Design, synthesis and in vitro cytotoxic studies of novel bis-pyrrolo[2,1][1,4] benzodiazepine-pyrrole and imidazole polyamide conjugates

Article abstract of DOI:10.1016/j.ejmech.2005.02.005

The design, synthesis and biological evaluation of novel pyrrolo [2,1][1,4] benzodiazepine (PBD) dimers 38-43 linked with pyrrole and imidazole polyamides from either side by a flexible methylene chain of variable length are described, which involved merc

Full text of DOI:10.1016/j.ejmech.2005.02.005

European Journal of Medicinal Chemistry 40 (2005) 641–654
www.elsevier.com/locate/ejmech
Design, synthesis and in vitro cytotoxic studies of novel
bis-pyrrolo[2,1][1,4] benzodiazepine-pyrrole
and imidazole polyamide conjugates
Rohtash Kumar, J. William Lown *
Department of Chemistry, University of Alberta, Edmonton, Alta., Canada, T6G 2G2
Received 6 September 2004; accepted 22 February 2005
Available online 02 April 2005
Abstract
The design, synthesis and biological evaluation of novel pyrrolo [2,1][1,4] benzodiazepine (PBD) dimers 38–43 linked with pyrrole and
imidazole polyamides from either side by a flexible methylene chain of variable length are described, which involved mercuric chloride
mediated cyclization of the corresponding amino diethyl thioacetals. The compounds were prepared with varying numbers of pyrrole and
imidazole containing polyamides to determine the structural requirements for optimal in vitro antitumor activity. These compounds were
tested against a panel of 60 human cancer cells by the National Cancer Institute, and demonstrated that, of the compounds bis-PBD-pyrrole
polyamides (38–40) and bis-PBD-imidazole polyamides (41–43) certain of the bis-PBD-pyrrole and imidazole polyamide conjugates are
active for individual cancer cell lines (Table 1). However, this study found that bis-PBD-pyrrole and imidazole polyamide conjugates 38–43
in general are potent against many human cancer cell lines.
© 2005 Elsevier SAS. All rights reserved.
Keywords: Cytotoxicity; Polyamide; Pyrrolo [2,1-c] [1,4] benzodiazepines; DNA minor groove binders; PBD-polyamide conjugates
1. Introduction
principles responsible for the sequence-selective recognition
of DNA by small organic molecules [3] including a range of
naturally occurring antitumor antibiotics. Three fundamental
issues that arise in the examination of DNA binding agents
are: the origin of binding affinity, binding selectivity and reac-
tion selectivity including DNA alkylation or cleavage. Each
factor can independently assert levels of control on the
sequence-selective recognition of DNA and the relative role
and origin of these effects remain a primary objective of many
investigations. A powerful complement to such tools in the
examination of naturally derived DNA binding agents is the
DNA has for many years been a traditional target for che-
motherapeutic intervention [1] in human cancers, especially
for those where high proliferation rates of some tumor cell
types have resulted in sensitivity to drugs, which block rep-
lication and transcription of their DNA [2]. Substantial
progress has been made in understanding the fundamental
* Corresponding author. Tel.: +1 780 492 3646; fax: +1 780 492 8231.
E-mail address: Lynne.lechelt@ualberta.ca (J.W. Lown).
0223-5234/$ - see front matter © 2005 Elsevier SAS. All rights reserved.
doi:10.1016/j.ejmech.2005.02.005

642
R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
preparation and subsequent examination of key partial struc-
ture modifications or variations in the natural product and
their corresponding unnatural enantiomers.
date only a few PBD dimers have been prepared to examine
interstrand cross-linking of DNA, which are linked from two
possible positions by a flexible methylene chain of variable
length. To our knowledge no attempt has been made to syn-
thesize bis-pyrrolo [2,1-c][1,4] benzodiazepines (PBD)-
pyrrole and imidazole polyamide conjugates 38–43 (Bis-
PBDs dimers with pyrrole and imidazole polyamide
conjugates).
In view of the commonly observed activity of these PBDs
dimers and PBDs polyamides conjugates we attempted to con-
jugate two PBDs units with pyrrole and imidazole bearing
polyamide from either side by a flexible methylene chain of
variable length. In order to investigate the structure–activity
relationship systematically as well as their cytotoxicity against
human cancer cells, we herein describe the first synthesis and
report of the cytotoxic activity of novel bis-pyrrolo [2,1-
c][1,4] benzodiazepines (PBD)-pyrrole and imidazole polya-
mide conjugates 38–43 (Bis-PBDs dimers with pyrrole and
imidazole polyamide conjugates), which contain two PBDs
moieties linked from two different positions with pyrrole and
imidazole bearing polyamides by a flexible methylene chain
of variable length.
In addition, DNA sequence specificity or selectivity has
recently become recognized as an important component of
many cytotoxic agents [4,5] e.g. CC-1065 and duocarmycins
[6], saramycin [7], distamycin [8–10], netropsin [8–10], pyr-
rolo [1,4] benzodiazepinone [11], bleomycin [12,13], several
of which are of clinical interest in the treatment of human
malignancies. In this context PBDs (pyrrolo [2,1-c][1,4] ben-
zodiazepines), represent a group of exceptionally potent natu-
rally occurring antitumor antibiotics, derived from Strepto-
myces species [14]. Their interactions with DNA are unique
since they bind within the minor groove of DNA forming a
covalent aminal bond between the C11 position of the central
B-ring and the N2 amino group of a guanine base [14,15].
They differ in the number, type and position of substituent in
both their aromaticA-ring and pyrrolidine C-rings, and in the
degree of saturation of the C-rings which can be either fully
saturated or unsaturated at either the C2–C3 (endocyclic) or
C2 (exocyclic) positions. There is either an imine or carbino-
lamine methyl ether moiety at the N10–C11 position [16–19].
This latter is an electrophilic center responsible for alkylat-
ing DNA.
In addition studies on netropsin, distamycin and related
compounds have led to the concept of polyamides as infor-
mation reading agents [20]. A predominantly 4–5 AT base
pair sequence is recognized by netropsin and distamycin in
the minor groove of DNA. In our group attempts have been
made to link PBD with pyrrole [21,22] and imidazole [23,24]
and glycosylated pyrrole and imidazole polyamide [25], the
well-established DNA minor groove binders. It was found
that some PBD-glycosylated polyamides [25] conjugates
exhibit good cytotoxicity against different human cancer cells.
Studies have also shown that some synthetic compounds,
which contain two PBD moieties linked from two possible
positions by a flexible methylene chain of variable length,
are significantly more potent than PBD naturally occurring
compounds both in vitro and in vivo [26,27]. Recently a large
number of structurally modified PBDs compounds have also
been prepared and evaluated for their biological activity, par-
ticularly their antitumor potential [28,29]. The first PBD dimer
comprising two unsubstituted PBD units joined through their
C7–C7′ positions was reported by Farmer et al. [30,31] in
1988. Dimers with this linkage had only weak DNA cross-
linking activity and no cytotoxicity data were reported. The
first dimer with an C8–C8′ linkage was reported [32–34] in
1992. Dimers of this type have significant DNA interstrand
cross-linking activity, which forms a symmetric interstrand
cross link with duplex DNA involving a four base pairs bond-
ing site but spanning six base pairs overall [35] and pro-
nounced in vitro cytotoxicity and in vivo antitumor activity.
