A novel, unusually efficacious duocarmycin carbamate prodrug that releases no residual byproduct

Article abstract of DOI:10.1021/jm300330b

A unique heterocyclic carbamate prodrug of seco-CBI-indole₂ that releases no residual byproduct is reported as a new member of a class of hydrolyzable prodrugs of the duocarmycin and CC-1065 family of natural products. The prodrug was designed to be activated by hydrolysis of a carbamate releasing the free drug without the cleavage release of a traceable extraneous group. Unlike prior carbamate prodrugs examined that are rapidly cleaved in vivo, the cyclic carbamate was found to be exceptionally stable to hydrolysis under both chemical and biological conditions providing a slow, sustained release of the exceptionally potent free drug. An in vivo evaluation of the prodrug found that its efficacy exceeded that of the parent drug, that its therapeutic window of efficacy versus toxicity is much larger than the parent drug, and that its slow free drug release permitted the safe and efficacious use of doses 150-fold higher than the parent compound.

Full text of DOI:10.1021/jm300330b

Article
pubs.acs.org/jmc
A Novel, Unusually Efficacious Duocarmycin Carbamate Prodrug
That Releases No Residual Byproduct
Amanda L. Wolfe,† Katharine K. Duncan,† Nikhil K. Parelkar,§ Scott J. Weir,§ George A. Vielhauer,§,‡
and Dale L. Boger*,†
†Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 N. Torrey Pines
Road, La Jolla, California 92037, United States
§
‡Department of Urology and University of Kansas Cancer Center, University of Kansas Medical Center, 3901 Rainbow Boulevard,
Kansas City, Kansas 66160, United States
S
* Supporting Information
ABSTRACT: A unique heterocyclic carbamate prodrug of seco-CBI-indole2 that
releases no residual byproduct is reported as a new member of a class of
hydrolyzable prodrugs of the duocarmycin and CC-1065 family of natural products.
The prodrug was designed to be activated by hydrolysis of a carbamate releasing the
free drug without the cleavage release of a traceable extraneous group. Unlike prior
carbamate prodrugs examined that are rapidly cleaved in vivo, the cyclic carbamate
was found to be exceptionally stable to hydrolysis under both chemical and
biological conditions providing a slow, sustained release of the exceptionally potent free drug. An in vivo evaluation of the
prodrug found that its efficacy exceeded that of the parent drug, that its therapeutic window of efficacy versus toxicity is much
larger than the parent drug, and that its slow free drug release permitted the safe and efficacious use of doses 150-fold higher than
the parent compound.
DNA in a sequence selective manner.5,6 Comprehensive studies
INTRODUCTION
■
of the natural products, their synthetic unnatural enantiomers,7
and key analogues have defined many of the fundamental
features that control the DNA alkylation selectivity, efficiency,
and catalysis, resulting in a detailed understanding of the
relationships between structure, reactivity, and biological
activity.6−8
Duocarmycin SA (1)1 and CC-1065 (2)2 are two parent
members of a class of highly potent naturally occurring
antitumor agents that also include duocarmycin A3 and
yatakemycin4 (Figure 1). This unique class of natural products
derives its antitumor properties from their ability to alkylate
CBI (1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4-one) is
one of the most extensively studied synthetic analogues of the
family since we first introduced it in 1989.9 The CBI alkylation
subunit not only is more synthetically accessible and
participates in the now characteristic DNA alkylation reaction
effectively10 but also has been found to be 4 times more stable
and 4 times more potent than the naturally occurring alkylation
subunit of 2, approaching the stability and potency of the
duocarmycin SA (1) alkylation subunit. Since analogues
incorporating the CBI alkylation subunit have also been
established to exhibit efficacious in vivo antitumor activity in
animal models, it is an excellent synthetic replacement on
which to examine the structure−function features of the natural
products, including new prodrug design and evaluation.11
During the course of the total syntheses of the natural
products and related analogues including CBI-indole2 (5),11 it
was established that the synthetic phenol precursors such as 4,
which have yet to undergo the Winstein Ar-3′ spirocyclization,
are equipotent to and indistinguishable from their cyclized
cyclopropane containing counterparts within in vitro cytotoxic
Received: March 8, 2012
Published: May 31, 2012
Figure 1. Natural products.
© 2012 American Chemical Society
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assays, DNA alkylation studies, and in vivo antitumor models.
Because of this indistinguishable behavior both in vitro and in
vivo and because their extraordinary potency creates special
precautions for their handling, protection of the phenol
precursors not only permits safe handling during their
preparation but also provides an effective site on which to
create prodrugs that can be designed for controlled release in
vivo.12 Such prodrugs incorporating phenol acylation have been
developed to simultaneously improve solubility, pharmacoki-
netics, storage life, handling safety, and efficacy in vivo.12−14
Two such carbamate-based drugs, a derivative of duocarmycin
effectively taming the extraordinary potency of this class of
antitumor drugs.
CHEMISTRY
■
Synthesis. Prodrug (+)-6 was synthesized16 in 11 steps
from known intermediate 717 as shown in Scheme 1. The
Scheme 1
A12c,d (t1/2 = 20 h, calf serum) and carzelesin (U-80,244, t1/2
<
1 h, human plasma),12a,b which are rapidly cleaved in vivo (1−
20 h), entered clinical trials but have ultimately not progressed.
In related studies, we disclosed the carbamate prodrugs 3a−f of
(+)-CBI-indole2 as shown in Figure 2, many of which were
phenol of 7 was protected as its benzyl ether and 8 was
hydrolyzed using LiOH to provide the carboxylic acid 9 in good
overall yield. Carboxylic acid 9 was subjected to a Curtius
rearrangement using diphenylphosphorylazide (DPPA) and
Et3N in freshly distilled t-BuOH, providing the Boc protected
aniline 10 in 79% yield. The use of nondistilled t-BuOH
resulted in low yields due to competing hydrolytic release of the
free aniline. Regioselective C1 iodination of 10 and subsequent
N-alkylation of 11 with 1,3-dichloropropene proceeded
effectively, providing the cyclization precursor 12. Finally, a
selective 5-exo-trig free radical cyclization18 of 12 using
substoichiometric quantities of Bu3SnH (0.9 equiv) provided
13 in 83% yield with only trace amounts of further reduced
(debrominated) material observed.
Figure 2. Carbamate prodrug design.
found to be essentially equipotent to (+)-CBI-indole2 (5) in
vitro.12e This work established that the free drug is rapidly
released in a cellular assay and is able to spirocyclize, alkylate
DNA, and express its biological activity efficiently in a manner
essentially indistinguishable from the free drug itself. Herein,
we describe a novel intramolecular heterocyclic carbamate
(+)-CBI-indole2 prodrug (6, Figure 3) that is subject to an
Compound 13, which has served as a key precursor in the
divergent synthesis19 of a series of compounds,20 was further
elaborated to aniline 15 using triphenylsilylamine21 as an
ammonia surrogate for a Pd(0) catalyzed aryl amination22 with
LiHDMS in THF and ligand 14 (Scheme 2). Fortunately, a
solution of LiHMDS could be used in place of solid LiHMDS,
which alleviated the need for use of a glovebox as reported.22
Other amination reactions, including the use of benzophenone
imine and copper-promoted couplings with acetamidine,
yielded only trace amounts of the desired amination product.
