03 0.01
0.08 0.01
Breast
Note: The half-maximal inhibitory concentration (IC50) is the concentration required to reduce the viability of cells by 50%, after 48 h of treatment with
indicated compounds, relative to the dimethyl sulfoxide (DMSO) control; standard deviation (SD) from two independent experiments, each performed in
four replicates, determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.
noline from Zn. Qualitative thin-layer chromatography (TLC)
was done with precoated silica gel aluminum sheets (EMD
silica gel 60 F254) with detection by UV or by spraying with a
cerium ammonium molybdate (CAM) solution (5 g of
(NH4)6Mo7O24·4H2O, 1 g of Ce(SO4)2, and 100 mL of 10%
H2SO4) or a 0.5% aqueous KMnO4 solution followed by
heating. Melting points are uncorrected. Flash chromatogra-
phy was performed using silica gel SiliaFlash P60 from
Silicycle (40–66 µm). Optical rotation was measured in a 1
dm cell at 20–25 °C and 589 nm with a concentration (c)
in g/100 mL. IR spectra were recorded in KBr pellets or as
Fig. 2. Pancratistatin analogues cause cytotoxicity in cancer cells in
a dose-dependent manner.
1
a thin film. H and 13C NMR spectra were recorded at 300
or 600 MHz and 75 or 150 MHz, respectively, and were
calibrated on the solvent residual peak or tetramethylsilane
(TMS) signal (CDCl3, 7.28 ppm; DMSO-d6, 2.51 ppm); the
1
chemical shifts are reported in ppm. Copies of H and 13C
NMR spectra for compounds 7–9, 12, 13, and 15–29) are
available in the Supplementary data.
4-Methoxy-1,3-benzodioxole-5-carbaldehyde (11)
Dimethyl sulfate (22.7 mL, 240 mmol) was added to a
mixture of K2CO3 (55.2 g, 400 mmol) and phenol 10 (made
from piperonal by literature procedure;17 33.2 g, 200 mmol)
in acetone (260 mL). The reaction was stirred under reflux
until consumption of the starting material was observed
(TLC, approximately 4 h). Then the reaction mixture was
cooled and inorganic salts were removed by filtration and
rinsed with acetone (2 × 100 mL). The solution was evapo-
rated, redissolved in CH2Cl2, and washed sequentially with a
10% solution of NaOH, water, and saturated solution of NaCl.
The organic solution was then dried over anhydrous Na2SO4
and evaporated to obtain compound 11 (30.2 g, 84%) as pale
brown crystals, which was used without further purification;
mp 102–104 °C (EtOH) (lit.18 mp 103–105 °C (EtOH)). Rf =
1
0.65 (hexanes/EtOAc, 9:1). H NMR (CDCl3, 300 MHz) d:
Experimental section
10.24 (s, 1H), 7.49 (d, J = 8.3 Hz, 1H), 6.62 (d, J =
8.3 Hz, 1H), 6.05 (s, 2H), 4.14 (s, 3H).
General
Reactions were carried out under inert atmosphere in oven-
dried glassware unless stated otherwise. LiCl was fused
under vacuum immediately before use. Solvents were dis-
tilled: CH2Cl2, DMF, i-Pr2NEt, and pyridine from CaH2,
MeOH from magnesium methoxide, tetrahydrofuran (THF)
and DME from Na/benzophenone, toluene from Na, and qui-
5-(2,2-Dibromovinyl)-4-methoxy-1,3-benzodioxole (12)
Triphenylphosphine (64.0 g, 244 mmol) in CH2Cl2
(100 mL) was added dropwise to a stirring solution of CBr4
(40.5 g, 122 mmol) in CH2Cl2 (150 mL) at 0 °C (ice bath).
After 15 min of stirring, a solution of aldehyde (11; 11.0 g,
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Can. J. Chem. Vol. 90, 2012
61.0 mmol) in CH2Cl2 (50 mL) was added dropwise. Upon
completion, the reaction was reduced in volume to 100 mL
and slowly poured into vigorously stirred hexanes
(1400 mL). The mixture was then filtered through a short
plug of silica, washed with a mixture of hexanes/EtOAc
(10:1, 200 mL), and evaporated. Subjection of this material
to flash column chromatography (eluent hexanes/EtOAc,
9:1) and concentration of the relevant fractions gave 12
(16.11 g, 78.6%) as a white solid; mp 38–40 °C (pentane).
Rf = 0.9 (hexanes/EtOAc, 9:1). IR (KBr, cm–1) n: 3448,
2981, 2948, 2934, 2900, 2876, 2838, 2770, 1625, 1605,
1471, 1427, 1384, 1350, 1265, 1213, 1126, 1072, 1045,
(20 mL) was added dropwise. The reaction mixture was
stirred for 1 h and was allowed to warm to room tempera-
ture overnight. The reaction mixture was cooled to 0 °C by
ice bath and quenched with 1 N HCl (1 mL), followed by
ice-cold water (1 mL) and 1 N HCl (2 mL). The reaction
mixture was filtered through a plug of Celite and extracted
with EtOAc (3 × 20 mL). The combined organic phases
were dried over Na2SO4, filtered, and evaporated. The crude
product was purified by column chromatography (gradient
hexanes/EtOAc 5:1 to 3:1) to give the product as colourless
oil (1.02 g, 44%). Repetition of this procedure on a 4.12 g
24
scale of 13 provided 15 in a 38% yield. ½aꢀD +78.4 (c 1.0,
1
979, 960, 939, 929, 848, 829, 788, 767, 729, 644. H NMR
CHCl3). Rf = 0.3 (hexanes/EtOAc, 2:1). IR (film, cm–1) n:
3482, 3093, 2986, 2935, 2900, 1620, 1599, 1470, 1434,
1404, 1383, 1332, 1307, 1260, 1225, 1186, 1183, 1071,
(CDCl3, 300 MHz) d: 7.49 (s, 1H), 7.24 (d, J = 8.3 Hz, 1H),
6.57 (d, J = 8.3 Hz, 1H), 5.96 (s, 2H), 4.02 (s, 3H). 13C
NMR (CDCl3, 75 MHz) d: 149.7, 141.1, 136.1, 132.3,
122.5, 121.4, 102.3, 101.2, 89.0, 59.9. MS (+EI) m/z (%):
338 ([81Br + 81Br, M]+, 49), 336 ([81Br + 79Br, M]+, 100),
334 ([79Br + 79Br, M+], 51), 242 (55), 240 (57), 176 (53),
175 (42), 131 (29). HR-MS (+EI) calcd for C10H8Br2O3:
333.8820; found: 333.8845. Anal. calcd for C10H8Br2O3: C
35.75, H 2.40; found: C 35.99, H 2.41.
1
985. H NMR (CDCl3, 300 MHz) d: 7.83 (d, J = 8.3 Hz,
2H), 7.39 (d, J = 8.3 Hz, 2H), 6.88 (d, J = 8.1 Hz, 1H),
6.47 (d, J = 8.3 Hz, 1H), 5.96 (s, 2H), 4.48 (d, J =
6.2 Hz, 1H), 4.19 (t, J = 5.7 Hz, 1H), 4.06 (s, 3H), 3.98–
3.95 (m, 1H), 3.44–3.41 (m, 1H), 3.26 (d, J = 6.8 Hz, 1H),
3.24 (dd, J = 5.0, 2.1 Hz, 1H), 3.08 (d, J = 8.7 Hz, 1H),
2.48 (s, 3H), 1.51 (s, 3H), 1.34 (s, 3H). 13C NMR (CDCl3,
75 MHz) d: 149.7, 145.4, 144.4, 136.3, 134.1, 130.1, 127.9,
127.2, 110.1, 108.8, 102.8, 101.3, 87.6, 80.5, 75.5, 70.3,
69.0, 60.0, 42.3, 40.2, 31.9, 27.3, 25.1, 21.7. MS (+FAB)
m/z (%): 514 ([M + H]+, 24), 513 ([M]+, 13), 258 (12),
238 (11), 230 (14), 179 (26), 155 (28), 149 (23), 43 (100).
HR-MS (+FAB) calcd. for C26H28NO8S+ [M + 1]+:
514.1457; found: 514.1502.