More recently, we have also reported C2–C2′ linked dimers
and their cytotoxicity [22,36].A number of these dimeric com-
pounds have been selected for preclinical studies but unfor-
tunately most of them did not proceed beyond that stage. To
2. Results and discussion
2.1. Synthesis
In our previous work the (2S)-N-[5-methoxy-4-[3-
(carboxy) propyloxy]-2-nitrobenzoyl] pyrrolidine-2-carbo-
xaldehyde diethyl thioacetal (1) was synthesized [22] by using
convenient routes in good yield. Condensation of the (2S)-N-
[5-methoxy-4-[3-(carboxy) propyloxy]-2-nitrobenzoyl]
pyrrolidine-2-carboxaldehyde diethyl thioacetal (1) with
[2-[N-(benzyloxycarbonyl) amino]ethyl]amine in the pres-
ence of EDCI and HOBt in dry DMF at room temperature
gave the [2-(4-{4-[2-(bis-ethylsulfanyl-methyl)-pyrrolidine-
1-carbonyl]-2-methoxy-5-nitro-phenoxy}-butyrylamino)-
ethyl]-carbamic acid benzyl ester (2) in 70% yield. Removal
of the CBZ group from this compound 2 with EtSH and
BF3·OEt2 gave the free amino N-(2-amino-ethyl)-4-{4-[2-
(bis-ethylsulfanyl-methyl)-pyrrolidine-1-carbonyl]-2-methoxy-
5-nitro-phenoxy}-butyramide (3) in 70% yield (Scheme 1).
Owing to the sensitivity of this amine to oxidation, it was
used for the next reaction immediately.
The (2S)-N-[5-methoxy-4-[3-(carboxy) propyloxy]-2-
nitrobenzoyl] pyrrolidine-2-carboxaldehyde diethyl thioac-
etal (1) was then coupled with the amine moiety of 1-methyl-
4-nitro-1H-pyrrole-2-carboxylic acid methyl ester (4), using
EDCI and HOBt as the coupling agents, in dry DMF at room
temperature for about 12 h to afford the corresponding
coupled compound 5 in 80% yield which, upon hydrolysis
with 0.5 N NaOH at room temperature then acidification, pro-
duced the corresponding acid 6 in 70% yield. The polyamide
acid 6 was treated with the N-(2-amino-ethyl)-4-{4-[2-(bis-
ethylsulfanyl-methyl)-pyrrolidine-1-carbonyl]-2-methoxy-5-

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
643
Schema 1
.
nitro-phenoxy}-butyramide (3) under standard EDCI, HOBt
coupling conditions and via its acid chloride (Scheme 2).
Unfortunately both reactions failed to produce the desired
products, due to the unreactive carboxyl group in the acid 6.
In that case we needed to increase the reactivity of the car-
boxyl group in the pyrrole and imidazole polyamide esters
by introducing a more electrophilic primary carboxylic group
through a suitable linker.
In our previous report the pyrrole and imidazole polya-
mide methyl esters 8–13 with a primary carboxylic group were
synthesized [37,38] by using convenient routes in good yield.
Condensation of the (2S)-N-[5-methoxy-4-[3-(carboxy)
propyloxy]-2-nitrobenzoyl] pyrrolidine-2-carboxaldehyde
diethyl thioacetal (1) with the amine moiety of pyrrole and
imidazole polyamide methyl esters 8–13, using EDCI and
HOBt as the coupling agents, in dry DMF afforded the cor-
responding coupled pyrrole and imidazole polyamide methyl
esters 20–25 in good yield which upon hydrolysis with 0.5 N
NaOH produced the corresponding coupled pyrrole and imi-
dazole polyamide acids 26–31 in good yield. The correspond-
ing amino compounds 14–19 were then prepared by hydro-
genation of the nitro polyamides 8–13. The reaction of free
amine N-(2-amino-ethyl)-4-{4-[2-(bis-ethylsulfanyl-methyl)-
pyrrolidine-1-carbonyl]-2-methoxy-5-nitro-phenoxy}-buty-
Schema 2
.

644
R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
Schema 3
.
ramide (3) with 1.0 equiv. of the nitro thio-PBD coupled pyr-
role and imidazole polyamide acids 26–31 using EDCI, HOBt
as the coupling agents in dry DMF at room temperature for
about 12 h, afforded the corresponding coupled bis nitro thio-
PBD-pyrrole and imidazole polyamides 32–37 in 70–80%
yield (Scheme 3). The nitro groups of compounds 32–37 were
reduced with hydrogen in the presence of Pd/C catalyst to
their corresponding respective amino compounds which were
then subjected to deprotective cyclization with HgCl2/HgO
in aqueous acetonitrile at room temperature affording the cor-
responding Bis-PBD-pyrrole and imidazole compounds
38–43 in 40–45% yield. Since the conjugation of the polya-
mides with PBD resulted in highly polar imines, this neces-
sitated the use of methanol in combination with dichlo-
romethane as eluent during the purification of the imines by
column chromatography and therefore the product was
obtained as a mixture of imine and its methyl ether in approxi-
mately a 1:1 ratio. NMR and mass spectra confirmed the pres-

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
645
ence of both forms. Since the methyl ether form is fully
equivalent to the imine form in terms of their reaction with
DNA, the compounds were isolated as the mixture of imines
and methyl ethers. This did not present any problems in evalu-
ating their biological activities. The final compounds were
isolated as pale yellow crystalline compounds in 40–45%
overall yields (Scheme 3).
breast cancer cell MDA-MB-435 (Log10 GI50 value
< –8.00).
Compound 40, having three pyrrole units linked between
two PBD moieties, shows notably higher cytotoxicity against
the renal cancer cells 786-O and CAKI-1 with Log10 GI50
value –6.25 and –7.00. This compound 40 has also shown
promising activity against non-small cell lung cancer cell NCI-
H522 with Log10 GI50 value –6.66. Its counterpart com-
pound 43 bearing three imidazole units linked between two
PBD moieties shows somewhat lower cytotoxic potency
against the same cell lines.
2.2. Anticancer cytotoxicity
This series of bis-pyrrolo [2,1][1,4] benzodiazepine-
pyrrole and imidazole compounds 38–43, which contain one
or more pyrrole and imidazole units for comparative pur-
poses, was selected by the US National Cancer Institute for
evaluation in the in vitro preclinical antitumor screening pro-
gram against sixty human tumor cell lines derived from nine
cancer types, leukemia, non-small cell lung cancer, colon can-
cer, CNS cancer, melanoma, ovarian cancer, renal cancer,
prostate cancer and breast cancer. For each compounds, dose
response curves for each cell line were measured at a mini-
mum of five concentrations at 10-fold dilutions in a protocol
of 48 h continuous drug exposure, and a sulfurhodamine B
(SRB) protein assay was used to estimate cell viability or
growth. The concentration causing 50% cell growth inhibi-
tion (GI50), total cell growth inhibition (TGI, 0% growth),
and 50% cell death (LC50, –50% growth) compared with the
control was calculated. The biological evaluation results for
the compounds 38–43 are presented in Table 1 as log10
GI50 values (the concentration of the drug resulting in inhi-
bition of cell growth to 50% of control).