Bu4NF deprotection of the resulting amine and debenzylation
of the phenol under hydrogenation conditions produced aniline
15. Aniline 15 was converted to the cyclic carbamate 16 by a
double acylation with triphosgene, which proceeded cleanly and
in quantitative yield. At this point, compound 16 was resolved
into its two enantiomers using chiral phase HPLC with 20% i-
PrOH/hexanes as the eluent. We chose to resolve 16 instead of
6 itself in order to permit access to additional resolved
Figure 3. Cyclic carbamate prodrug 6.
analogous hydrolysis mechanism of activation15 but that is
substantially more stable and upon activation does not release
any extraneous or traceable functionality into the surrounding
cellular environment. Significantly, the resulting drug is
accordingly less potent both in vitro and in vivo but
substantially safer to handle and more efficacious in vivo,
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prodrug 16 was assessed under a variety of acidic, basic, and
nucleophilic conditions. The cyclic carbamate of 16 proved to
be robust to hydrolysis under acidic conditions (1:1 TFA/
CH2Cl2, 4 N HCl in EtOAc) and was stable over a period of 48
h at 23 °C, although the Boc protecting group was readily
cleaved under such conditions. Similarly, compound 6 was also
found to be stable to the above acidic conditions for up to 72 h
at 23 °C. As shown in Figure 4, 16 was also stable to organic
Scheme 2
Figure 4. N-Boc prodrug 16 stability under basic conditions.
Footnotes in the figure indicate the following: (a) excess base (>100
equiv) used; (b) percent of 16 hydrolyzed as determined by LC/MS
analysis at 254 nm absorption. All reactions were run at 23 °C.
bases in aprotic solvents (entries 1−3), but the cyclic carbamate
was slowly hydrolyzed in the presence of NaHCO3 in protic
solvents in a reaction that proceeded at a greater rate as the
polarity of the solution increased (entries 3−6). Compound 16
was found to be completely stable in the presence of the
nucleophiles BnSH and BnOH (100 equiv) in MeOH and
THF at 23 °C for 48 h and was stable to BnNH2 in THF but
was rapidly cleaved with BnNH2 (100 equiv) in MeOH in 24 h.
Finally, the stability of the full prodrug 6 was examined in pH
7.0 phosphate buffer (t1/2 > 4 weeks, no cleavage observed) and
in human plasma (t1/2 > 48 h, 5% free drug release), indicating
that the cyclic carbamate is remarkably stable under both
conditions but subject to slow release in human plasma. By
contrast, the open chain carbamates explored in earlier studies,
including those leading to carzelesin, were designed for much
more rapid release (1−20 h). We also found that 6 is incapable
of alkylating DNA in cell-free systems23 (see Supporting
Information), indicating that any in vitro cytotoxic activity or in
vivo antitumor activity of 16 or 6 is due to release of the free
drug and not the prodrug itself.
analogues and to avoid the lower solubility of the full prodrug 6
in the chromatography solvents. Each enantiomer of 16 was
subjected to Boc deprotection with 4 N HCl in EtOAc and
immediate N-acylation with 17, providing (+)- and ent-(−)-6 in
52% yield.
The parent compound of 6 was prepared as shown in
Scheme 3 through a four-step sequence. The aniline of
Scheme 3
BIOLOGICAL PROPERTIES
■
In Vitro Cytotoxic Activity. Both (+)- and ent-(−)-6 and
their N-Boc precursors 16 were tested for cell growth inhibition
in a cytotoxic assay with the L1210 murine leukemia cell line
(Figure 5). The natural enantiomer of the prodrug (+)-6 was
intermediate 15 was differentially protected as a Fmoc
carbamate. Subsequent Boc deprotection and coupling with
carboxylic acid 17 gave 19, which was Fmoc deprotected and
cyclized upon treatment with piperidine to provide the parent
compound 20 as a racemic mixture.
Stability of the Cyclic Carbamate Prodrug. In order to
determine the ability of the free drug to be released under
physiological conditions, the chemical reactivity of N-Boc-
Figure 5. In vitro cytotoxic activity.
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found to be approximately 200-fold less potent (IC50 of 6.6
nM) than the free drug seco-CBI-indole2 4 (IC50 of 30 pM) and
6-fold more potent that its unnatural enantiomer. The racemic
parent drug ( )-20 was found to have an IC50 of 210 pM,
suggesting that the active enantiomer is approximately 3- to 4-
fold less active than 4 and indicating that the prodrug (+)-6 is
30- to 70-fold less potent than the parent drug 20. Consistent
with expectations, the full prodrug 6 proved to be 100−1000
times more potent than its N-Boc precursor 16, which in turn is
50- to 100-fold less active than N-Boc-CBI (natural enantiomer
IC50 = 80 nM).9 These data are consistent with the remarkable
stability of the prodrug to chemical hydrolysis conditions, in pH
7 phosphate buffer, and in human plasma and its ineffective in
vitro DNA alkylation reaction21 (not shown), indicating that
the release of free drug is similarly slow under the conditions of
an in vitro cellular assay as well. Despite the lower potency
relative to the free drug 4 and the racemic parent compound
20, it is notable that the cyclic carbamate prodrug (+)-6 now
displays an in vitro cellular potency (IC50 of 1−10 nM) on par
with most clinically used antitumor drugs.
In Vivo Antitumor Activity. Even though results of the in
vitro cellular assay showed that (+)-6 is substantially less potent
than its parent drug, the slow release of the compound could
prove to be advantageous in vivo because of the inherent
potency and toxicity of the parent compound. Therefore, the in
vivo antitumor activity of (+)-6 was assessed alongside seco-
CBI-indole2 (4) in an antitumor model consisting of L1210
murine leukemia cells implanted ip into DBA/2J mice (Figure
6), which has been used traditionally as an initial antitumor
behavior of prodrug (+)-6. As anticipated, (+)-CBI-indole2 (4)
proved to be toxic at doses of 100−500 μg/kg, leading to
premature death of the animals, and productive antitumor
activity was observed only at the dose of 60 μg/kg (T/C =
197), albeit producing only 1/10 long-term (250 days)
survivors in this extended study (Figure 6). By contrast, the
prodrug (+)-6 exhibited productive antitumor activity over the
entire and much larger dose range examined (30-fold range).