5-Ethynyl-4-methoxy-1,3-benzodioxole (13)
To a solution of 12 (19.38 g, 57.68 mmol) in THF
(350 mL) was added a solution of n-BuLi (52.9 mL,
2.5 mol/L, and 130 mmol) at –78 °C. After 20 min of stir-
ring at –78 °C, the reaction mixture was warmed to room
temperature over a period of 2 h. A saturated solution of
NH4Cl (40 mL) was poured into the reaction mixture, which
was later extracted by CH2Cl2 (3 × 50 mL). The combined
organic phases were dried over Na2SO4, filtered, and evapo-
rated. The crude compound was purified by flash chromatog-
raphy (hexanes/EtOAc 2:1) and white crystals of acetylene
13 were obtained (8.5 g, 81.7%); mp 77–78 °C (pentane). Rf =
0.9 (hexanes/EtOAc, 2:1). IR (KBr, cm–1) n: 3278, 3254,
3000, 2945, 2901, 2846, 2794, 2097 (weak), 1620, 1600,
1469, 1433, 1336, 1267, 1229, 1077, 1043, 979, 950, 930,
(3aS,4R,5R,6R,7S,7aR)-6-[(Z)-2-(4-Methoxy-1,3-
benzodioxol-5-yl)vinyl]-2,2-dimethyl-8-[(4-methylphenyl)
sulfonyl]hexahydro-4,5-epimino-1,3-benzodioxol-7-ol (16)
To a solution of compound 15 (427 mg, 0.83 mmol) in
MeOH (20 mL) was added quinoline (11 mg, 0.09 mmol).
The solution was charged with a Lindlar catalyst (5%,
100 mg) and allowed to stir under H2 (1 atm) for 45 min.
After consumption of the starting material was observed
(NMR), the reaction mixture was filtered through a pad of
Celite, washed with CH3OH (3 × 30 mL), evaporated, and
used without further purification. An analytical sample was
purified by column chromatography (hexanes/EtOAc, 3:1) to
1
797. H NMR (CDCl3, 300 MHz) d: 7.01 (d, J = 7.9 Hz,
1H), 6.50 (d, J = 8.29 Hz, 1H), 5.98 (s, 2H), 4.11 (s, 3H),
3.20 (s, 1H). 13C NMR (CDCl3, 75 MHz) d: 150.1, 144.8,
136.2, 128.0, 108.1, 102.9, 101.3, 79.9, 79.4, 60.0. MS
(+EI) m/z (%): 176 ([M]+, 100), 175 (29), 131 (16), 53
(18). HR-MS (+EI) calcd for C10H8O3: 176.0473; found:
176.0475. Anal. calcd for C10H8O3: C 68.18, H 4.58;
found: C 68.27, H 4.55.
20
provide 16 as a waxy solid. ½aꢀD +2.7 (c 1.78, CHCl3). Rf =
0.4 (hexanes/ EtOAc, 2:1). IR (film, cm–1) n: 3482, 2987,
2934, 2900, 1621, 1598, 1470, 1434, 1404, 1382, 1332,
1
1260, 1218, 1162, 1070, 1043. H NMR (CDCl3, 300 MHz)
d: 7.74 (d, J = 8.3 Hz, 2H), 7.32 (d, J = 8.3 Hz, 2H), 6.60
(d, J = 11.4 Hz, 1H), 6.55 (d, J = 8.1 Hz, 1H), 6.46 (d, J =
8.0 Hz, 1H), 5.98 (d, J = 1.2 Hz, 1H), 5.96 (d, J = 1.2 Hz,
1H), 5.73 (t, J = 11.2 Hz, 1H), 4.43 (d, J = 6.2 Hz, 1H),
4.16–4.13 (m, 1H), 3.96 (s, 3H), 3.72–3.68 (m, 1H), 3.18 (d,
J = 6.4 Hz, 1H), 3.15–3.10 (m, 1H), 3.08 (d, J = 6.4 Hz,
1H), 2.84 (d, J = 9.2 Hz, 2H), 2.43 (s, 3H), 1.49 (s, 3H),
1.30 (s, 3H). 13C NMR (CDCl3, 75 MHz) d: 148.9, 145.2,
141.3, 136.6, 134.3, 130.0, 128.1, 127.9, 127.8, 122.5,
122.2, 109.7, 102.6, 101.0, 75.8, 70.1, 69.3, 59.7, 43.1,
40.5, 37.8, 27.1, 24.7, 21.7. MS (+FAB) m/z (%) [M]+: 517
(16), 516 (42), 515 (29), 514 (15), 386 (25), 285 (14), 284
(15), 269 (15), 203 (30), 165 (61), 91 (100). HR-MS
(3aS,4R,5R,6R,7S,7aR)-6-[(4-Methoxy-1,3-benzodioxol-5-
yl)ethynyl]-2,2-dimethyl-8-[(4-methylphenyl)sulfonyl]
hexahydro-4,5-epimino-1,3-benzodioxol-7-ol (15)
To a solution of alkyne 13 (1.580 g, 8.95 mmol) in toluene
(30 mL) at –50 °C, n-BuLi (3.80 mL, 2.35 mol/L, and
8.95 mmol) was added dropwise. After 15 min of stirring,
Me2AlCl (9.0 mL, 1.0 mol/L, and 8.95 mmol) was added
dropwise. The reaction mixture was warmed to 0 °C within
1 h and stirred for an additional 40 min at 0 °C. The reaction
mixture was allowed to warm to room temperature and stirred
for 40 min. The reaction mixture was cooled to –30 °C and a
solution of epoxide 14 (1.510 g, 4.47 mmol) in toluene
Published by NRC Research Press

Pagination not final/Pagination non finale
Vshyvenko et al.
7
(+FAB) calcd for C26H31NO8S [M + 1]+: 516.1692; found:
516.1666. Anal. calcd for C26H29NO8S: C 60.57, H 5.67;
found: C 60.33, H 5.55.
9.8, 1.5 Hz, 1H), 4.59 (d, J = 8.8 Hz, 1H), 4.28 (m, 1H),
4.11 (s, 1H), 4.02–3.99 (m, 1H), 3.97 (s, 3H), 3.80–3.70 (m,
1H), 2.79–2.78 (m, 1H), 2.61–2.56 (m, 1H), 2.41 (s, 3H),
1.73 (s, 1H), 1.45 (s, 3H), 1.34 (s, 3H), 0.88 (s, 9H), 0.10
(s, 3H), 0.05 (s, 3H). 13C NMR (CDCl3, 75 MHz) d: 147.5,
142.1, 139.5, 138.6, 135.5, 129.4, 129.8, 126.8, 125.3,120.5,
119.7,109.2,104.8,100.7,79.1,78.3,70.3,59.7,53.7,42.4,39.1,
27.8,26.3,25.7,21.5,18.0,–5.03,–5.056. MS (+FAB) m/z (%)
[M]+: 629 (3), 129 (13), 111 (12), 99 (13), 57 (100). HR-
MS (+FAB) calcd for C32H43NO8SSi [M]+: 629.2479;
found: 629.2472.