It can be seen from the comparison of the cytotoxic data
presented in Table 1 for the compounds bis-PBD-pyrrole
polyamides (38–40) and bis-PBD-imidazole polyamides (41–
43) that certain of the bis-PBD-pyrrole and imidazole polya-
mide conjugates are active for individual cancer cell lines.
However, this study found that bis-PBD-pyrrole and imida-
zole polyamide conjugates 38–43 in general are potent against
many human cancer cell lines.
In summary, we have described the first synthesis of the
bis-PBD-pyrrole and imidazole polyamide conjugates 38–43
and also their anticancer evaluation against 60 human tumor
cell lines in nine cancer panels. More details of the biophysi-
cal, intracellular uptake and intracellular localization proper-
ties and additional biological evaluation will be reported in
due course.
2.3. Experimental
In general all the compounds are active against most of
the cell lines with the MG_MID (mean graph mid point) val-
ues ranging from –5.49 to –6.77 (activity is defined as a log10
GI50 of < 100 µM). Compound 38 which has one pyrrole unit
between two PBD units shows good cytotoxic activities
against non-small cell lung cancer cell line NCI-H522 (Log10
GI50 value < –8.00), colon cancer cell line SW-620 (Log10
GI50 value < –8.00). This compound 38 has also shown prom-
ising activity against leukemia cancer cells HL-60, K-562,
MOLT-4, RPMI-8226 and SR with Log10 GI50 values –6.26,
–7.34, –7.05, –6.48 and –7.29, while its counterpart com-
pound 41, which has one imidazole unit between two PBD
units, shows somewhat some better cytotoxicity against the
same cell lines. This compound 41 also shows higher cyto-
toxicity against renal cancer cells 786-O, ACHN, CAKI-1,
RXF 393, SN12C AND UO-31 with Log10 GI50 values rang-
ing from –7.84 to < –8.00, and breast cancer cell MCF7
(Log10 GI50 value < –8.00).
Compound 39, bearing two pyrrole units linked between
two PBD moieties displayed almost the same cytotoxicity with
the Log10 GI50 values ranging from –6.35 to –6.67 against
leukemia, non-small cell lung, ovarian, renal and breast can-
cer cells, cytotoxicity compared with the compound 42 which
has two imidazole unit linked between two PBD moieties
displays lower cytotoxicity against the same cell lines, in
addition compound 42 shows higher cytotoxicity against the
Kieselgel 60 (230–400 mesh) of E. Merck was used for
flash column chromatography, and precoated silica gel
GF-254 sheets of E-Merck were used for TLC, with the sol-
vent system indicated in the procedure. TLC plates were visu-
alized by using UV light. All compounds obtained commer-
cially were used without further purification unless otherwise
stated. Methanol and ethanol were freshly distilled over mag-
nesium turnings; tetrahydrofuran was distilled over sodium
benzophenone ketyl under an atmosphere of dry argon, ether
was dried over sodium; methylene chloride was freshly dis-
tilled from calcium hydride, triethylamine was treated with
potassium hydroxide then distilled from barium oxide and
stored over 3 Å molecular sieves. Dry dimethylformamide
and all commercially available chemicals were purchased
1
from Aldrich Chemical Co. The H NMR spectra were
recorded on a Bruker WH-300 spectrometer. Proton chemi-
cal shifts are reported in parts per million (d) downfield from
tetramethylsilane (SiMe4) as an internal standard. Coupling
constants (J values) are given in hertz and spin multiplicities
are described as follows: s (singlet), d (doublet), dd (doublet
of doublets), t (triplet), q (quartet), p (pentet) or m (multip-
let). FAB (fast atom bombardment) mass spectra with glyc-
erol as the matrix were determined on Associate Electrical
Ind. (AEI) MS-9 and MS-50 focusing high-resolution mass
spectrometers.

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R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
Table 1
In vitro cytotoxicity (log10GI50s) of novel bis-pyrrolo [2,1-c] [1,4] benzodiazepine-pyrrole and imidazole polyamide conjugates (38–43) against nine panels of
human cell lines
log10GI50(µM) a
38
39
40
41
42
43
Panels/cancer cell lines
Leukemia
CCRF-CEM
HL-60 (TB)
K-562
MOLT-4
RPMI-8226
SR
–
–6.60
–
–5.88
–6.31
–5.61
–6.52
–5.32
–
–5.67
–5.91
–6.46
–6.00
–6.90
–
–
–6.17
–
–5.85
–6.10
–5.64
–
–6.26
–7.34
–7.05
–6.48
–7.29
< –8.00
< –8.00
< –8.00
–6.61
–5.56
–5.53
–5.43
–5.63
< –8.00
Non-small cell lung cancer
A549/ATCC
EKVX
HOP-62
HOP-92
NCI-H226
NCI-H23
NCI-H322M
NCI-H460
NCI-H522
Colon cancer
COLO 205
HCC-2998
HCT-116
HCT-15
–4.56
–5.73
–4.68
–5.28
–
–4.98
–4.55
–4.53
< –8.00
–4.68
–
–5.47
–6.26
–4.69
–5.81
–
–5.83
–4.77
–5.62
–
–6.79
–5.87
–7.07
–4.70
–7.50
< –8.00
–4.71
< –8.00
< –8.00
–4.53
–
–4.68
–
–5.70
–5.16
–5.67
–5.69
–
–5.21
–5.41
–4.77
–5.55
–
–5.44
–5.50
–5.60
–5.60
–
–5.37
–6.67
–5.27
–
–5.46
–
–4.77
–
–5.02
–5.86
–5.79
–5.70
–5.61
–5.40
–5.73
–4.91
–5.67
–5.89
–6.14
–5.43
–4.97
–5.59
–5.51
–6.23
–5.90
–4.97
–4.95
–4.89
< –8.00
–5.29
–5.35
–5.79
–5.70
–5.58
–4.68
–5.35
–5.13
–5.53
–5.79
–5.61
–5.65
–4.73
–5.72
–5.22
–5.72
–5.42
–4.64
–7.52
HT29
KM12
SW-620
CNS cancer
268
SF-295
SF-539
SNB-19
–4.81
–4.51
–
–4.70
–5.34
–6.36
–5.77
–5.88
–5.38
–4.88
–5.39
–5.76
–5.29
–4.82
–5.55
–4.66
–4.42
–6.39
< –8.00
< –8.00
–6.64
–5.98
–5.47
–5.46
–5.74
–5.46
–4.80
–4.91
–5.59
–5.66
–5.65
–5.32
–4.83
–5.46
–5.67
SNB-75
U251
< –8.00
Melanoma
LOX IMVI
MALME-3M
M14
–6.92
–6.29
–5.11
–4.79
–5.14
–
–5.77
–
–5.80
–6.37
–5.84
–5.55
–5.45
–6.63
–5.85
–5.67
< –8.00
–5.87
–8.00
–5.54
–5.74
–8.00
–6.50
–7.27
–5.82
–
–5.75
–
–5.71
–4.92
–5.47
–4.57
–5.75
–
–5.32
–
–5.72
–5.53
–5.59
–5.49
–5.69
–5.46
–5.67
–5.63
–5.65
–5.78
SK-MEL-2
SK-MEL-28
SK-MEL-5
UACC-257
UACC-62
Ovarian cancer
IGROV 1