The most efficacious activity was observed at the highest dose
of 9000 μg/kg, producing 5/10 long-term cures (>250 days, T/
C > 980) and indicating that even higher doses may be not only
tolerable but potentially even more efficacious. This highest
dose represents one that is 150 times greater than the optimal
dose observed with (+)-4, in line with the 100- to 200-fold
differences in their cytotoxic potencies. In addition, the dose
range over which (+)-6 exhibited productive activity was much
larger, the in vivo antitumor activity was more efficacious (T/C
> 980), and long-term cures (5/10 > 250 day survivors) were
observed even without an effort at dosing optimization.
CONCLUSIONS
■
A novel heterocyclic carbamate prodrug 6 of (+)-CBI-indole2,
which can be released via hydrolysis, was synthesized and
evaluated for its in vitro cytotoxic activity and in vivo antitumor
activity. Compared to its open chain counterparts explored in
earlier studies, the cyclic carbamate prodrug was found to be
remarkably stable to chemical hydrolysis conditions as well as in
pH 7.0 phosphate buffer and human plasma. Accordingly, 6 was
less potent in vitro and in vivo compared to the parent drug 4
but was found to be substantially safer and more efficacious in
vivo, being superior in extending life expectancy of tumor-
bearing animals even at 150-fold higher doses. Notable
elements of the cyclic carbamate prodrug behavior include
not only its hydrolysis liberation of the free drug that releases
no residual byproduct (CO2) but also its remarkable stability
relative to its acyclic counterparts explored in early studies. This
results in an apparent slow, sustained release of free drug that
permits the safer and more efficacious use of larger doses of
drug (as much as 150-fold), effectively taming the extraordinary
potency of this class of antitumor drugs
EXPERIMENTAL SECTION
■
General. Reagents and solvents were purchased reagent-grade and
used without further purification. Pooled human plasma, with sodium
citrate as an anticoagulant, was purchased from Innovative Research
and stored at −20 °C. THF was freshly distilled from sodium
benzophenone ketyl. t-BuOH was freshly distilled from calcium
hydride. All reactions were performed in oven-dried glassware under
an Ar atmosphere. Evaporation and concentration in vacuo were
performed at 20 °C. TLC was conducted using precoated SiO2 60
F254 glass plates from EMD with visualization by UV light (254 or
366 nm). Chiral phase HPLC was performed using a Shimadzu HPLC
on a semipreparative Diacel ChiralCel OD column (0.46 cm × 25 cm)
with a flow rate of 7 mL/min and with UV detection at λ = 254 nm.
Optical rotations were determined on a Rudolf Research Analytical
Autopol III automatic polarimeter (λ = 589 nm, 25 °C). NMR (1H or
13C) were recorded on Bruker DRX-500 and DRX-600 NMP
spectrophotometers at 298 K. Residual solvent peaks were used as
an internal reference. Coupling constants (J) (H,H) are given in Hz.
Coupling patterns are designated as singlet (s), doublet (d), triplet (t),
quadruplet (q), multiplet (m), or broad singlet (br). IR spectra were
recorded on a Thermo Scientific Nicolet 380 FT-IR spectropho-
tometer and measured neat. High resolution mass spectral data were
acquired on an Agilent Technologies high resolution LC/MSD-TOF,
and the detected masses are given as m/z with m representing the
Figure 6. In vivo antitumor activity (L1210, ip). Footnotes in the
figure indicate the following: (a) dose (μg/kg of animal) administered
ip on days 1, 5, and 9; (b) MSP = mean survival period (days); (c) T/
C = [treated/(control (MSP))] × 100; (d) number of live animals
after 250 days (terminated).
model for comparisons in this class.11,12,14,15 A dose range of
300−9000 μg/kg for prodrug (+)-6 (scaled to its in vitro
cytotoxic activity IC50) and 60−500 μg/kg for seco-CBI-indole2
(4) and a dosing schedule (administered three times ip on days
1, 5, and 9) for both compounds were employed. A subtle but
additional important empirical observation made in the studies
is that the prodrug administration is tolerated at the injection
sites of the animals much better than the free drug.
The optimal does range for 4 was previously established
(60−100 μg/kg) and was extended for the study herein to
highlight its narrow therapeutic window versus the potential
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molecular ion. The purity of each tested compound (>95%) was
determined on an Agilent 1100 LC/MS instrument using a ZORBAX
SB-C18 column (3.5 mm, 4.6 mm × 50 mm, with a flow rate of 0.75
mL/min and detection at 220 and 254 nm) with a 10−98%
acetonitrile/water/0.1% formic acid gradient.
products were combined to give 9 (2.09 g, 100%) as a pale yellow
solid. 1H NMR (DMSO-d6, 500 MHz) δ 8.23 (s, 1H), 8.10 (d, J = 6.0
Hz, 1H), 7.91 (d, J = 6.5 Hz, 1H), 7.61 (d, J = 7.0 Hz, 2H), 7.55 (s,
1H), 7.43−7.39 (m, 3H), 7.33 (t, J = 7.0 Hz, 1H), 5.35 (s, 2H). 13C
NMR (DMSO-d6, 125 MHz) δ 166.7, 153.8, 136.2, 135.9, 135.0,
129.8, 128.8, 128.2, 127.8, 127.7, 127.5, 124.7, 123.8, 115.5, 107.0,
70.4. IR (film) νmax 3368, 2969, 1680 cm−1. ESI-TOF HRMS m/z
357.0125 (M + H+, C18H13BrO3 requires 357.0121).