(3aS,4R,5R,6R,7S,7aR)-7-{Tert-butyl[dimethylsylil]oxy}-6-
[(Z)-2-(4-methoxy-1,3-benzodioxol-5-yl)vinyl]-2,2-
dimethyl-8-[(4-methylphenyl)sulfonyl]hexahydro-4,5-
epimino-1,3-benzodioxol (17)
To a solution of alcohol 16 (1.07 g, 2.07 mmol) in 30 mL
of CH2Cl2 was added Et3N (0.58 mL, 4.15 mmol) at 0 °C
and tert-butyldimethylsilyl triflate (0.53 mL, 2.29 mmol)
was added dropwise. After the complete consumption of the
starting material was observed (TLC), the reaction mixture
was quenched by water (10.0 mL) and extracted with
CH2Cl2 (3 × 20 mL). The combined organic phases were
washed with 10% citric acid (5 mL), brine (5 mL), dried
over Na2SO4, and concentrated to afford 17 as pale yellow
oil (1.26 g, 97%). The compound was used without further
purification. An analytical sample was purified by silica gel
N-[(3aS,4R,5R,6S,7S,7aS)-7-{[Tert-butyl(dimethyl)silyl]
oxy}-6,6′-bis(hydroxymethyl)-7′-methoxy-2,2-dimethyl-
3,4,5,6,7,7a-hexahydro-5,5′-bi-1,3-benzodioxol-4-yl]-4-
methylbenzenesulfonamide (19)
To a solution of 18 (254 mg, 0.404 mmol) in MeOH
(50 mL) was added a few crystals of Sudan Red 7B. The sol-
ution was cooled down to –80 °C and a oxygen–ozone mix-
ture was bubbled through until the disappearance of the pink
color. The consumption of the starting material was also
checked by TLC. A stream of nitrogen was bubbled through
the reaction mixture for 5 min. NaBH4 (250 mg, 6.67 mmol)
was slowly added and reaction mixture was gradually
warmed from –80 °C to room temperature. The solvent was
removed in vacuo and the residue was redissolved in CH2Cl2
(50 mL), neutralized by 10% citric acid, and washed with
water (50 mL). The organic phase was separated, dried over
Na2SO4, filtered, and evaporated. The crude product was sub-
jected to column chromatography (hexanes/EtOAc, 1:1) to
give 19 as a white crystalline solid (164.3 mg, 61%) and 20
as a mixture of anomers (80 mg, 30%); mp 121–123 °C
24
chromatography (hexanes/EtOAc, 4:1). ½aꢀD –18.5 (c 2.0,
CHCl3). Rf = 0.85 (hexanes/EtOAc, 2:1). IR (KBr, cm–1) n:
3446, 2986, 2954, 2931, 2887, 2856, 1622, 1600, 1470,
1
1435, 1382, 1332, 1257, 1218, 1163, 1071, 1043. H NMR
(CDCl3, 300 MHz) d: 7.78 (d, J = 8.2 Hz, 2H), 7.30 (d, J =
8.2 Hz, 2H), 6.67 (d, J = 8.0 Hz, 1H), 6.60 (d, J = 11.5 Hz,
1H), 6.39 (d, J = 8.0 Hz, 1H), 5.96–5.93 (m, 2H), 5.65 (t,
J = 11.5 Hz, 1H), 4.39 (d, J = 5.9 Hz, 1H), 3.98 (s, 3H),
3.89 (t, J = 6.03 Hz, 1H), 3.67 (t, J = 6.3 Hz, 1H), 3.12 (d,
J = 6.6 Hz, 1H), 2.98–2.91 (m, 2H), 2.45 (s, 3H), 1.52 (s,
3H), 1.34 (s, 3H), 0.78 (s, 9H), 0.00 (s, 3H), –0.08 (s, 3H).
13C NMR (CDCl3, 75 MHz) d: 148.7, 144.4, 141.4, 136.3,
135.0, 129.7, 129.6, 127.8, 127.6, 122.9, 122.4, 109.3,
102.3, 100.8, 71.8, 71.4, 59.6, 43.4, 39.7, 39.2, 27.7, 25.7,
25.5, 21.6, 18.0, –4.6, –4.8. MS (+FAB) m/z (%) [M]+: 628
(8), 514 (13), 343 (17), 256 (10), 228 (10), 215 (19), 165
(36), 73(100). HR-MS (+FAB) calcd for C32H44NO8SSi
[M + 1]+: 630.2557; found: 630.2492. Anal. calcd for
C32H43NO8SSi: C 61.02, H 6.88; found: C 61.28, H 7.02.
20
(CHCl3). ½aꢀD –30.8 (c 1.09, CHCl3). Rf = 0.45 (hexanes/
EtOAc, 1:1). IR (KBr, cm–1) n: 3472, 3386, 3172, 2927,
2855, 1622, 1482, 1385, 1332, 1255, 1220, 1158, 1095,
1057, 837. 1H NMR (CDCl3, 600 MHz) d: 7.51 (d, J =
7.8 Hz, 2H), 7.13 (d, J = 7.8 Hz, 2H), 6.57 (s, 1H), 5.96 (s,
1H), 5.90 (s, 1H), 5.44 (d, J = 7.0 Hz, 1H), 4.77 (d, J =
11.8 Hz, 1H), 4.42 (d, J = 11.9 Hz, 1H), 4.27–4.24 (m,
1H), 4.17–4.10 (m, 2H), 4.00 (s, 3H), 3.95–3.86 (m, 1H),
3.70 (dd, J = 11.9, 6.1 Hz, 1H), 3.59 (dd, J = 11.8, 6.3 Hz,
1H), 3.37 (dd, J = 11.8, 3.9 Hz, 1H), 2.92 (br s, 2H), 2.39
(s, 3H), 2.00–1.96 (m, 1H), 1.56 (s, 3H), 1.32 (s, 3H), 0.92
(s, 9H), 0.14 (s, 3H), 0.11 (s, 3H). 13C NMR (CDCl3, 150
MHz) d: 149.1, 142.1, 141.6, 139.2, 135.1, 132.4, 128.8,
126.8, 124.6, 109.7, 103.5, 101.0, 79.9, 79.0, 71.5, 61.3,
60.0, 57.0, 55.2, 47.3, 38.2, 27.4, 25.9, 25.8, 21.5, 18.0,
21.6, 21.0, 18.0, –4.8, –5.0. MS (+FAB) m/z (%): 664 ([M –
H]+, 6), 648 (7), 372 (11), 302 (11), 254 (21), 248 (12), 73
(100). HR-MS (+EI) calcd for C32H47NO10SSi [M]+:
665.2690; found: 665.2803. Anal. calcd for C32H47NO10SSi:
C 57.72, H 7.11; found: C 57.76, H 6.99.
N-((1R,2S,3S,4S,4aR,11bR)-4-{[Tert-butyl(dimethyl)silyl]
oxy}-3,3-dimethoxy-7-methoxy-1,2,3,4,4a,11b-
hexahydrophenanthro[2,3-d][1,3]dioxol-1-yl)
benzenesulfonamide (18)
Olefin 17 (100 mg, 0.561 mmol), quinoline (15 mg,
0.12 mmol), and silica gel (500 mg), which has been acti-
vated in advance by heating under vacuum for 24 h at 150 °C,
were suspended in CH2Cl2 (10 mL). The solvent was re-
moved in vacuo and the flask containing silica gel support-
ing the absorbed reactants was heated at 120 °C under a
nitrogen atmosphere and stirred for 36 h. The reaction mix-
ture was then separated by column chromatography (hex-
anes/EtOAc, 4:1) to give 74 mg (74%) of olefin 18 as a
(3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-11-(hydroxymethyl)-6-methoxy-2,2-dimethyl-4-
[(4-methylphenyl)sulfonyl]-3,3b,4,5,10b,11,12,12a-
octahydrobis[1,3]dioxolo[4,5-c:4′,5′-j]phenanthridin-5-ol
(20)
24
clear and colourless oil. ½aꢀD –25.1 (c 1.0, CHCl3). Rf =
0.45 (hexanes/EtOAc, 2:1). IR (KBr, cm–1) n: 3275, 2983,
2953, 2932, 2889, 2857, 1633, 1614, 1599, 1479, 1384,
1
1361, 1331, 1221, 1158, 1092, 841. H NMR (CDCl3, 300
To a solution of alcohol 19 (100 mg, 0.15 mmol) in
CH2Cl2 (100 mL) was added MnO2 (268 mg, 3 mmol). The
reaction mixture was vigorously stirred until total consump-
MHz) d: 7.43 (d, J = 8.1 Hz, 2H), 7.13 (d, J = 8.1 Hz,
2H), 6.65 (dd, J = 9.8, 3.4 Hz, 1H), 6.22 (s, 1H), 5.92 (d,
J = 1.5 Hz, 1H), 5.82 (d, J = 1.5 Hz, 1H), 5.74 (dd, J =
Published by NRC Research Press

Pagination not final/Pagination non finale
8
Can. J. Chem. Vol. 90, 2012
tion of the starting material was observed (TLC). The reac-
tion mixture was filtered through a plug of Celite® and
washed with CH2Cl2 (3 × 100 mL). The solvent was re-
moved in vacuo, affording 20 as a white solid compound
(87 mg, 87%, mixture of anomers); mp 106–116 °C
(CHCl3). Rf = 0.5 and 0.65 (hexanes/EtOAc, 1:1). IR (KBr,
cm–1) n: 3452, 2985, 2954, 2931, 2894, 2857, 1624, 1483,
1384, 1341, 1251, 1163, 1077, 839. NMR of the major
anomer: 1H NMR (CDCl3, 300 MHz) d: 7.55 (d, J =
8.1 Hz, 2H), 7.03 (d, J = 8.1 Hz, 2H), 6.65 (s, 1H), 6.08 (s,
1H), 5.92–5.91 (m, 2H), 5.26 (dd, J = 9.9, 5.4 Hz, 1H), 4.38
(t, J = 3.0 Hz, 1H) 4.25–4.21 (m, 2H), 4.11 (s, 3H), 3.74
(dd, J = 11.0, 7.9 Hz, 1H), 3.37 (dd, J = 11.0, 3.6 Hz, 1H),
2.87 (dd, J = 12.9, 5.1 Hz, 1H), 2.33 (s, 3H), 2.16 (br s,
1H), 2.15 (br s, 1H), 2.06 (s, 1H), 1.44 (s, 3H), 1.37 (s,
3H), 0.92 (s, 9H), 0.15 (s, 3H), 0.10 (s, 3H). 13C NMR
(CDCl3, 150 MHz) d: 150.0, 142.9, 140.5, 138.3, 134.0,
130.0, 128.9, 127.2, 121.3, 109.4, 101.0, 100.2, 78.9, 76.4,
73.5, 68.1, 59.9, 59.3, 53.1, 48.1, 34.4, 27.9, 26.2, 25.7,
21.4, 17.9, –4.8, –5.1. MS (+EI) m/z (%) [M – Ts – H2O]+:
491 (15), 432 (13), 302 (22.2), 302 (10.5), 247 (31.1), 246
(19.9), 43 (100). HR-MS (+EI) calcd for C32H45NO10SSi
[M]+: 663.2533; found: 663.2549. Anal. calcd for
C32H45NO10SSi: C 57.90, H 6.83; found: C 58.02, H 7.03.