OVCAR-3
OVCAR-4
OVCAR-5
OVCAR-8
SK-OV-3
Renal cancer
786-0
–4.86
–4.85
–6.73
–4.37
–5.59
–
–5.39
–4.64
–5.64
–6.53
–5.74
–5.86
–5.46
–4.80
–6.62
–5.69
–5.92
–5.81
–5.72
–4.80
–5.41
–5.83
–4.78
–5.57
–6.07
–5.04
–
–
–5.83
–5.84
–5.48
–5.41
–4.71
–5.79
–5.79
–5.59
–5.47
–4.71
–5.99
–
–6.35
–4.67
–5.76
–6.64
–5.77
–5.79
–
–6.47
–4.68
–5.82
–6.14
–6.82
–5.85
–5.83
–6.58
< –8.00
–6.46
< –8.00
< –8.00
–7.65
< –8.00
–5.83
< –8.00
–6.40
–4.69
–5.75
–5.84
–5.69
–5.55
–
–6.11
–4.90
–5.74
–6.43
–5.75
–5.61
–
A498
ACHN
CAKI-1
RXF-393
SN12C
–5.62
–5.66
–5.49
–4.96
–5.51
–5.75
TK-10
UO-31
–5.69
–5.69
–5.56
(continued on next page)

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
647
Table 1
(continued)
log10GI50(µM) a
38
39
40
41
42
43
Panels/cancer cell lines
Prostate cancer
PC-3
–4.79
–5.27
–5.92
–5.32
–5.33
–5.84
–5.31
< –8.00
–5.21
–5.32
–5.76
–5.28
DU-145
Breast cancer
MCF 7
RES
MDA-MB231/ATCC
HS578T
MDA-MB-435
MDA-N
–5.13
–4.71
–6.07
–
–4.85
–
–5.90
–5.81
–5.77
–5.60
–
–6.15
–5.55
–6.43
–5.53
–5.66
–
< –8.00
–5.57
–5.39
–5.91
–7.31
–
–5.91
–5.53
–5.68
–5.10
< –8.00
–
–5.85
–5.68
–5.64
–5.48
–5.70
–
–
BT-549
T-47D
MGM b
–6.25
–4.71
–5.51
–5.39
–4.92
–5.69
–6.46
–5.23
–5.60
–6.24
–
–6.77
–5.36
–
–5.49
–5.35
–
–5.55
a The cytotoxicity log10GI50 values are the concentrations corresponding to 50% growth inhibition.
b Mean graph midpoint (µM) for growth inhibition against all human cancer cell lines tested.
2.3.1. [2-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitrophenoxy}-butyrylamino)ethyl]-
carbamic acid benzyl ester (2)
vent was removed under reduced pressure and the residue
was quenched with 5% NaHCO3 solution (50 ml) and then
extracted with ethyl acetate (3 × 50 ml). The combined organic
phase was dried over Na2SO4 the solvent was removed under
reduced pressure and the crude product was purified by flash
column chromatography on silica gel using 10% methanol/
dichloromethane as eluent to afford the free amine N-(2-
amino-ethyl)-4-{4-[2-(bis-ethylsulfanylmethyl)pyrrolidine-
1-carbonyl]-2-methoxy-5-nitrophenoxy}-butyramide (3) in
72% yield (1.60 g) as a yellow solid, after developing in 15%
methanol/dichloromethane, showed a single spot on TLC with
an Rf of 0.30, m.p. 95–98 °C. 1H NMR (300 MHz, DMSO-
d6) d 1.21 (s, 6H, –(CH3)2), 1.53–1.61 (m, 4H, –CH2CH–,
–CH2–), 1.95–2.18 (m, 4H, –CH2CO–, –CH2–), 2.48 (m, 4H,
2× –CH2CH3), 2.95 (m, 2H, –NH2CH2–), 3.32–3.48 (m, 4H,
–NHCH2–, –NCH2–), 3.73 (s, 3H, –OCH3), 3.89–3.94 (m,
3H, –OCH2–, –NCH–), 4.19 (m, 1H, –CHS2–), 5.58 (brs,
2H, –CH2NH2–), 7.61–7.77 (m, 2H, Ar–H), 8.10 (t, 1H,
–NHCH2–). HR-MS m/z Calcd. for C23H36N4O6S2 528.2402,
found 529.2402 (M + 1).
A
solution of [2-[N-(benzyloxycarbonyl)amino]-
ethyl]amine (1.43 g, 7.36 mmol) in dry DMF was added in to
mixture of the (2S)-N-[5-methoxy-4-[3-(carboxy)
a
propyloxy]-2-nitrobenzoyl] pyrrolidine-2-carboxaldehyde
diethyl thioacetal (1) (3.0 g, 6.16 mmol), hydroxybenzotria-
zole (0.833 g, 6.16 mmol), and EDCI (2.95 g, 15.38 mmol)
the reaction mixture was stirred at RT for 6 h and after comple-
tion of the reaction the solvent was removed under reduced
pressure to afford a dark oil which was purified by flash col-
umn chromatography on silica gel using 10% ethylacetate/
dichloromethane as eluent to afford the coupled [2-(4-{4-[2-
(bis-ethylsulfanylmethyl)pyrrolidine-1-carbonyl]-2-methoxy-
5-nitrophenoxy}-butyrylamino)ethyl] carbamic acid benzyl
ester (2) in 78% yield (3.20 g) as a yellow oil, after develop-
ing in 10% ethyacetate/dichloromethane, showed a single spot
on TLC with an Rf of 0.32. 1H NMR (300 MHz, DMSO-d6) d
1.20 (s, 6H, –(CH3)2), 1.56–1.60 (m, 4H, –CH2CH–, –CH2–),
1.95–2.18 (m, 4H, –CH2CO–, –CH2–), 2.50 (m, 4H, 2×
–CH2CH3), 3.22–3.47 (m, 6H, 2× –NHCH2–, –NCH2–), 3.75
(s, 3H, –OCH3), 3.86–3.94 (m, 3H, –OCH2–, –NCH–), 4.20
(m, 1H, –CHS2–), 5.35 (s, 2H, –CH2OCO–), 7.20 (m, 5H,
Ar–H), 7.61–7.75 (m, 2H, Ar–H), 8.10 (m, 2H, 2×
–NHCH2–). HR-MS m/z Calcd. for C31H42N4O8S2 662.2405,
found 663.2806 (M + 1).
2.4. General procedure A
A solution of the nitropolyamide methyl esters (pyrrole or
imidazole) in MeOH or DMF was hydrogenated over 10%
Pd/C at 50 psi pressure for 2 h and the catalyst was removed
by filtration through a Celite pad. The filtrate was concen-
trated to dryness under reduced pressure (at RT) to afford the
corresponding amine. Owing to the sensitivity of the amine
to oxidation, it was used for the next reaction immediately. It
was dissolved in dry DMF and a mixture of the (2S)-N-[5-
methoxy-4-[3-(carboxy) propyloxy]-2-nitrobenzoyl] pyrro-
lidine-2-carboxaldehyde diethyl thioacetal (1) (1.0 equiv.),
hydroxybenzotriazole (1.0 equiv.), and EDCI (2.5 equiv.), in
dry DMF was added. This mixture was stirred at RT for 12 h
and after completion of the reaction the solvent was removed
under reduced pressure to afford a dark oil which was puri-
fied by flash column chromatography on silica gel by using
2.3.2. N-(2-Amino-ethyl)-4-{4-[2-(bis-ethylsulfanylmeth-
yl)pyrrolidine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-
butyramide (3)
To a stirred solution of EtSH (8.21 g) and BF3·OEt2 (9.60 g)
in dry dichloromethane was added dropwise compound [2-(4-
{4-[2-(bis-ethylsulfanylmethyl) pyrrolidine-1-carbonyl]-2-
methoxy-5-nitrophenoxy}butyrylamino)ethyl]carbamic acid
benzyl ester (2) (2.8 g, 4.22 mmol) in dry dichloromethane at
room temperature. Stirring was continued until reaction was
completed by TLC. After completion of the reaction the sol-

648
R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
methanol–dichloromethane as eluent to afford the PBD-nitro
dithioacetal pyrrole or imidazole polyamide methyl esters as
a yellow solid in good yield.