Ethyl 5-Bromo-4-hydroxy-2-naphthoate (7). A solution of
potassium tert-butoxide (20.0 g, 0.78 mol) at 55 °C in t-BuOH (249
mL) was treated with a premixed solution of diethyl succinate (40.4
mL, 0.243 mol) and 3-bromobenzaldehyde (18.9 mL, 0.162 mol)
dropwise. Upon completion of the addition, the reaction mixture was
warmed to 85 °C and stirred for 2 h. After 2 h, the reaction mixture
was cooled to 25 °C. The reaction mixture was acidified to pH < 4
with 2 N aqueous HCl and concentrated. The aqueous suspension was
then extracted with ethyl acetate (3×). The organic layers were
combined and washed with saturated aqueous NaHCO3 (5×). The
basic aqueous washes were combined and reacidified with 2 N aqueous
HCl to pH 1. Finally, the aqueous phase was extracted with ethyl
acetate (3×). The organic layers were combined, dried over Na2SO4,
and concentrated under reduced pressure, which afforded the desired
half ester (39.1 g, 77%) as an orange oil. The half ester (39.1 g, 0.124
mol) was dissolved in acetic anhydride (178 mL), and NaOAc (18.7 g,
0.137 mol) was added. The reaction mixture was warmed to 140 °C
and stirred for 6 h. Upon completion, the reaction mixture was cooled
to 25 °C and poured into H2O. The aqueous layer was extracted with
ethyl acetate (3×). The organic layers were combined, dried over
Na2SO4, and concentrated under reduced pressure. The residue was
dissolved in anhydrous ethanol (620 mL). K2CO3 (104 g, 0.624 mol)
was added, and the reaction mixture was warmed at 80 °C for 1 h. The
reaction mixture was cooled and acidified to pH 1 with 2 N aqueous
HCl. The ethanol was removed under reduced pressure, and the
aqueous suspension was extracted with ethyl acetate (3×). The organic
extracts were combined, dried over Na2SO4, and concentrated under
reduced pressure. Flash chromatography (SiO2, 16 cm × 30 cm, 0−
15% EtOAc/hexanes gradient elution) provided 7 (5.4 g, 15% over
three steps) as a yellow solid and its 7-bromo isomer (12.4 g, 34% over
tert-Butyl (4-(Benzyloxy)-5-bromonaphthalen-2-yl)-
carbamate (10). Carboxylic acid 9 (950 mg, 2.66 mmol) was
dissolved in freshly distilled t-BuOH (0.01 M) over 4 Å molecular
sieves. Et3N (467 μL, 3.35 mmol) and diphenylphosphorylazide (602
μL, 2.79 mmol) were added. The reaction mixture was warmed to 85
°C under Ar and stirred for 14 h. Upon completion, the mixture was
filtered through cotton to remove the molecular sieves and
concentrated under reduced pressure. The residue was diluted with
10% aqueous HCl and extracted with EtOAc (3×). The organic
extracts were combined and washed with H2O (2×) and saturated
aqueous NaCl. The organic phase was dried over Na2SO4 and
concentrated under reduced pressure. Flash chromatography (SiO2, 5
cm × 12 cm, 5% EtOAc/hexanes elution) provided 10 (1.02 g, 89%)
as a tan solid. 1H NMR (CDCl3, 600 MHz) δ 7.62 (m, 2H), 7.58 (d, J
= 7.8 Hz, 2H), 7.49 (s, 1H), 7.39 (t, J = 7.2 Hz, 2H), 7.33 (t, J = 6.0
Hz, 1H), 7.16 (t, J = 7.8 Hz, 1H), 7.03 (s, 1H), 6.58 (s, 1H), 5.22 (s,
2H), 1.54 (s, 9H). 13C NMR (CDCl3, 150 MHz) δ 155.2, 152.5,
137.6, 136.4, 136.3, 131.3, 130.0, 128.4, 127.9, 127.3, 126.9, 120.4,
120.2, 120.1, 116.6, 107.8, 101.8, 71.4, 28.3. IR (film) νmax 3325, 2977,
1702, 1156 cm−1. ESI-TOF HRMS m/z 428.0856 (M + H+,
C22H22BrNO3 requires 428.0856).
tert-Butyl (4-(Benzyloxy)-5-bromo-1-iodonaphthalen-2-yl)-
carbamate (11). Carbamate 10 (1.20 g, 2.80 mmol) was dissolved
in freshly distilled THF (0.17 M) under Ar and in the absence of light,
and TsOH·H2O (53 mg, 0.28 mmol) and N-iodosuccinamide (753
mg, 3.30 mmol) were added. The reaction mixture was allowed to stir
at 25 °C for 2 h. After 2 h, the reaction was quenched with the
addition saturated aqueous NaHCO3 and the mixture was diluted with
ethyl acetate. The organic layer was washed with saturated aqueous
NaCl, dried over Na2SO4, and concentrated under reduced pressure.
Flash chromatography (SiO2, 5 cm × 16 cm, 5% EtOAc/hexanes
elution) provided 11 (1.47 g, 94%) as an orange solid. 1H NMR
(CDCl3, 500 MHz) δ 8.16 (s, 1H), 8.09 (d, J = 8.0 Hz, 1H), 7.70 (d, J
= 7.5 Hz, 1H), 7.62 (d, J = 7.0 Hz, 2H), 7.39 (t, J = 7.2 Hz, 2H), 7.34
(d, J = 7.0 Hz, 1H), 7.32 (s, 1H), 7.25−7.22 (m, 2H), 5.28 (s, 2H),
1.58 (s, 9H). 13C NMR (CDCl3, 125 MHz) δ 155.8, 152.4, 139.0,
136.7, 135.9, 132.2, 131.9, 130.0, 128.5, 128.3, 128.0, 127.9, 121.4,
120.2, 120.1, 117.0, 101.9, 81.3, 71.3, 28.3. IR (film) νmax 3378, 2978,
1730, 1225, 1145 cm−1. ESI-TOF HRMS m/z 553.9820 (M + H+,
C22H21BrINO3 requires 553.9822).
1
three steps). H NMR (CDCl3, 500 MHz) δ 8.16 (s, 1H), 8.07 (s,
1H), 7.89 (d, J = 6.5 Hz, 1H), 7.73 (d, J = 6.5 Hz, 1H), 7.63 (s, 1H),
7.29 (t, J = 10 Hz, 1H), 4.43 (q, J = 6.0 Hz, 2H) 1.44 (t, J = 6.0 Hz,
3H). 13C NMR (CDCl3, 125 MHz) δ 165.9, 152.7, 136.3, 133.6, 130.6,
129.2, 126.7, 123.6, 122.7, 115.2, 112.3, 61.3, 14.3. IR (film) νmax 3367,
2979, 1690, 1227 cm−1. ESI-TOF HRMS m/z 294.9959 (M + H+,
C13H11BrO3 requires 294.9964).
Ethyl 4-(Benzyloxy)-5-bromo-2-naphthoate (8). Naphthol 7
(3.20 g, 11.0 mmol) was dissolved in anhydrous DMF (78 mL).