found: 688.2642. Anal. calcd for C34H47NO11SSi: C 57.85,
H 6.71; found: C 57.80, H 6.67.
{(3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-6-methoxy-2,2-dimethyl-4-[(4-methylphenyl)
sulfonyl]-5-oxo-3a,3b,4,5,10b,11,12,12a-octahydrobis[1,3]
dioxolo[4,5-c:4′,5′-j]phenanthridin-11-yl}methyl acetate
(22)
A predried (by MgSO4) solution of N-methylmorpholine-
N-oxide (100 mg, 0.85 mmol) in CH2Cl2 (20 mL) was added
to hemiaminal 21 (30 mg, 0.042 mmol) in CH2Cl2 (10 mL),
followed by activated crushed molecular sieves (0.5 g, 4 Å).
After stirring for 15 min, a few crystals of tetrapropylammo-
nium perruthenate were added and the reaction was stirred
until consumption of the starting material was observed
(TLC). The reaction mixture was filtered through a plug of
Celite and washed with CH2Cl2 (3 × 50 mL). The combined
organic phases were evaporated and subjected to column
chromatography (hexanes/EtOAc, 2:1) affording 22 as a col-
24
ourless oil (25 mg, 84%). ½aꢀD +41.4 (c 0.9, CHCl3). Rf =
0.65 (eluent hexanes/EtOAc, 1:1). IR (KBr, cm–1) n: 3450,
2984, 2953, 2930, 2857, 1742, 1710, 1614, 1484, 1360,
1
1253, 1169, 1090, 1021, 839. H NMR (CDCl3, 300 MHz)
d: 8.20 (d, J = 8.3 Hz, 2H), 7.29 (d, J = 8.3 Hz, 2H), 6.73
(s, 1H), 6.06 (s, 1H), 6.02 (s, 1H), 4.83 (dd, J = 7.9, 5.5 Hz,
1H), 4.46 (s, 1H), 4.26 (t, J = 11.1 Hz, 1H), 4.17–4.13 (m,
1H), 4.12–4.08 (m, 1H), 4.04 (s, 3H), 4.00–3.95 (m, 1H),
3.47 (dd, J = 13.1, 3.3 Hz, 1H), 2.66–2.63 (m, 1H), 2.43 (s,
3H), 2.08 (s, 1H), 1.48 (s, 3H), 1.33 (s, 3H), 0.89 (s, 9H),
0.15 (s, 3H), 0.13 (s, 3H). 13C NMR (CDCl3, 150 MHz) d:
170.8, 163.7, 153.4, 144.8, 143.4, 138.8, 136.7, 136.1,
128.9, 128.7, 126.8, 117.8, 109.1, 102.0, 100.1, 79.0, 76.3,
66.9, 60.9, 60.7, 60.4, 59.7, 43.4, 36.5, 27.9, 26.1, 25.7,
21.6, 20.9, 18.0, –4.9, –5.1. MS (+EI) m/z (%): 639 (3.1),
549 (6.6), 492 (3.4), 434 (2.9), 374 (3.9), 43 (100). HR-MS
(+EI) calcd for C33H42NO11SSi+ [M – CH3]+: 688.2248;
found: 688.2436.
{(3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-5-hydroxy-6-methoxy-2,2-dimethyl-4-[(4-
methylphenyl)sulfonyl]-3,3b,4,5,10b,11,12,12a-
octahydrobis[1,3]dioxolo[4,5-c:4′,5′-j]phenanthridin-11-yl}
methyl acetate (21)
Pyridine (50 mg, 0.63 mmol) was added to a solution of
hemiaminal 20 (48 mg, 0.072 mmol) in CH2Cl2 (3 mL), fol-
lowed by the addition of acetic anhydride (29.5 mg,
0.29 mmol). The reaction mixture was stirred until consump-
tion of the starting material was observed (TLC). The reac-
tion mixture was quenched with water (5 mL) and extracted
by CH2Cl2 (3 × 4 mL). The combined organic phases were
dried over Na2SO4, filtered, and evaporated. The crude prod-
uct was subjected to column chromatography (eluent hexanes/
EtOAc, 4:1) affording 21 as a colourless oil (43 mg, 87%
mixture of anomers). Rf = 0.9 and 0.8 (hexanes/EtOAc,
1:1). IR (KBr, cm–1) n: 3462, 2984, 2953, 2931, 2896,
2857, 1743, 1624, 1481, 1371, 1342, 1330, 1250, 1222,
1164, 1077, 840. NMR of the major anomer: 1H NMR
(CDCl3, 600 MHz) d: 7.56 (d, J = 8.2 Hz, 2H), 7.08 (d, J =
8.0 Hz, 2H), 6.66 (s, 1H), 6.012 (s, 1H), 5.94 (m, 2H),
5.31 (dd, J = 10.0, 5.1 Hz, 1H), 4.37 (m, 1H) 4.22–4.21
(m, 1H), 4.08 (s, 3H), 3.96 (s, 1H), 3.91 (dd, J = 11.4,
4.4 Hz, 1H), 3.05 (s, 1H), 2.91 (dd, J = 13.2, 4.5 Hz,
1H), 2.35 (s, 3H), 2.15 (s, 1H), 1.93 (s, 3H), 1.47 (s, 3H),
1.38 (s, 3H), 0.93 (s, 9H), 0.15 (s, 3H), 0.11 (s, 3H). 13C
NMR (CDCl3, 150 MHz) d: 170.8, 150.0, 143.0, 140.6,
138.5, 134.1, 129.8, 128.9, 127.2, 121.3, 109.3, 101.0,
100.1, 78.9, 76.3, 73.3, 67.2, 61.2, 59.9, 53.2, 45.6, 33.8,
29.7, 26.3, 25.7, 25.5, 21.4, 20.8, 17.9, –5.0, –5.2. MS
(+FAB) m/z (%) [M – H2O]+: 688 (47), 230 (12), 117
(17), 302 (11), 247 (31), 117 (17), 73 (100). HR-MS
(+FAB) calcd for C34H46NO10SSi [M – H2O]+: 688.2612;
((3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-6-methoxy-2,2-dimethyl-5-oxo-
3,3b,4,5,10b,11,12,12a-octahydrobis[1,3]dioxolo[4,5-
c:4′,5′-j]phenanthridin-11-yl)methyl acetate (23)
To a solution of 22 (64 mg, 0.09 mmol) in dry DME
(7 mL) at –50 °C was added a solution of Na/naphthalene in
DME (0.5 mol/L) dropwise until a light green colour persisted
and total consumption of the starting material was observed
(TLC). The solution was stirred for 10 min before the reac-
tion was quenched with a saturated solution of NH4Cl (aq,
1 mL). The reaction was warmed to room temperature and
extracted with CH2Cl2 (6 × 15 mL). The combined organic
phases were dried over Na2SO4, filtered, and concentrated.