4.20 (m, 1H, –CHS2–), 6.89 (d, 1H, J = 1.8 Hz, Py–H), 7.10
(d, 1H, J = 1.8 Hz, Py–H), 7.61 (s, 1H, Ar–H), 7.78 (s, 1H,
Ar–H), 9.94 (s, 1H, –NH–), 11.10 (brs, 1H, –COOH). HR-MS
m/z Calcd. for C27H36N4O8S2 608.1905; found 609.2005 (M
+ 1).
2.4.1. 4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)1-
methyl-1H-pyrrole-2-carboxylic acid methyl ester (5)
This compound was prepared starting from 1-methyl-4-
nitro-1H-pyrrole-2-carboxylic acid methyl ester (0.415 g,
2.25 mmol) and (2S)-N-[5-methoxy-4-[3-(carboxy) propy-
loxy]-2-nitrobenzoyl] pyrrolidine-2-carboxaldehyde diethyl
thioacetal (1) (1.0 g, 2.05 mmol), according to the general
procedure A as a yellow solid in 78% yield (1.0 g), after devel-
oping in 5% methanol/dichloromethane, showed a single spot
on TLC with an Rf of 0.34. 1H NMR (DMSO-d6, 300 MHz):
d 1.24 (s, 6H, –(CH3)2), 1.56–1.61(m,4H, –CH2CH–,
–CH2–), 1.95–2.18 (m, 4H, –CH2CO–, –CH2–), 2.50 (m, 4H,
2× –CH2CH3), 3.37 (m, 2H, –NCH2), 3.62 (s, 3H, –NCH3),
3.74 (s, 3H, –OCH3), 3.85 (s, 3H, –COOCH3), 3.90–3.96 (m,
3H, –OCH2–, –NCH–), 4.21 (m, 1H, –CHS2–), 6.71 (d, 1H,
J = 1.8 Hz, Py–H), 6.96 (d, 1H, J = 1.8 Hz, Py–H), 7.61 (s,
1H, Ar–H), 7.78 (s, 1H, Ar–H), 9.95 (s, 1H, –NH–). HR-MS
m/z Calcd. for C28H38N4O8S2 622.2107; found 623.2105 (M
+ 1).
2.5.2. {[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)-1-
methyl-1H-pyrrole-2-carbonyl]-amino}-acetic acid methyl
ester (20)
This compound was prepared starting from compound 8
(0.55 g, 2.28 mmol) and (2S)-N-[5-methoxy-4-[3-(carboxy)
propyloxy]-2-nitrobenzoyl] pyrrolidine-2-carboxaldehyde
diethyl thioacetal (1) (1.0 g, 2.05 mmol), according to the
general procedure A (1.0 g, 72% yield) as a yellow solid, it
showed a single spot on TLC with an Rf of 0.34, after devel-
oping in 5% methanol/dichloromethane. 1H NMR (DMSO-
d6, 300 MHz): d 1.21 (s, 6H, –(CH3)2), 1.55–1.62 (m, 4H,
–CH2CH–, –CH2–), 1.99–2.20 (m, 4H, –CH2CO–, –CH2–),
2.48 (m, 4H, 2× –CH2CH3), 3.35 (m, 2H, –NCH2), 3.60 (s,
3H, –NCH3), 3.71 (s, 3H, –COOCH3), 3.80 (s, 3H, –OCH3),
3.89–3.96 (m, 5H, –NCH–, –NHCH2, –OCH2–), 4.20 (m, 1H,
–CHS2–), 6.82 (d, 1H, J = 1.8 Hz, Py–H), 7.10 (d, 1H,
J = 1.8 Hz, Py–H), 7.62 (s, 1H, Ar–H), 7.77 (s, 1H, Ar–H),
8.20 (t, 1H, J = 6.5 Hz, –NHCH2–), 9.96 (s, 1H, –NH–).
HR-MS m/z Calcd. for C30H41N5O9S2 679.2345; found
638.2380 (M + 1).
2.5. General procedure B
2.5.3. [(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitro phenoxy}-butyryl-
amino)-1-methyl-1H-pyrrole-2-carbonyl]-amino}-1-methyl-
1H-pyrrole-2-carbonyl)-amino]-acetic acid methyl ester
(21)
A mixture of PBD-nitro dithioacetal pyrrole or imidazole
polyamide methyl esters in methanol and 10 ml of 0.5 N
NaOH was placed in a flask, then the reaction mixture was
stirred at room temperature until the ester completely disap-
peared as shown by TLC. The reaction was cooled in ice with
stirring and neutralized with 0.5 N HCl slowly to pH 2. The
reaction mixture was extracted with pure ethyl acetate or ethyl
acetate/THF (1:1) three times and dried over sodium sulfate
and the solvent was removed under reduced pressure. The
residue was purified by column chromatography using
MeOH/dichloromethane as eluent to afford the PBD-nitro
dithioacetal pyrrole or imidazole polyamide acids in good
yield.
This compound was prepared starting from compound 9
(0.820 g, 2.25 mmol) and the acid 1 (1.0 g, 2.05 mmol) accord-
ing to the general procedure A (1.2 g, 73% yield) as a yellow
solid, after developing in 5% methanol/dichloromethane,
1
showed a single spot on TLC with an Rf of 0.28. H NMR
(DMSO-d6, 300 MHz): d 1.20 (s, 6H, –(CH3)2), 1.56–1.62
(m,4H, –CH2CH–, –CH2–), 1.95–2.18 (m, 4H, –CH2CO–,
–CH2–), 2.48 (m, 4H, 2× –CH2CH3), 3.37 (m, 2H, –NCH2),
3.62 (s, 3H, –NCH3), 3.69 (s, 3H, –NCH3), 3.73 (s, 3H,
–COOCH3), 3.82 (s, 3H, –OCH3), 3.89–3.96 (m, 5H, –NCH–,
–NHCH2, –OCH2–), 4.19 (m, 1H, –CHS2–), 6.82 (d, 1H,
J = 1.8 Hz, Py–H), 6.96 (d, 1H, J = 1.8 Hz, Py–H), 7.08 (d,
1H, J = 1.8 Hz, Py–H), 7.40 (d, 1H, J = 1.8 Hz, Py–H), 7.63
(s, 1H, Ar–H), 7.75 (s, 1H, Ar–H), 8.18 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.94 (s, 1H, –NH–), 9.98 (s, 1H, –NH–). HR-MS
m/z Calcd. for C36H47N7O10S2 801.2826; found 802.2852 (M
+ 1).