K2CO3 (3.05 g, 22.0 mmol), benzyl bromide (1.59 mL, 13.2 mmol),
and Bu4NI (163 mg, 0.440 mmol) were added. The solution was
stirred at 25 °C for 16 h. The reaction mixture was poured into H2O
and extracted with ethyl acetate (3×). The organic extracts were
combined, dried over Na2SO4, and concentrated under reduced
pressure. The solid was recrystallized with 5% EtOAc/hexanes and the
mother liquor was further purified by flash chromatography (SiO2, 6
cm × 15 cm, 10−20% EtOAc/hexanes gradient elution), affording
tert-Butyl (4-(Benzyloxy)-5-bromo-1-iodonaphthalen-2-yl)-
(3-chloroallyl)carbamate (12). Compound 11 (1.65 g, 2.99
mmol) and Bu4NI (55 mg, 0.15 mmol) were dissolved in anhydrous
DMF (0.16 M), and the solution was cooled to 0 °C. Once cooled,
60% NaH in mineral oil (239 mg, 5.98 mmol) was added and the
reaction mixture was allowed to stir at 0 °C for 30 min. 1,3-
Dichloropropene (0.84 mL, 8.97 mmol) was added dropwise, and the
solution was warmed to room temperature. After 1 h, the reaction
mixture was quenched with the addition of saturated aqueous NH4Cl
and diluted with ethyl acetate. The organic layer was washed with
H2O, saturated aqueous NaCl, dried over Na2SO4, and concentrated
under reduced pressure. Flash chromatography (SiO2, 5 cm × 8 cm,
10% EtOAc/hexanes elution) provided an E/Z mixture of alkene 12
1
additional 8 (3.30 g combined, 77%) as a brown crystalline solid. H
NMR (CDCl3 500 MHz) δ 8.17 (s, 1H), 7.87 (d, J = 7.5 Hz, 1H), 7.85
(d, J = 8.0 Hz, 1H), 7.61 (d, J = 7.5 Hz, 2H), 7.57 (s, 1H), 7.40 (t, J =
7.5 Hz, 2H), 7.35−7.32 (m, 1H), 7.29 (t, J = 7.5 Hz, 1H), 5.30 (s,
2H), 4.43 (q, J = 7.0 Hz, 2H), 1.44 (t, J = 7.0 Hz, 3H). 13C NMR
(CDCl3, 125 MHz) δ 166.0, 154.5, 136.1, 136.0, 135.0, 129.3, 128.3,
128.0 (2C), 127.8, 126.8, 125.8, 124.1, 116.7, 106.9, 71.2, 61,1. IR
(film) νmax 2980, 1712, 1413, 1236 cm−1. ESI-TOF HRMS m/z
385.0433 (M + H+, C20H17BrO3 requires 385.0434).
4-(Benzyloxy)-5-bromo-2-naphthoic Acid (9). Ester 8 (2.29 g,
5.94 mmol) was dissolved in a 3:1:1 mixture of THF/CH3OH/H2O
(0.1 M). LiOH·H2O was added, and the reaction mixture was stirred
at 25 °C for 24 h. Upon completion, the reaction mixture was acidified
to pH 1 with the addition of 10% aqueous HCl. A precipitate formed
during the acidification, and it was collected by vacuum filtration. The
remaining aqueous layer was then extracted with ethyl acetate (3×).
The organic extracts were combined, dried over Na2SO4, and
concentrated under reduced pressure. The filtered and extracted
1
(1.818 g, 96%) as a yellow foam. H NMR (acetone-d6, 600 MHz) δ
8.35 (m, 2H), 7.92 (d, J = 7.2 Hz, 2H), 7.61 (br, 4H) 7.45 (t, J = 8.4
Hz, 2H), 7.42−7.40 (m, 4H), 7.35−7.33 (m, 2H), 7.18 (d, J = 18.0
Hz, 2H), 6.21−6.08 (m, 3H), 5.39 (s, 4H), 4.60 (dd, J = 18.9, 5.4 Hz,
1H), 4.42 (dd, J = 15.0, 7.2 Hz, 1H), 4.27 (dd, J = 15.6, 6.6 Hz, 1H),
3.99 (dd, J = 14.1, 6.6 Hz, 1H), 1.55 (br, 4H), 1.28 (br, 14H). 13C
NMR (acetone-d6, 150 MHz) δ 157.28, 157.27, 154.8, 154.6, 145.9,
145.8, 139.2, 139.1, 138.2, 138.1, 136.13, 136.12, 135.4 (2C), 130.9,
130.2, 130.1, 129.9, 129.86, 129.81, 129.7, 129.4, 126.2, 125.4, 123.2,
5882
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122.2, 118.2, 112.4, 112.3, 98.0, 97.2, 82.2, 81.8, 72.8, 72.6, 50.6, 47.3,
29.3. IR (film) νmax 2974, 2928, 1697, 1156, 749 cm−1. ESI-TOF
HRMS m/z 627.9750 (M + H+, C25H24BrClINO3 requires 627.9746).
tert-Butyl 1,2-Dihydro-5-(benzyloxy)-6-bromo-1-(chloro-
methyl)-1H-benzo[e]indole-3(2H)-carboxylate (13). Alkene 12
(1.81 g, 2.89 mmol) and AIBN (140 mg, 0.86 mmol) were dissolved in
benzene (0.05 M). Freshly prepared Bu3SnH (701 μL, 2.60 mmol)
was added, and the system was purged of oxygen using Ar and the
freeze/pump/thaw method. The reaction mixture was warmed to 80
°C for 12 h. Upon completion, the reaction mixture was concentrated
under reduced pressure and purified by flash chromatography (10% w/
w KF fused SiO2, 5 cm × 16 cm, 0−10% EtOAc/hexanes gradient
elution) to provide 13 (1.32 g, 90%) as a white solid. 1H NMR
(acetone-d6, 600 MHz) δ 7.98 (br, 1H), 7.81 (d, J = 8.4 Hz, 1H),
7.65−7.63 (m, 3H), 7.41 (t, J = 7.2 Hz, 2H), 7.35−7.30 (m, 2H), 5.31
(s, 2H), 4.21−4.16 (m, 2H), 4.12−4.09 (m, 1H), 3.96 (dd, J = 11.1,
3.0 Hz, 1H), 3.71 (dd, J = 8.4, 11.4 Hz, 1H), 1.58 (s, 9H). 13C NMR
(acetone-d6, 150 MHz) δ 157.8, 153.8, 144.3, 138.4, 135.0, 132.6,
130.1 129.7, 129.4, 124.9, 124.3, 121.6, 119.4, 117.2, 100.7, 82.4, 72.8,
54.4, 48.6, 43.1, 29.5. IR (film) νmax 2926, 1692, 1330, 1135, 752 cm−1.
ESI-TOF HRMS m/z 502.0772 (M + H+, C25H25BrClNO3 requires
502.0779).
1H), 3.77 (dd, J = 8.2, 11.4 Hz, 1H), 1.58 (s, 9H). 13C NMR (acetone-
d6, 150 MHz) δ 178.5, 153.7, 152.8, 148.37, 148.31, 146.1, 137.0,
136.9, 131.8, 131.2, 118.8, 116.8, 110.0, 105.5, 100.3, 82.7, 54.5, 48.5,
42.5, 29.4. IR (film) νmax 2924, 1701, 1606, 1405, 1332, 1140 cm−1.
ESI-TOF HRMS m/z 375.1105 (M + H+, C19H19ClN2O4 requires
375.1106).
The enantiomers were resolved on a semipreparative Diacel
chiralcel OD column (0.46 cm × 25 cm) with 20% i-PrOH/hexanes
elution; α = 1.38. (1S)-16: [α]23 −31 (c 0.75, THF), natural
D
enantiomer. (1R)-16: [α]23 +32 (c 0.80, THF), unnatural
D
enantiomer.