The products 23 and 24 were isolated by column chroma-
tography (gradient hexanes / EtOAc, 2:1 to 1:1) as a clear
and colourless oil for 23 (30 mg, 62%) and a white crystal-
line solid for 24 (2 mg; 5%). Repetition of this procedure
on a 1.01 g gram scale provided 23 in a 48% and 24 in a
20
10% yield. ½aꢀD +32.4 (c 1.0, CHCl3). Rf = 0.28 (hexanes/
EtOAc, 1:1). IR (KBr, cm–1) n: 3417, 3228, 3109, 2987,
2953, 2932, 2897, 2858, 1743, 1676, 1617, 1481, 1385,
1366, 1339, 1250, 1222, 1169, 1088, 1071, 1057, 1033,
1
840. H NMR (CDCl3, 600 MHz) d: 6.69 (s, 1H), 6.06 (s,
Published by NRC Research Press

Pagination not final/Pagination non finale
Vshyvenko et al.
9
1 H), 6.01 (s, 1H), 5.92 (s, 1H), 4.57 (s, 1H), 4.23 (d, J =
7.5 Hz, 1H), 4.19–4.18 (m, 1H), 4.16–4.14 (m, 1H), 4.07
(s, 3H), 3.42 (dd, J = 13.8, 8.3 Hz, 1H), 3.31 (dd, J =
13.8, 3.6 Hz, 1H), 2.66–2.65 (m, 1H), 2.11 (s, 3H), 1.45
(s, 3H), 1.39 (s, 3H), 0.91 (s, 9H), 0.16 (s, 3H), 0.15 (s,
3H). 13C NMR (CDCl3, 150 MHz) d: 170.9, 163.5, 152.4,
145.4, 137.5, 135.5, 116.2, 109.9, 101.8, 99.9, 78.3, 77.8,
67.0, 61.1, 60.9, 52.5, 41.9, 35.1, 28.2, 26.1, 25.69, 25.65,
20.9, 18.0, –5.0, –5.1. MS (+FAB) m/z (%): 552 (12), 551
(36), 550 ([M + 1]+, 100), 246 (10), 220 (11), 117 (101).
0.3 (hexanes / ethyl acetate, 1:1). IR (KBr, cm–1) n: 3449,
3270, 2988, 2911, 1743, 1672, 1625, 1600, 1466, 1443,
1
1357, 1307, 1245, 1231, 1166, 1087, 1071, 1032, 844. H
NMR (DMSO-d6, 600 MHz) d: 13.35 (s, 1H), 8.55 (s, 1H),
6.61 (s, 1H), 6.08 (s, 1H), 6.06 (s, 1H), 5.50 (d, J = 3.6 Hz,
1H), 4.34 (br s, 1H), 4.25 (d, J = 4.8 Hz, 1H), 4.20 (dd, J =
11.4, 3.6 Hz, 1H), 4.16–4.15 (m, 1H), 4.13 (d, J = 11.4 Hz,
1H), 3.51 (dd, J = 15.0, 8.4 Hz, 1H), 3.19 (dd, J = 14.4,
3.6 Hz, 1H), 2.03 (s, 3H), 1.41 (s, 3H), 1.31 (s, 3H). 13C
NMR (DMSO-d6, 150 MHz) d: 170.9, 169.8, 152.7, 146.3,
135.0, 132.6, 109.0, 107.7, 102.4, 97.6, 77.9, 76.9, 65.2,
61.2, 53.4, 34.3, 28.3, 26.4, 21.2. MS (+EI) m/z (%): 422
([M + 1], 24), 421 ([M+], 100), 248 (34), 247 (60), 232 (27),
231 (32), 218 (11), 206 (17), 145 (14). HR-MS calcd for
C20H23NO9 [M]+: 421.13728; found: 421.13792. Anal. calcd
for C20H23NO9: C 57.00, H 5.50; found: C 57.18, H 5.48.
HR-MS (+FAB) calcd for C27H40NO9Si [M
550.2472; found: 550.2459.
+
1]+:
((3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-6-hydroxy-2,2-dimethyl-5-oxo-
3,3b,4,5,10b,11,12,12a-octahydrobis[1,3]dioxolo[4,5-
c:4′,5′-j]phenanthridin-11-yl)methyl acetate (25)
[(1S,2S,3R,4S,4aR,11bR)-2,3,4,7-Tetrahydroxy-6-oxo-
1,2,3,4,4a,5,6,11b-octahydro[1,3]dioxolo[4,5-j]
phenanthridin-1-yl]methyl acetate (8)
To a solution of 23 (39 mg, 0.071 mmol) in DMF (5 mL)
was added LiCl (50 mg, 1.2 mmol) followed by three cycles
of freeze–pump–thaw. The reaction mixture was heated to
120 °C for 2.5 h. The reaction mixture was cooled to room
temperature, diluted with water (100 mL), and extracted with
diethyl ether (10 × 15 mL). The combined organic phases
were dried over Na2SO4, filtered, and concentrated. The
product was isolated by column chromatography (hexanes/
EtOAc, 3:1) as a clear and colourless oil that solidifies after
drying (25.7 mg, 68%). Repetition of this procedure on a
0.296 g gram scale provided 25 in a 67% yield; mp 68–69 °C
Compound 26 (38 mg, 0.09 mmol) was taken up in a
CH2Cl2–CH3OH mixture (1:1, 4 mL) and cooled to 0 °C.
Trifluoroacetic acid (2 mL) was added dropwise and the re-
action mixture was stirred until consumption of the starting
material was observed (TLC). The reaction mixture was dried
in vacuo, triturated with CH2Cl2 (3 × 15 mL) and finally
dried under high vacuum. The final product was isolated by
column chromatography on silica gel (deactivated by 10% w/w
of water, eluent CH2Cl2–CH3OH, 10:1) to give 8 as a white
20
(CHCl3). ½aꢀ –35.2 (c 1.0, CHCl3). Rf = 0.8 (hexanes/
EtOAc, 1:1).D IR (KBr, cm–1) n: 3402, 3348, 3285, 3212,
3087, 2987, 2953, 2933, 2899, 2859, 2795, 1743, 1627,
1601, 1464, 1389, 1366, 1353, 1341, 1301, 1250, 1226,
1171, 1081, 1032, 940, 838, 778. 1H NMR (CDCl3, 600
MHz) d: 12.68 (s, 1H), 6.53 (s, 1H), 6.26 (s, 1H), 6.07 (s,
2H), 4.57 (s, 1H), 4.30 (dd, J = 11.1, 3.3 Hz, 1H), 4.22 (t,
J = 11.0 Hz, 1H), 4.21–4.18 (m, 2H), 3.50 (dd, J = 14.4,
7.8 Hz, 1H), 3.36 (dd, J = 14.4, 3.7 Hz, 1H), 2.70–2.69
(m, 1H), 2.11 (s, 3H), 1.49 (s, 3H), 1.41 (s, 3H), 0.91 (s,
9H), 0.17 (s, 3H), 0.16 (s, 3H). 13C NMR (CDCl3, 150
MHz) d: 170.8, 169.9, 153.1, 146.9, 133.9, 133.0, 110.0,
107.3, 102.3, 97.3, 78.4, 77.6, 67.0, 61.10, 53.2, 41.7,
33.9, 28.3, 26.0, 25.7, 20.9, 18.0, –5.0, –5.1. MS (+EI) m/z
(%): 536 ([M + 1]+, 9), 535 (25), 360 (17), 256 (11), 231
(10), 218 (19), 205 (21), 149 (25), 43 (100). HR-MS (+EI)
calcd for C26H37NO9Si [M]+: 535.2238; found: 535.2248.
crystalline compound (31 mg, 90%); mp > 200 °C
24
(CH2Cl2–CH3OH). ½aꢀ
+36.8 (c 0.2, THF). Rf = 0.3
(CH2Cl2–CH3OH 10:1D). IR (KBr, cm–1) n: 3459, 3287,
3214, 2991, 2923, 1750, 1709, 1670, 1628, 1595, 1466,
1436, 1384, 1342, 1264, 1227, 1196, 1090, 1070, 1034.