2.5.1. 4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)1-
methyl-1H-pyrrole-2-carboxylic acid (6)
This compound was prepared according to the general pro-
cedure
ethylsulfanylmethyl)pyrrolidine-1-carbonyl]-2-methoxy-5-
nitrophenoxy}-butyryl amino)-1-methyl-1H-pyrrole-2-
B by employing compound 4-(4-{4-[2-(bis-
carboxylic acid methyl ester (5) (0.8 g, 1.28 mmol) and 0.5 N
NaOH in 83% yield (0.65 g) as a yellow solid, after develop-
ing in 10% methanol/dichloromethane, showed a single spot
on TLC with an Rf of 0.30. 1H NMR (DMSO-d6, 300 MHz):
d 1.22 (s, 6H, –(CH3)2), 1.55–1.62 (m, 4H, –CH2CH–,
–CH2–), 1.95–2.18 (m, 4H, –CH2CO–, –CH2–), 2.50 (m, 4H,
2× –CH2CH3), 3.37 (m, 2H, –NCH2), 3.65 (s, 3H, –NCH3),
3.76 (s, 3H, –OCH3), 3.89–3.95 (m, 3H, –OCH2–, –NCH–),
2.5.4. [4-(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-butyryl-
amino)-1-methyl-1H-pyrrole-2-carbonyl]-amino}-1-methyl-
1H-pyrrole-2-carbonyl)-amino]-1-methyl-1H-pyrrole-2-
carbonyl)-amino]-acetic acid methyl ester (22)
Prepared according to the general procedure A by using
compound 10 (1.09 g, 2.25 mmol) and the (2S)-N-[5-

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
649
methoxy-4-[3-(carboxy) propyloxy]-2-nitrobenzoyl] pyr-
rolidine-2-carboxaldehyde diethyl thioacetal (1) (1.0 g,
2.05 mmol) in 74% yield (1.4 g) as a yellow solid, showed a
single spot on TLC with an Rf of 0.31, after developing in 7%
methanol/dichloromethane. 1H NMR (DMSO-d6, 300 MHz):
d 1.22 (s, 6H, –(CH3)2), 1.55–1.67 (m, 4H, –CH2CH–,
–CH2–), 1.93–2.18 (m, 4H, –CH2CO–, –CH2–), 2.49 (m, 4H,
2× –CH2CH3), 3.35 (m, 2H, –NCH2), 3.61 (s, 3H, –NCH3),
3.66 (s, 3H, –NCH3), 3.70 (s, 3H, –NCH3), 3.75 (s, 3H,
–COOCH3), 3.82 (s, 3H, –OCH3), 3.89–3.96 (m, 5H, –NCH–,
–NHCH2, –OCH2–), 4.22 (m, 1H, –CHS2–), 6.81 (d, 1H,
J = 1.8 Hz, Py–H), 6.96 (d, 1H, J = 1.8 Hz, Py–H), 7.08 (d,
1H, J = 1.8 Hz, Py–H), 7.20 (d, 1H, J = 1.8 Hz, Py–H), 7.35
(d, 1H, J = 1.8 Hz, Py–H), 7.55 (d, 1H, J = 1.8 Hz, Py–H),
7.62 (s, 1H,Ar–H), 7.76 (s, 1H,Ar–H), 8.20 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.92 (s, 1H, –NH–), 9.95 (s, 1H, –NH–), 9.98 (s,
1H, –NH–). HR-MS m/z Calcd. for C42H53N9O11S2 923.3306;
found 924.3340 (M + 1).
5H, –NCH–, –NHCH2, –OCH2–), 4.20 (m, 1H, –CHS2–),
7.20 (s, 1H, Im–H), 7.45 (s, 1H, Im–H), 7.63 (s, 1H, Ar–H),
7.75 (s, 1H, Ar–H), 8.18 (t, 1H, J = 6.5 Hz, –NHCH2–), 9.96
(s, 1H, –NH–), 9.98 (s, 1H, –NH–). HR-MS m/z Calcd. for
C34H45N9O10S2 803.2731; found 804.2764 (M + 1).
2.5.7. [4-(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-butyryl-
amino)-1-methyl-1H-imidazole-2-carbonyl]-amino}-1-
methyl-1H-imidazole-2-carbonyl)-amino]-1-methyl-1H-
imidazole-2-carbonyl)-amino]-acetic acid methyl ester
(25)
This compound was prepared according to the method
described for the compound 20 by employing compound 13
(1.10 g, 2.25 mmol) and the (2S)-N-[5-methoxy-4-[3-
(carboxy) propyloxy]-2-nitrobenzoyl] pyrrolidine-2-car-
boxaldehyde diethyl thioacetal (1) (1.10 g, 2.05 mmol) in 76%
yield (1.45 g) as a yellow solid, it showed a single spot on
TLC with an Rf of 0.31, after developing in 10% methanol/
2.5.5. {[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)-1-
methyl-1H-imidazole-2-carbonyl]-amino}-acetic acid
methyl ester (23)
This compound was prepared according to the method
described for the compound 20 by employing compound 11
(0.547 g, 2.25 mmol) and the (2S)-N-[5-methoxy-4-[3-
1
dichloromethane with few drops of aq. ammonia. H NMR
(DMSO-d6, 300 MHz): d 1.22 (s, 6H, –(CH3)2), 1.56–1.67
(m, 4H, –CH2CH–, –CH2–), 1.95–2.20 (m, 4H, –CH2CO–,
–CH2–), 2.49 (m, 4H, 2× –CH2CH3), 3.36 (m, 2H, –NCH2),
3.63 (s, 3H, –NCH3), 3.65 (s, 3H, –NCH3), 3.69 (s, 3H,
–NCH3), 3.74 (s, 3H, –COOCH3), 3.83 (s, 3H, –OCH3), 3.89–
3.96 (m, 5H, –NCH–, –NHCH2, –OCH2–), 4.18 (m, 1H,
–CHS2–), 7.10 (s, 1H, Im–H), 7.38 (s, 1H, Im–H), 7.56 (s,
1H, Im–H), 7.64 (s, 1H, Ar–H), 7.77 (s, 1H, Ar–H), 8.20 (t,
1H, J = 6.5 Hz, –NHCH2–), 9.95 (s, 1H, –NH–), 9.97 (s, 1H,
–NH–), 10.00 (s, 1H, –NH–). HR-MS m/z Calcd. for
C39H50N12O11S2 926.3162; found 927.3198 (M + 1).
(carboxy)
propyloxy]-2-nitrobenzoyl]
pyrrolidine-2-
carboxaldehyde diethyl thioacetal (1) (1.0 g, 2.05 mmol) in
82% yield (1.15 g) as a yellow solid, showed a single spot on
TLC with an Rf of 0.31, after developing in 5% methanol/
1
dichloromethane and with two drops of aq. ammonia. H
NMR (DMSO-d6, 300 MHz): d 1.21 (s, 6H, –(CH3)2), 1.55–
1.66 (m, 4H, –CH2CH–, –CH2–), 1.95–2.19 (m, 4H,
–CH2CO–, –CH2–), 2.45 (m, 4H, 2× –CH2CH3), 3.37 (m,
2H, –NCH2), 3.66 (s, 3H, –NCH3), 3.73 (s, 3H, –COOCH3),
3.83 (s, 3H, –OCH3), 3.89–3.98 (m, 5H, –NCH–, –NHCH2,
–OCH2–), 4.19 (m, 1H, –CHS2–), 7.37 (s, 1H, Im–H), 7.64
(s, 1H, Ar–H), 7.76 (s, 1H, Ar–H), 8.19 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.97 (s, 1H, –NH–). HR-MS m/z Calcd. for
C29H40N6O9S2 680.2295; found 681.2332 (M + 1).