N-(2-(10-(Chloromethyl)-5-oxo-5,8,9,10-tetrahydro-4H-
pyrrolo[3′,2′:5,6]naphtho[1,8-de][1,3]oxazine-8-carbonyl)-1H-
indol-5-yl)-1H-indole-2-carboxamide (6). Compound 16 (7.5 mg,
0.020 mmol) was dissolved in 4 N HCl in EtOAc (0.5 mL), and the
mixture was allowed to stir at room temperature for 25 min. The
solvent was removed under a stream of nitrogen, and the residue was
redissolved in anhydrous DMF (0.4 mL). EDCI (11.4 mg, 0.06 mmol)
and 17 (7.0 mg, 0.022 mmol) were added, and the reaction mixture
was allowed to stir at 25 °C for 24 h. The reaction mixture was
quenched with the addition of H2O and diluted with ethyl acetate. The
organic phase was washed with 2 N aqueous HCl (3×), saturated
aqueous NaHCO3 (5×), and saturated aqueous NaCl. The organic
extract was dried over Na2SO4 and concentrated under reduced
pressure. The residue was purified by PTLC (SiO2, 40% THF/
toluene) to provide 6 (6.08 mg, 52%, typically 52−60%) as a tan solid.
1H NMR (DMSO-d6, 600 MHz) δ 11.85 (s, 1H), 11.75 (s, 1H), 11.14
tert-Butyl 1,2-Dihydro-6-amino-1-(chloromethyl)-5-hy-
droxy-1H-benzo[e]indole-3(2H)-carboxylate (15). An oven-
dried microwave vial was charged with Pd2(dba)3 (10.9 mg, 11
μmol), 2-dicyclohexylphosphinobiphenyl (14, 8.3 mg, 0.023 mmol),
and (C6H5)3SiNH2 (72.1 mg, 0.261 mmol). The vial was evacuated
and filled with Ar. Compound 13 (120 mg, 0.238 mmol) was added,
and the vial was evacuated again. Toluene (2.3 mL) was added, and
the vessel was purged with Ar. Finally, LiHMDS (0.29 mL, 1 M in
THF) was added, and the vessel was sealed. The mixture was
submerged in a 100 °C oil bath for 24 h. After 24 h, the reaction
mixture was cooled to room temperature, diluted with diethyl ether,
filtered through a plug of Celite, and concentrated. The residue was
dissolved in THF (15 mL) and cooled to 0 °C. Bu4NF (0.36 mL, 1 M
in THF) was added dropwise. The reaction mixture was allowed to stir
for 30 min before being quenched with the addition of saturated
aqueous NH4Cl and diluted with ethyl acetate. The organic layer was
washed with saturated aqueous NaCl, dried over NasSO4, and
concentrated. The residue was purified by flash chromatography
(SiO2, 4 cm × 8 cm, 5−10% EtOAc/hexanes gradient elution). The
product was carried on to the next reaction mixture without
characterization because of coelution of triphenyl byproduct. The
amine (104 mg theoretical, 0.238 mmol) was dissolved in anhydrous
CH3OH (6 mL) under Ar. Then 10% Pd/C (29 mg, 0.024 mmol) was
added and the atmosphere was exchanged with H2. The reaction
mixture was allowed to stir at 25 °C for 5 h. The reaction mixture was
diluted with diethyl ether, filtered through Celite, and concentrated
under reduced pressure. Flash chromatography (SiO2, 3 cm × 8 cm,
50−70% Et2O/hexanes gradient elution) provided 15 (56 mg, 67%
(br, 1H), 10.20 (s, 1H), 8.25 (s, 1H), 7.91 (s, 1H), 7.67 (d, J = 8.4 Hz,
1H), 7.59 (dd, J = 9.0, 1.8 Hz, 1H), 7.48 (t, J = 9.0 Hz, 2H), 7.43 (m,
4H), 7.27 (s, 1H), 7.21 (t, J = 7.8 Hz, 1H), 7.07 (t, J = 7.8 Hz, 1H),
6.66 (dd, J = 5.7, 3.0 Hz, 1H), 4.87 (t, J = 10.2 Hz, 1H), 4.61 (dd, J =
10.8, 2.4 Hz, 1H), 4.03−4.02 (m, 1H), 4.00−3.98 (m, 2H). 13C NMR
(DMSO-d6, 150 MHz) δ 160.2, 159.4, 149.8, 146.5, 143.4, 136.6,
134.8, 133.3, 131.8, 131.7, 130.7, 129.5, 129.1, 127.9, 127.03, 127.00,
126.9, 123.4, 121.5, 119.5, 119.4, 118.7, 115.3, 112.8, 112.29, 112.21,
108.7, 106.1, 104.4, 103.3, 99.8, 54.7, 47.2, 40.8. IR (film) νmax 3255,
1731, 1603, 1514, 1400, 1232, 794, 733 cm−1. ESI-TOF HRMS m/z
576.1431 (M + H+, C32H22ClN5O4 requires 576.1433). (1S)-6: [α]23
D
+18.4 (c 0.21, THF), natural enantiomer. (1R)-6: [α]23 −18.5 (c
D
0.24, THF), unnatural enantiomer.