1H NMR (DMSO-d6, 600 MHz) d: 13.26 (s, 1H), 7.40 (s,
1H), 6.59 (s, 1H), 6.08 (s, 1H), 6.06 (s, 1H), 5.17 (m,
1H), 5.10–5.09 (m, 2H), 4.40 (m, 1H), 4.16 (dd, J = 10.9,
3.5 Hz, 1H), 4.11 (s, 1H), 3.84 (m, 1H), 3.71 (m, 1H), 3.52
(dd, J = 13.6, 9.8 Hz, 1 H), 3.26 (m, 1H), 2.66 (m, 1H),
2.03 (s, 3H). 13C NMR (DMSO-d6, 150 MHz) d: 171.0,
169.9, 152.8, 146.3, 135.7, 132.5, 107.5, 102.4, 97.8, 73.1,
71.2, 68.9, 61.8, 51.6, 40.5, 36.5, 21.3. MS (+EI) m/z (%):
381 ([M]+, 12), 321 (16), 279 (12), 277 (99), 276 (11), 256
(12), 247 (11), 205 (13), 201 (16), 199 (12), 185 (13), 183
(11), 179 (21), 167 (14), 149 (37), 129 (12), 123 (11), 69
(100). HR-MS (+EI) calcd for C17H19NO9: 381.1060;
found: 381.1055.
[(3aS,3bR,10bR,11S,12S,12aR)-6,12-Dihydroxy-2,2-
dimethyl-5-oxo-3,3b,4,5,10b,11,12,12a-octahydrobis[1,3]
dioxolo[4,5-c:4′,5′-j]phenanthridin-11-yl]methylacetate(26)
Compound 25 (52 mg, 0.097 mmol) was taken up in THF
(2.5 mL) and cooled to 0 °C. A solution of tetrabutylammo-
nium fluoride (TBAF) in THF (1 mol/L, 0.107 mL,
0.107 mmol) was added dropwise over 1 min. The reaction
mixture was stirred until total consumption of the starting
material was observed (TLC). The reaction mixture was
quenched with water (5 mL) and extracted with CH2Cl2 (6 ×
15 mL). The combined organic phases were dried over
Na2SO4, filtered, and evaporated. The final product was iso-
lated by column chromatography (hexanes/EtOAc, 1:1) to
give 26 as a white crystalline solid (38 mg, 95%); mp >
(1S,2S,3R,4S,4aR,11bR)-2,3,4,7-Tetrahydroxy-1-
(hydroxymethyl)-1,3,4,4a,5,11b-hexahydro[1,3]dioxolo[4,5-
j]phenanthridin-6(2H)-one (7)
Compound 26 (48 mg, 0.11 mmol) was taken up in a
CH2Cl2–CH3OH mixture (1:1, 4 mL) and two drops of con-
centrated HCl were added. The reaction was stirred until total
consumption of the starting material was observed (TLC).
The reaction mixture was neutralized by the dropwise addi-
tion of a saturated solution of NaHCO3 and evaporated to
dryness. The final product was isolated by column chroma-
tography on silica gel (deactivated by 10% w/w of water, elu-
ent CH2Cl2–CH3OH, 5:1) to give 7 as a white crystalline
compound (36 mg, 92%); mp > 200 °C (CH2Cl2–CH3OH).
24
200 °C (CH2Cl2–CH3OH). ½aꢀD +6.2 (c 0.48, DMSO). Rf =
Published by NRC Research Press

Pagination not final/Pagination non finale
10
Can. J. Chem. Vol. 90, 2012
20
crystalline powder (94 mg, 64%); mp 90–92 °C (CHCl3).
½aꢀD +48.0 (c 0.5, abs DMSO). Rf = 0.4 (CH2Cl2–CH3OH
20
5:1). IR (KBr, cm–1) n: 3386, 2917, 1670, 1638, 1598, 1466,
1439, 1384, 1352, 1304, 1228, 1089, 1077, 1064, 1032. H
½aꢀD –15.1 (c 1, CHCl3). Rf = 0.4 (hexanes/EtOAc, 1:1). IR
(KBr, cm–1) n: 3467, 3416, 3387, 3069, 2986, 2952, 2932,
1
NMR (DMSO-d6, 300 MHz) d: 13.25 (s, 1H), 7.31 (s, 1H),
6.55 (s, 1H), 6.07 (s, 1H), 6.06 (s, 1H), 5.02 (m, 3H), 4.47
(dd, J = 6.4, 4.0 Hz, 1H), 4.18 (m, 1H), 3.91 (m, 1H),
3.82 (m, 1H), 3.67 (m, 1H), 3.43 (m, 1H), 3.14 (m, 1H),
2.37 (br. s, 1H). 13C NMR (DMSO-d6, 150 MHz) d: 169.9,
152.8, 146.1, 136.7, 132.2, 107.4, 102.4, 97.9, 73.2, 71.4,
69.5, 57.7, 51.8, 44.3, 36.9. MS (+EI) m/z (%): 339 [M]+:
(0.4), 85 (39), 84 (80), 83 (57), 68 (13), 66 (100). HR-MS
(+EI) calcd for C15H17NO8: 339.0954; found: 339.0925.
2896, 2857, 1722, 1674, 1616, 1502, 1481, 1453, 1385,
1
1339, 1273, 1221, 1093, 1071, 1028, 838. H NMR (CDCl3,
300 MHz) d: 8.06 (d, J = 7.2 Hz, 2H), 7.63–7.58 (m, 1H),
7.48 (t, J = 7.5 Hz, 2H), 6.74 (s, 1H), 6.01–6.02 (m, 3H),
4.68 (s, 1H), 4.53–4.45 (m, 2H), 4.20–4.16 (m, 2H), 4.07 (s,
3H), 3.53 (dd, J = 13.8, 7.5 Hz, 1H), 3.40–3.35 (m, 1H),
2.86–2.83 (m, 1H), 1.50 (s, 3H), 1.40 (s, 3H), 0.90 (s, 9H),
0.18 (s, 6H). 13C NMR (CDCl3, 75 MHz) d: 166.4, 163.6,
152.4, 145.4, 135.6, 133.2, 129.8, 129.6, 128.4, 116.1, 110.0,
101.7, 99.9, 78.3, 77.9, 67.2, 61.6, 60.9, 52.5, 42.1, 35.2, 28.2,
26.1, 25.6, 17.9, –4.9, –5.0. MS (+FAB) m/z (%): 614 ([M +
2]+, 12), 613 (36), 612 (89), 179 (12), 105 (100). HR-MS
calcd for C32H42NO9Si+ [M]+: 612.2251; found: 612.2653.