2.5.8. {[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)-1-
methyl-1H-pyrrole-2-carbonyl]-amino}-acetic acid (26)
This compound was prepared starting from {[4-(4-{4-[2-
(bis-ethylsulfanylmethyl)pyrrolidine-1-carbonyl]-2-methoxy-
5-nitrophenoxy}-butyrylamino)-1-methyl-1H-pyrrole-2-
carbonyl]-amino}-acetic acid methyl ester (20) (0.8 g,
1.17 mmol) and 0.5 N NaOH (1.5 ml), according to the gen-
eral procedure B (1.0 g, 80% yield) as a yellow solid in 89%
yield (0.7 g), after developing in 10% methanol/
dichloromethane, showed a single spot on TLC with an Rf of
2.5.6. [(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-butyryl-
amino)-1-methyl-1H-imidazole-2-carbonyl]-amino}-1-
methyl-1H-imidazole-2-carbonyl)-amino]-acetic acid
methyl ester (24)
This compound was prepared starting from compound 12
(0.825 g, 2.25 mmol) and the acid 1 (1.0 g, 2.05 mmol) accord-
ing to the general procedure A (1.2 g, 72% yield) as a yellow
solid, after developing in 7% methanol/dichloromethane with
few drops of aq. ammonia, showed a single spot on TLC with
an Rf of 0.30. 1H NMR (DMSO-d6, 300 MHz): d 1.20 (s, 6H,
–(CH3)2), 1.55–1.66 (m, 4H, –CH2CH–, –CH2–), 1.94–2.18
(m, 4H, –CH2CO–, –CH2–), 2.48 (m, 4H, 2× –CH2CH3), 3.35
(m, 2H, –NCH2), 3.64 (s, 3H, –NCH3), 3.68 (s, 3H, –NCH3),
3.73 (s, 3H, –COOCH3), 3.82 (s, 3H, –OCH3), 3.89–3.95 (m,
1
0.30, m.p. 160–162 °C. H NMR (DMSO-d6, 300 MHz): d
1.20 (s, 6H, –(CH3)2), 1.55–1.60 (m,4H, –CH2CH–, –CH2–),
1.99–2.18 (m, 4H, –CH2CO–, –CH2–), 2.47 (m, 4H, 2×
–CH2CH3), 3.36 (m, 2H, –NCH2), 3.62 (s, 3H, –NCH3), 3.78
(s, 3H, –OCH3), 3.89–3.96 (m, 5H, –NCH–, –NHCH2,
–OCH2–), 4.20 (m, 1H, –CHS2–), 6.80 (d, 1H, J = 1.8 Hz,
Py–H), 7.15 (d, 1H, J = 1.8 Hz, Py–H), 7.63 (s, 1H, Ar–H),
7.75 (s, 1H, Ar–H), 8.18 (t, 1H, J = 6.5 Hz, –NHCH2–), 9.96
(s, 1H, –NH–), 10.80 (brs, 1H, –COOH). HR-MS m/z Calcd.
for C29H39N5O9S2 665.2189; found 666.2220 (M + 1).

650
R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
2.5.9. [(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-
butyrylamino)-1-methyl-1H-pyrrole-2-carbonyl]-amino}-1-
methyl-1H-pyrrole-2-carbonyl)-amino]-acetic acid (27)
2.5.11. {[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro phenoxy}-butyrylamino)-1-
methyl-1H-imidazole-2-carbonyl]-amino}-acetic acid (29)
This compound was prepared according to the method
described for the compound 28 by employing compound
{[4-(4-{4-[2-(bis-ethylsulfanylmethyl)pyrrolidine-1-
This compound was prepared starting from compound
[(4-{[4-(4-{4-[2-(bis-ethylsulfanylmethyl)-pyrrolidine-1-
carbonyl]-2-methoxy-5-nitrophenoxy}-butyrylamino)-1-
methyl-1H-pyrrole-2-carbonyl]-amino}-1-methyl-1H-pyr-
role-2-carbonyl)-amino]-acetic acid methyl ester (21) (1.0 g,
1.24 mmol) and the 0.5 N NaOH (1.5 ml) according to the
general procedure B (0.85 g, 86% yield) as a yellow solid,
showed a single spot on TLC with an Rf of 0.28, after devel-
oping in 10% methanol/dichloromethane, m.p. 175–178 °C.
1H NMR (DMSO-d6, 300 MHz): d 1.21 (s, 6H, –(CH3)2),
1.55–1.62 (m,4H, –CH2CH–, –CH2–), 1.99–2.20 (m, 4H,
–CH2CO–, –CH2–), 2.48 (m, 4H, 2× –CH2CH3), 3.35 (m,
2H, –NCH2), 3.65 (s, 3H, –NCH3), 3.69 (s, 3H, –NCH3), 3.80
(s, 3H, –OCH3), 3.89–3.96 (m, 5H, –NCH–, –NHCH2,
–OCH2–), 4.20 (m, 1H, –CHS2–), 6.84 (d, 1H, J = 1.8 Hz,
Py–H), 6.96 (d, 1H, J = 1.8 Hz, Py–H), 7.10 (d, 1H,
J = 1.8 Hz, Py–H), 7.35 (d, 1H, J = 1.8 Hz, Py–H), 7.62 (s,
1H, Ar–H), 7.72 (s, 1H, Ar–H), 8.18 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.95 (s, 1H, –NH–), 9.98 (s, 1H, –NH–), 10.80
(brs, 1H, –COOH),. HR-MS m/z Calcd. for C35H45N7O10S2
787.2669; found 788.2705 (M + 1).
carbonyl]-2-methoxy-5-nitro-phenoxy}-butyrylamino)-1-
methyl-1H-imidazole-2-carbonyl]-amino}-acetic acid methyl
ester (23) (1.0 g, 1.47 mmol) and the 0.5 N NaOH solution
(1.5 ml) in 87% yield (0.85 g) as a yellow solid, it showed a
single spot on TLC with an Rf of 0.28, after developing in
10% methanol/dichloromethane with few drops of aq. ammo-
1
nia, m.p. 152–155 °C. H NMR (DMSO-d6, 300 MHz): d
1.20 (s, 6H, –(CH3)2), 1.55–1.65 (m, 4H, –CH2CH–, –CH2–),
1.95–2.18 (m, 4H, –CH2CO–, –CH2–), 2.48 (m, 4H, 2×
–CH2CH3), 3.35 (m, 2H, –NCH2), 3.67 (s, 3H, –NCH3), 3.83
(s, 3H, –OCH3), 3.89–3.99 (m, 5H, –NCH–, –NHCH2,
–OCH2–), 4.20 (m, 1H, –CHS2–), 7.40 (s, 1H, Im–H), 7.65
(s, 1H, Ar–H), 7.77 (s, 1H, Ar–H), 8.19 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.97 (s, 1H, –NH–), 10.80 (brs, 1H, –COOH).