N-(2-(5-Amino-4-oxo-1,2,9,9a-tetrahydrocyclopropa[c]-
benzo[e]indole-2-carbonyl)-1H-indol-5-yl)-1H-indole-2-car-
boxamide (20). Intermediate 15 (10 mg, 0.028 mmol) was
suspended in H2O (0.4 mL) and cooled to 0 °C. Fmoc-Cl (9.6 mg,
0.037 mmol) in dioxane (0.2 mL) was added, and the reaction mixture
was allowed to slowly warm to room temperature over 17 h. The
reaction mixture was diluted with H2O and extracted with EtOAc
(2×). The organic layers were combined, dried over Na2SO4, and
concentrated under reduced pressure. The residue was dissolved in 4
N HCl in EtOAc (0.8 mL), and the mixture was allowed to stir at
room temperature for 25 min. The solvent was removed under a
stream of nitrogen, and the residue was redissolved in anhydrous DMF
(0.8 mL). EDCI (10.7 mg, 0.056 mmol) and 17 (10.7 mg, 0.34 mmol)
were added, and the reaction mixture was allowed to stir at 25 °C for
24 h. The reaction mixture was quenched with the addition of H2O
and diluted with EtOAc. The organic phase was washed with 2 N
aqueous HCl (3×), saturated aqueous NaHCO3 (5×), and saturated
aqueous NaCl. The organic extract was dried over Na2SO4 and
concentrated under reduced pressure. The crude residue was dissolved
in DMF (0.8 mL), and piperidine (160 μL) was added. The reaction
mixture was allowed to stir at room temperature for 1 h after which the
solvent was removed under reduced pressure. The residue was purified
by PTLC (SiO2, 60% THF/toluene) to provide 20 (4.1 mg, 29% over
four steps) as a yellow solid. 1H NMR (DMSO-d6, 600 MHz) δ 11.81
(s, 1H), 11.72 (s, 1H), 10.17 (s, 1H), 8.21 (s, 1H), 7.67 (d, J = 7.8 Hz,
1H), 7.60 (d, J = 9.0 Hz, 1H), 7.47 (d, J = 9.0 Hz, 2H), 7.42 (s, 1H),
7.25 (s, 1H), 7.21 (t, J = 7.8 Hz, 1H), 7.17 (t, J = 8.4 Hz, 1H), 7.07 (t,
J = 7.8 Hz, 1H), 6.81 (s, 1H), 6.58 (d, J = 8.4 Hz, 1H), 6.20 (d, J = 7.2
Hz, 1H), 4.60−4.57 (m, 1H), 4.45 (d, J = 10.2 Hz, 1H), 3.07 (m, 1H),
1.61 (t, J = 4.8 Hz, 1H), 1.51−1.49 (m, 1H). 13C NMR (DMSO-d6,
1
over three steps) as a tan solid. H NMR (acetone-d6, 600 MHz) δ
7.48 (br, 1H), 7.12 (t, J = 7.8 Hz, 1H), 6.84 (d, J = 7.8 Hz, 1H), 6.44
(d, J = 6.6 Hz, 1H), 4.13−4.05 (m, 2H), 3.92−3.87 (m, 2H), 3.55 (t, J
= 10.8 Hz, 1H), 1.54 (s, 9H). 13C NMR (acetone-d6, 150 MHz) δ
158.6, 153.7, 148.8, 134.8, 130.0, 126.8, 115.3, 112.5, 111.4, 108.7,
99.6, 81.7, 54.1, 48.3, 43.4, 29.3. IR (film) νmax 3391, 2974, 1706, 1583,
1406, 1142 cm−1. ESI-TOF HRMS m/z 349.1323 (M + H+,
C18H21ClN2O3 requires 349.1313).
tert-Butyl 10-(Chloromethyl)-5-oxo-9,10-dihydro-4H-
pyrrolo[3′,2′:5,6]naphtho[1,8-de][1,3]oxazine-8(5H)-carboxy-
late (16). Naphthol 15 (56 mg, 0.160 mmol) and triphosgene (47 mg,
0.160 mmol) were dissolved in toluene (3.2 mL) at 25 °C. The
reaction mixture was stirred for 1 h before being diluted with H2O and
ethyl acetate. The organic layer was washed with saturated aqueous
NaCl, dried over Na2SO4, and concentrated under reduced pressure.
Flash chromatography (SiO2, 2 cm × 6 cm, 20−50% EtOAc/hexanes
1
gradient elution) provided 16 (60 mg, 100%) as a yellow solid. H
NMR (acetone-d6, 600 MHz) δ 9.86 (s, 1H), 7.66 (br, 1H), 7.37 (t, J
= 8.4 Hz, 1H), 7.32 (d, J = 8.4 Hz, 1H), 6.66 (d, J = 7.8 Hz, 1H),
4.19−4.18 (m, 2H), 4.07−4.05 (m, 1H), 3.98 (dd, J = 11.1, 3.6 Hz,
5883
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150 MHz) δ 188.8, 161.1, 159.2, 158.4, 150.5, 142.1, 136.4, 133.4,
132.4, 131.5, 131.4, 129.7, 126.7, 126.6, 123.2, 121.2, 119.6, 119.5,
113.6, 112.9, 112.6, 112.0, 110.5, 107.7, 106.8, 103.1, 63.1, 53.6, 32.4,
29.8, 24.1. ESI-TOF HRMS m/z 514.1872 (M + H+, C31H23N5O3
requires 514.1874).
(4) Igarashi, Y.; Futamata, K.; Fujita, T.; Sekine, A.; Senda, H.; Naoki,
H.; Furumai, T. Yatakemycin, a Novel Antifungal Antibiotic Produced
by Streptomyces sp. TP-A0356. J. Antibiot. 2003, 56, 107−113.
(5) For duocarmycin SA, see the following: (a) Boger, D. L.;
Johnson, D. S.; Yun, W. (+)- and ent-(−)-Duocarmycin SA and (+)-
and ent-(−)-N-BOC-DSA DNA Alkylation Properties. Alkylation Site
Models That Accommodate the Offset AT-Rich Adenine N3
Alkylation Selectivity of the Enantiomeric Agents. J. Am. Chem. Soc.
In Vivo Antitumor Activity. B6D2F1 mice were injected
intraperitoneal (ip) with syngeneic L1210 cells (1 × 106) on day 0.
Ten mice were randomly assigned to control vehicle or treatment
groups for compounds (+)-4 and (+)-6 at doses of 60, 100, 250, and
500 μg/kg per injection for (+)-4 or 300, 1000, 3000, and 9000 μg/kg
per injection for (+)-6. Compounds (+)-4 and (+)-6 were formulated
in 100% DMSO and injected ip on study days 1, 5, and 9. Following
injection of tumor cells, animals were monitored daily and weighed
two times per week. Percent survival (T/C) for treated and control
groups was determined by dividing the total survival days for each
treatment group by the total survival days for the control group and
multiplying by 100. All animal studies were carried out in the animal
facilities of The University of Kansas Medical Center, KS, with strict
adherence to the guidelines of the IACUC Animal Welfare Committee
of KUMC (IACUC Approval No. 2009-1837).
1994, 116, 1635−1656.
For yatakemycin, see the following:
(b) Parrish, J. P.; Kastrinsky, D. B.; Wolkenberg, S. E.; Igarashi, Y.;
Boger, D. L. DNA Alkylation Properties of Yatakemycin. J. Am. Chem.
Soc. 2003, 125, 10971−10976. (c) Trzupek, J. D.; Gottesfeld, J. M.;
Boger, D. L. Alkylation of Duplex DNA in Nucleosome Core Particles
by Duocarmycin SA and Yatakemycin. Nat. Chem. Biol. 2006, 2, 79−
82. (d) Tichenor, M. S.; MacMillan, K. S.; Trzupek, J. D.; Rayl, T. J.;
Hwang, I.; Boger, D. L. Systematic Exploration of the Structural
Features of Yatakemycin Impacting DNA Alkylation and Biological
Activity. J. Am. Chem. Soc. 2007, 129, 10858−10869. For CC-1065,
see the following: (e) Hurley, L. H.; Lee, C.-S.; McGovren, J. P.;
Warpehoski, M. A.; Mitchell, M. A.; Kelly, R. C.; Aristoff, P. A.
Molecular Basis for Sequence-Specific DNA Alkylation by CC-1065.