(3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-11-(hydroxymethyl)-6-methoxy-2,2-dimethyl-
3b,4,10b,11,12,12a-hexahydrobis[1,3]dioxolo[4,5-c:4′,5′-j]
phenanthridin-5(3aH)-one (24)
((3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-6-hydroxy-2,2-dimethyl-5-oxo-
3a,3b,4,5,10b,11,12,12a-octahydrobis[1,3]dioxolo[4,5-
c:4′,5′-j]phenanthridin-11-yl)methyl benzoate (28)
To a solution of 23 (170 mg, 0.309 mmol) in methanol
(5 mL) was added NaOH (aq 40%, 0.5 mL) and stirred until
total consumption of the starting material was observed
(TLC). The reaction was quenched with a saturated solution
of NH4Cl (1 mL). The mixture was evaporated and the resi-
due was extracted with CH2Cl2 (6 × 15 mL). The combined
organic phases were dried over Na2SO4, filtered, and concen-
trated. The final product was isolated by column chromatog-
raphy (gradient hexanes / EtOAc, 2:1 to 1:1) to give 24 as a
white crystalline solid (125 mg; 80%); mp 148–149 °C
To a solution of 27 (74.3 mg, 0.122 mmol) in dry DMF
(5 mL) was added LiCl (50 mg, 1.2 mmol) followed by three
cycles of freeze–pump–thaw. The reaction mixture was heated
to 120 °C for 3.5 h. The reaction was then cooled to room tem-
perature, diluted with distilled water (50 mL), and extracted
with diethyl ether (6 × 15 mL). The combined organic phases
were dried over Na2SO4, filtered, and concentrated. The prod-
uct was isolated by column chromatography (hexanes/EtOAc,
2:1) to give 28 as a white crystalline solid (60 mg, 83%);
mp 141–145 °C (CHCl3). Rf = 0.8 (hexanes/EtOAc, 1:1).
20
(CHCl3). ½aꢀD +32.6 (c 0.63, CHCl3). Rf = 0.2 (hexanes/
EtOAc, 1:1). IR (KBr, cm–1) n: 3416, 2987, 2952, 2932,
2893, 2857, 1660, 1618, 1501, 1481, 1439, 1384, 1350,
1
1295, 1250, 1220, 1169, 1083, 1056, 841. H NMR (CDCl3,
20
½aꢀD –63.0 (c 1.0, CHCl3). IR (KBr, cm–1) n: 3449, 2953,
600 MHz) d: 6.61 (s, 1H), 6.05 (s, 1H), 6.00 (s, 1H), 5.86 (s,
1H), 4.68(s, 1H), 4.20 (d, J = 4.5 Hz, 1H), 4.14 (dd, J = 8.2,
5.0 Hz, 1H), 4.08 (s, 3H), 3.95 (dt, J = 10.1, 6.0 Hz, 1H),
3.67–3.66 (m, 1H), 3.49 (dd, J = 13.8, 8.4 Hz, 1H), 3.28
(dd, J = 13.8, 3.7 Hz, 1H), 2.53–2.52 (m, 1H), 1.91 (s, 1H),
1.46 (s, 3H), 1.40 (s, 3H), 0.91 (s, 9H), 0.18 (s, 6H). 13C
NMR (CDCl3, 150 MHz) d: 163.6, 152.2, 145.4, 137.3,
136.3, 116.3, 101.7, 99.8, 78.3, 78.0, 67.4, 60.9, 58.8, 52.7,
45.2, 35.3, 28.1, 26.2, 25.7, 17.9, –4.92, –4.94. MS (+FAB)
m/z (%): 508 ([M + 1], 14), 507 ([M+], 37), 450 (16), 449
(10), 434 (17), 433 (16), 392 (22), 374 (12), 345 (12), 261
(100). HR-MS (+FAB) calcd for C25H37NO8Si [M]+:
507.2289; found: 507.2287.
2931, 2901, 2858, 1721, 1674, 1655, 1637, 1627, 1603,
1461, 1385, 1351, 1341, 1304, 1271, 1219, 1113, 1081,
1
837. H NMR (CDCl3, 600 MHz) d: 12.70 (s, 1H), 8.07
(d, J = 8.0 Hz, 2H), 7.62–7.60 (m, 1H), 7.49–7.47 (m,
2H), 6.59 (s, 1H), 6.11 (s, 1H), 6.06 (s, 1H), 6.02 (s, 1H),
4.68 (s, 1H), 4.55–4.54 (m, 2H), 4.23–4.21 (m, 2H), 3.63–
3.59 (m, 1H), 3.43 (dd, J = 14.4, 3.5 Hz, 1H), 2.89 (s br,
1H), 1.53 (s, 3H), 1.42 (s, 3H), 0.91 (s, 9H), 0.16–0.15 (m,
6H). 13C NMR (CDCl3, 150 MHz) d: 170.0, 166.4, 153.2,
146.9, 134.0, 133.2, 133.1, 129.8, 129.6, 129.4, 110.1,
107.3, 102.3, 97.3, 78.4, 77.7, 67.2, 61.6, 53.2, 41.8, 34.0,
30.3, 28.4, 26.1, 25.7, 17.9, –4.9, –5.0. MS (+FAB) m/z
(%): 599 ([M + 2]+, 11), 598 (29), 597 (8), 596 (4), 179
(11), 73 (100). HR-MS (+FAB) calcd for C31H40N1O9Si+
[M + 1]+: 598.2472; found: 598.2446.
((3aS,3bR,10bR,11S,12S,12aS)-12-{[Tert-butyl(dimethyl)
silyl]oxy}-6-methoxy-2,2-dimethyl-5-oxo-
3a,3b,4,5,10b,11,12,12a-octahydrobis[1,3]dioxolo[4,5-
c:4′,5′-j]phenanthridin-11-yl)methyl benzoate (27)
[(3aS,3bR,10bR,11S,12S,12aR)-6,12-Dihydroxy-2,2-
dimethyl-5-oxo-3a,3b,4,5,10b,11,12,12a-octahydrobis[1,3]
dioxolo[4,5-c:4′,5′-j]phenanthridin-11-yl]methyl benzoate
(29)
Compound 28 (42 mg, 0.07 mmol) was dissolved in THF
(2.5 mL) and cooled to 0 °C. A 1 mol/L solution of TBAF in
THF (0.077 mL, 0.077 mmol) was added dropwise and the
reaction mixture was stirred until total consumption of the
starting material was observed (TLC). The reaction mixture
was quenched by distilled water (5 mL) and extracted with
CH2Cl2 (6 × 15 mL). The combined organic phases were
dried over Na2SO4, filtered, and evaporated. The final prod-
uct was isolated by column chromatography (hexanes/EtOAc,
To a solution of 24 (121 mg, 0.24 mmol) in CH2Cl2
(25 mL) was added triethylamine (0.04 mL, 0.48 mmol) at
0 °C followed by benzoyl chloride (0.03 mL, 0.26 mmol)
and crystals of 4-dimethylaminopyridine (DMAP). The reac-
tion mixture was stirred at 0 °C until total consumption of
the starting material was observed (TLC). The reaction mix-
ture was quenched with distilled water (10 mL) and extracted
with CH2Cl2 (3 × 15 mL). The combined organic phases
were washed with a solution of citric acid (10%, 10 mL),
dried over sodium sulfate, filtered, and concentrated. The fi-
nal product was isolated by column chromatography (gra-
dient hexanes/EtOAc 2:1 to 1:1) to give 27 as a white
Published by NRC Research Press

Pagination not final/Pagination non finale
Vshyvenko et al.
11
1:1) to give 29 as a white crystalline solid (27.2 mg, 80%);
HTB-81), were cultured in Dulbecco’s modified Eagle’s me-
dium (Cellgro) supplemented with 10% FBS, 100 mg/L pen-
icillin G, and 100 mg/L streptomycin. Human mammary
carcinoma cells, MCF-7 (ATCC No. HTB-22), were cultured
using Dulbecco’s modified Eagle’s medium supplemented
with 10% FBS, 100 mg/L penicillin G, 100 mg/L streptomy-
cin, 1.0 mmol/L GlutaMAX, and 1.0 mmol/L sodium pyru-
vate (Gibco).
21
mp > 200 °C (THF). ½aꢀD –40.1 (c 1.0, THF). Rf = 0.3
(hexanes/EtOAc, 1:1). IR (KBr, cm–1) n: 3446, 3255, 2986,
2930, 2905, 2854, 1720, 1672, 1626, 1601, 1466, 1384,
1
1356, 1343, 1311, 1222, 1166, 1088, 1071, 1027, 713. H
NMR (DMSO-d6, 600 MHz) d: 13.36 (s, 1H), 8.57 (s, 1H),
7.97–7.96 (m, 2H), 7.69–7.67 (m, 1H), 7.56–7.53 (m, 2H),
6.74 (s, 1H), 6.07 (s, 1H), 5.99 (s, 1H), 5.56 (dd, J =
4.3 Hz, 1H), 4.53 (dd, J = 11.3, 4.3 Hz, 1H), 4.46–4.45 (m,
1H), 4.43–4.39 (m, 1H), 4.29 (d, J = 5.0 Hz, 1H), 4.19 (dd,
J = 8.3, 5.3 Hz, 1H), 3.63 (dd, J = 14.4, 8.4 Hz, 1H), 3.26
(dd, J = 14.4, 3.6 Hz, 1H), 2.97–2.95 (m, 1H), 1.45 (s, 3H),
1.33 (s, 3H). 13C NMR (DMSO-d6, 150 MHz) d: 169.9,
166.1, 152.7, 146.3, 135.0, 133.9, 132.5, 130.0, 129.7,
129.2, 109.0, 107.7, 102.3, 97.9, 78.0, 77.1, 65.5, 62.0,
53.4, 34.3, 30.3, 28.3, 26.4. MS (+FAB) m/z (%): 484 ([M
+ 1]+, 4), 483 ([M]+, 26), 248 (13), 247 (59), 232 (13), 231
(28), 205 (11), 122 (16), 105 (100). HR-MS (+FAB) calcd
for C25H25NO9 [M]+: 483.1529; found: 483.1532.
MTT assay
To evaluate the cytotoxic effects of the C-1 homologues of
pancratistatin, mitochondrial dehydrogenase activities were
measured. Briefly, BxPC-3, NCI-H460, DU-145, and MCF-7
lines were assessed by seeding 4 × 103 cells/well into micro-
plates. The cells were grown for 24 h before treatment at
concentrations ranging from 0.001 to 10 µmol/L and incu-
bated for 48 h. MTT reagent (5 mg/mL, MP Biomedical, So-
lon, Ohio) was added to each well and incubated further for
2 h. The resulting formazan crystals were dissolved in
DMSO and the optical density (OD) was determined at a
wavelength of 490 nm. The experiments were repeated at
least twice for each compound per cell line. Cells treated
with 0.1% DMSO and narciclasine were used as negative
and positive controls, respectively.
[(1S,2S,3R,4S,4aR,11bR)-2,3,4,7-Tetrahydroxy-6-oxo-
1,2,3,4,4a,5,6,11b-octahydro[1,3]dioxolo[4,5-j]
phenanthridin-1-yl]methyl benzoate (9)
Compound 29 (32 mg, 0.066 mmol) was dissolved in a
mixture of CH2Cl2–CH3OH (1:1, 4 mL) and cooled to 0 °C.
Trifluoroacetic acid (1 mL) was added dropwise and the re-
action was stirred until total consumption of the starting ma-
terial was observed (TLC). The reaction mixture was dried in
vacuo, triturated with CH2Cl2 (3 × 15 mL), and finally dried
under high vacuum. The final product was isolated by col-
umn chromatography on silica gel (deactivated by 10% w/w
of water, gradient CH2Cl2–CH2Cl2/CH3OH (50:1) to
CH2Cl2/CH3OH (25:1)) to give 9 as a white crystalline
compound (25 mg, 85%); mp > 200 °C (CH2Cl2). Rf =
Cell culture
The human colorectal cancer cell line HCT 116 (ATCC,
Cat. No. CCL-247, Manassas, Virginia) was grown and cul-
tured with McCoy’s mMedium 5a (Gibco BRL, VWR, Mis-
sissauga, Ontario) supplemented with 2 mmol/L L-glutamine,
10% FBS, and 10 mg/mL gentamicin (Gibco BRL, VWR). A
human osterosarcoma cell line, Saos-2 (ATCC, Cat. No.
HTB-85, Manassas, Virginia), was grown in McCoy’s 5A
mMedium Modified (Sigma-Aldrich Canada, Mississauga,
Ontario) supplemented with 15% (v/v) FBS (Thermo Scien-
tific, Waltham, Massachusetts) and 10 mg/mL gentamicin
(Gibco BRL, VWR). All cells were grown at 37 °C and 5%
CO2.
20
0.6 (CH2Cl2/CH3OH 10:1). ½aꢀD –24.9 (c 1, THF). IR
(KBr, cm–1) n: 3423, 3386, 2956, 2921, 2852, 1716, 1672,
1627, 1600, 1466, 1384, 1363, 1340, 1278, 1095, 1072,
1038, 711. 1H NMR (DMSO-d6, 600 MHz) d: 13.27 (s,
1H), 8.00 (d, J = 7.7 Hz, 2H), 7.68 (t, J = 7.6 Hz, 1H),
7.57–7.54 (m, 2H), 7.43 (s, 1H), 6.72 (s, 1H), 6.07 (s,
1H), 6.01 (s, 1H), 5.19 (m, 2H), 5.13 (m, 1H), 4.66 (t, J =
10.7 Hz, 1H), 4.48 (dd, J = 10.9, 4.0 Hz, 1H), 4.24–4.23
(m, 1H), 3.88–3.87 (m, 1H), 3.75–3.73 (m, 1H), 3.62 (dd,
J = 13.6, 9.9 Hz, 1H), 3.31 (dd, J = 13.8, 4.2 Hz, 1H),
2.85–2.84 (m, 1H). 13C NMR (DMSO-d6, 150 MHz) d:
170.0, 166.3, 152.8, 146.3, 135.8, 133.8, 132.5, 130.3,
129.7, 129.2, 107.5, 102.4, 98.0, 73.1, 71.2, 69.2, 62.5,
51.6, 36.6. MS (+FAB) m/z (%): 444 ([M + 1]+, 4), 219
(12), 136 (11), 121 (11), 109 (15), 107 (17), 105 (23), 97
(18), 95 (29), 55 (100). HR-MS (+FAB) calcd for
C22H22NO9 [M + 1]+: 444.1295; found: 444.1262.
WST-1 assay for cell viability
The WST-1-based colorimetric assay was conducted as per
the manufacturer’s protocol (Roche Applied Science, Indian-
apolis, Indiana) to determine cell viability as a function of
active cell metabolism. Ninty-six well, clear bottom tissue
culture plates were seeded with approximately 2.0 × 103 HCT
116 cells/well or 6.0 × 103 Saos-2 cells/well, and treated
with various compounds at the indicated concentrations and
durations. Following treatment, the WST-1 reagent was
added to each well and incubated for 4 h at 37 °C. Absorb-
ance readings were taken at 450 nm on a Wallac Victor3
1420 multilabel counter (PerkinElmer, Woodbridge, Ontario)
and were expressed as percentages of the solvent control
group (Me2SO).
Cell culture
The human non-small cell lung cancer line, NCI-H460
(American Type Culture Collection (ATCC) No. HTB-177),
and human pancreatic adenocarcinoma cancer cell line,
BxPC-3 (ATCC No. CRL-1687,) were cultured in RPMI-
1640 medium supplemented with 10% fetal bovine serum
(FBS) (GIBCO BRL Carlsbad, California), 100 mg/L penicil-
lin G, and 100 mg/L streptomycin (Cellgro, Manassas, Vir-
ginia). Human prostate carcinoma cells, DU-145 (ATCC No.
Supplementary data
1
Supplementary data (copies of H and 13C NMR spectra
for compounds 7–9, 12, 13, and 15–29) are available with the
article through the journal Web site at http://nrcresearchpress.
com/doi/suppl/10.1139/v2012-073.
Published by NRC Research Press

Pagination not final/Pagination non finale
12
Can. J. Chem. Vol. 90, 2012
Acknowledgements
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The authors are grateful to the following agencies for fi-
nancial support of this work: Natural Sciences and Engineer-
ing Research Council of Canada (NSERC) (Idea to
Innovation and Discovery Grants), Canada Research Chair
Program, Canada Foundation for Innovation (CFI), TDC Re-
search, Inc., TDC Research Foundation, and Brock Univer-
sity. A.E.R. and A.K. are thankful for the support of the
National Center for Research Resources (5P20RR016480-12)
and the National Institute of General Medical Sciences (8
P20 GM103451-12). S.P. thanks the Knights of Columbus
Chapter 9671 (Windsor, Ontario). The Canadian Institutes of
Health Research (CIHR) Canada Graduate Scholarship
awarded to D.M. is gratefully acknowledged.
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