Biochemistry 1988, 27, 3886−3892. (f) Boger, D. L.; Johnson, D. S.;
Yun, W.; Tarby, C. M. Molecular Basis for Sequence Selective DNA
Alkylation by (+)- and ent-(−)-CC-1065 and Related Agents:
Alkylation Site Models That Accommodate the Offset AT-Rich
Adenine N3 Alkylation Selectivity. Bioorg. Med. Chem. 1994, 2, 115−
135. (g) Boger, D. L.; Munk, S. A.; Zarrinmayeh, H. (+)-CC-1065-
DNA Alkylation: Key Studies Demonstrating a Noncovalent Binding
Selectivity Contribution to the Alkylation Selectivity. J. Am. Chem. Soc.
1991, 113, 3980−3983. (h) Boger, D. L.; Zarrinmayeh, H.; Munk, S.
A.; Kitos, P. A.; Suntornwat, O. Demonstration of a Pronounced Effect
of Noncovalent Binding Selectivity on the (+)-CC-1065 DNA
Alkylation and Identification of the Pharmacophore of the Alkylation
Subunit. Proc. Nat. Acad. Sci. U.S.A. 1991, 88, 1431−1435. (i) Boger,
D. L.; Coleman, R. S.; Invergo, B. J.; Sakya, S. M.; Ishizaki, T.; Munk,
S. A.; Zarrinmayeh, H.; Kitos, P. A.; Thompson, S. C. Synthesis and
Evaluation of Aborted and Extended CC-1065 Functional Analogs:
(+)- and (−)-CPI-PDE-I1, (+)- and (−)-CPI-CDPI1, and (+)- and
(−)-CPI-CDPI3. Preparation of Key Partial Structures and Definition
of an Additional Functional Role of the CC-1065 Central and Right-
Hand Subunits. J. Am. Chem. Soc. 1990, 112, 4623−4632. (j) Boger, D.
L.; Munk, S. A.; Ishizaki, T. (+)-CC-1065 DNA Alkylation:
Observation of an Expected Relationship between Cyclopropane
Electrophile Reactivity and the Efficiency of DNA Alkylation. J. Am.
Chem. Soc. 1991, 113, 2779−2780. (k) Boger, D. L.; Coleman, R. S.;
Invergo, B. J.; Zarrinmayeh, H.; Kitos, P. A.; Thompson, S. C.; Leong,
T.; McLaughlin, L. W. A Demonstration of the Intrinsic Importance of
Stabilizing Hydrophobic Binding and Noncovalent van der Waals
Contacts Dominant in the Noncovalent CC-1065:DNA Binding.
Chem.-Biol. Interact. 1990, 73, 29−52. For duocarmycin A, see the
following: (l) Boger, D. L.; Ishizaki, T.; Zarrinmayeh, H.; Munk, S. A.;
Kitos, P. A.; Suntornwat, O. Duocarmycin−Pyrindamycin DNA
Alkylation Properties and Identification, Synthesis, and Evaluation of
Agents Incorporating the Pharmacophore of the Duocarmycin−
Pyrindamycin Alkylation Subunit. Identification of the CC-1065
Duocarmycin Common Pharmacophore. J. Am. Chem. Soc. 1990,
112, 8961−8971. (m) Boger, D. L.; Ishizaki, T.; Zarrinmayeh, H.
Isolation and Characterization of the Duocarmycin−Adenine DNA
Adduct. J. Am. Chem. Soc. 1991, 113, 6645−6649. (n) Boger, D. L.;
Yun, W.; Terashima, S.; Fukuda, Y.; Nakatani, K.; Kitos, P. A.; Jin, Q.
DNA Alkylation Properties of the Duocarmycins: (+)-Duocarmycin A,
epi-(+)-Duocarmycin A, ent-(−)-Duocarmycin A and epi,ent-(−)-Du-
ocarmycin A. Bioorg. Med. Chem. Lett. 1992, 2, 759−765. (o) Boger, D.
L.; Yun, W. Reversibility of the Duocarmycin A and SA DNA
Alkylation Reaction. J. Am. Chem. Soc. 1993, 115, 9872−9873.
ASSOCIATED CONTENT
* Supporting Information
■
S
A gel figure examining the DNA alkylation properties of 6, the
synthesis and characterization of N-Boc-ACBI (21) and N-
CO2Me-ACBI (24), and reactivity (solvolysis) data for 21 and
24. This material is available free of charge via the Internet at
http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Author
*Phone: (858) 784-7522. Fax: (858) 784-7550. E-mail:
boger@scripps.edu.
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We gratefully acknowledge the financial support of the National
Institutes of Health (Grant CA041986), The Skaggs Institute
for Chemical Biology, and a grant from the Kansas Biotech
Authority. A.L.W. and K.K.D are Skaggs Fellows. A.L.W. is the
recipient of a predoctoral NSF fellowship (2009−2012).
ABBREVIATIONS USED
■
CBI, 1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4-one;
DPPA, diphenylphosphorylazide; LiHMDS, lithium bis-
(trimethylsilyl)amide; THF, tetrahydrofuran; BnSH, benzylth-
iol; BnOH, benzyl alcohol; BnNH2, benzylamine; EtOAc, ethyl
acetate; DMF, dimethylformamide; MeOH, methanol; Fmoc-
Cl, fluorenylmethyloxycarbonyl chloride
REFERENCES
■
(1) Ichimura, M.; Ogawa, T.; Takahashi, K.; Kobayashi, E.;
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K.; Kawamoto, I.; Tomita, F.; Nakano, H. Duocarmycin, a New
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1917.
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Studies. Acc. Chem. Res. 1995, 28, 20−29. (c) Boger, D. L.; Johnson, D.
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Boger, D. L. Fundamental Relationships between Structure, Reactivity,
and Biological Activity for the Duocarmycins and CC-1065. J. Med.
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Prodrugs? Curr. Pharm. Des. 2002, 8, 1375−1389. (h) Tse, W. C.;
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(b) Boger, D. L.; Coleman, R. S. Total Synthesis of CC-1065, and the
Precise, Functional Agent CPI-CDPI2. J. Am. Chem. Soc. 1988, 110,
4796−4807. (c) Boger, D. L.; Machiya, K. Total Synthesis of
(+)-Duocarmycin SA. J. Am. Chem. Soc. 1992, 114, 10056−10058.
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Total Synthesis and Preliminary Evaluation of (+)- and ent-
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one (CBI): An Enhanced and Simplified Analog of the CC-1065 and
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M.; van Asperen, J.; Henrar, R. E. C.; Beijnen, J. H. Comparative
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Tumors. Cancer Res. 1994, 54, 2404−2410. (d) Nagamura, S.; Kanda,
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1509. (f) Li, L. S.; Sinha, S. C. Studies toward the Duocarmycin
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Journal of Medicinal Chemistry
Article
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Synthesis of N-Phenylsulfonyl-CI, N-Boc-CI, CI-CDPI1 and CI-
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(20) Wolfe, A. L. Unpublished studies.
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