HR-MS m/z Calcd. for C28H38N6O9S2 666.2142; found
667.2176 (M + 1).
2.5.12. [(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitro phenoxy}-
butyrylamino)-1-methyl-1H-imidazole-2-carbonyl]-amino}-
1-methyl-1H-imidazole-2-carbonyl)-amino]-acetic acid
(30)
2.5.10. [4-(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-
butyrylamino)-1-methyl-1H-pyrrole-2-carbonyl]-amino}-1-
methyl-1H-pyrrole-2-carbonyl)-amino]-1-methyl-1H-
pyrrole-2-carbonyl)-amino]-acetic acid (28)
This compound was prepared starting from compound
[(4-{[4-(4-{4-[2-(bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitro-phenoxy}-butyrylamino)-1-
methyl-1H-imidazole-2-carbonyl]-amino}-1-methyl-1H-
imidazole-2-carbonyl)-amino]-acetic acid methyl ester (24)
(1.1 g, 1.36 mmol) and 0.5 N NaOH solution (2.0 ml) accord-
ing to the general procedure B (0.92 g, 85% yield) as a yel-
low solid, after developing in 12% methanol/dichloromethane
with few drops of aq. ammonia, it showed a single spot on
TLC with an Rf of 0.35, m.p. 165–167 °C. 1H NMR (DMSO-
d6, 300 MHz): d 1.21 (s, 6H, –(CH3)2), 1.55–1.66 (m, 4H,
–CH2CH–, –CH2–), 1.97–2.18 (m, 4H, –CH2CO–, –CH2–),
2.48 (m, 4H, 2× –CH2CH3), 3.36 (m, 2H, –NCH2), 3.64 (s,
3H, –NCH3), 3.68 (s, 3H, –NCH3), 3.84 (s, 3H, –OCH3),
3.89–3.98 (m, 5H, –NCH–, –NHCH2, –OCH2–), 4.18 (m, 1H,
–CHS2–), 7.35 (s, 1H, Im–H), 7.45 (s, 1H, Im–H), 7.62 (s,
1H, Ar–H), 7.77 (s, 1H, Ar–H), 8.20 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.96 (s, 1H, –NH–), 9.98 (s, 1H, –NH–), 10.81
(s, 1H, –COOH). HR-MS m/z Calcd. for C33H43N9O10S2
789.2574; found 790.2605 (M + 1).
Prepared according to the general procedure B by using
compound [4-(4-{[4-(4-{4-[2-(bis-ethylsulfanylmethyl)-
pyrrolidine-1-carbonyl]-2-methoxy-5-nitro-phenoxy}-buty-
rylamino)-1-methyl-1H-pyrrole-2-carbonyl]-amino}-1-
methyl-1H-pyrrole-2-carbonyl)-amino]-1-methyl-1H-pyrrole-
2-carbonyl)-amino]-acetic acid methyl ester (22). (1.8 g,
1.94 mmol) and the 0.5 N NaOH (2.0 ml) in 84% yield (1.5 g)
as a yellow solid, after developing in 12% methanol/
dichloromethane, showed a single spot on TLC with an Rf of
1
0.30, m.p. 188–190 °C. H NMR (DMSO-d6, 300 MHz): d
1.20 (s, 6H, –(CH3)2), 1.55–1.68 (m, 4H, –CH2CH–, –CH2–),
1.96–2.18 (m, 4H, –CH2CO–, –CH2–), 2.49 (m, 4H, 2×
–CH2CH3), 3.36 (m, 2H, –NCH2), 3.62 (s, 3H, –NCH3), 3.65
(s, 3H, –NCH3), 3.69 (s, 3H, –NCH3), 3.80 (s, 3H, –OCH3),
3.89–3.95 (m, 5H, –NCH–, –NHCH2, –OCH2–), 4.18 (m, 1H,
–CHS2–), 6.82 (d, 1H, J = 1.8 Hz, Py–H), 6.96 (d, 1H,
J = 1.8 Hz, Py–H), 7.10 (d, 1H, J = 1.8 Hz, Py–H), 7.21 (d,
1H, J = 1.8 Hz, Py–H), 7.36 (d, 1H, J = 1.8 Hz, Py–H), 7.56
(d, 1H, J = 1.8 Hz, Py–H), 7.64 (s, 1H, Ar–H), 7.77 (s, 1H,
Ar–H), 8.20 (t, 1H, J = 6.5 Hz, –NHCH2–), 9.94 (s, 1H,
–NH–), 9.96 (s, 1H, –NH–), 9.98 (s, 1H, –NH–), 10.80 (s,
1H, –COOH). HR-MS m/z Calcd. for C41H51N9O11S2
909.3149; found 910.3183 (M + 1).
2.5.13. [4-(4-{[4-(4-{4-[2-(Bis-ethylsulfanylmethyl)pyrroli-
dine-1-carbonyl]-2-methoxy-5-nitrophenoxy}-butyryl-
amino)-1-methyl-1H-imidazole-2-carbonyl]-amino}-1-
methyl-1H-imidazole-2-carbonyl)-amino]-1-methyl-1H-
imidazole-2-carbonyl)-amino]-acetic acid (31)
This compound was prepared according to the method
described for the compounds 28 by employing compound

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
651
[4-(4-{[4-(4-{4-[2-(bis-ethylsulfanylmethyl)pyrrolidine-1-
carbonyl]-2-methoxy-5-nitrophenoxy}-butyryl amino)-1-
methyl-1H-imidazole-2-carbonyl]-amino}-1-methyl-1H-
imidazole-2-carbonyl)-amino]-1-methyl-1H-imidazole-2-
carbonyl)-amino]-acetic acid methyl ester (25) (1.4 g,
1.51 mmol) and 0.5 N NaOH solution (2.0 ml) in 87% yield
(1.20 g) as a yellow solid, showed a single spot on TLC with
an Rf of 0.28, after developing in 12% methanol/
dichloromethane with few drops of aq. ammonia, m.p. 179–
acetal pyrrole polyamide acid 27 (0.8 g, 1.01 mmol) and the
PBD-nitro dithoacetal amine 3 (0.59 g, 1.11 mmol) in 76%
yield (1.0 g) as a yellow color solid, it showed a single spot
on TLC with an Rf of 0.37, after developing in 5%
methanol/dichloromethane. 1H NMR (DMSO-d6, 300 MHz):
d 1.21 (s, 12H, 4× –CH3), 1.56–1.60 (m, 8H, 2× –CH2CH–,
2× –CH2–), 1.96–2.20 (m, 8H, 2× –CH2CO–, 2× –CH2–),
2.47 (m, 8H, 4× –CH2CH3), 3.36 (m, 4H, 2× –NCH2), 3.48
(m, 4H, 2× –NHCH2), 3.62 (s, 3H, –NCH3), 3.66 (s, 3H,
–NCH3), 3.75 (s, 3H, –OCH3), 3.76 (s, 3H, –OCH3), 3.86–
3.99 (m, 8H, –NHCH2, 2× –NCH–, 2× –OCH2–), 4.18 (m,
2H, 2× –CHS2–), 6.84 (d, 1H, J = 1.8 Hz, Py–H), 6.96 (d,
1H, J = 1.8 Hz, Py–H), 7.08 (d, 1H, J = 1.8 Hz, Py–H), 7.25
(d, 1H, J = 1.8 Hz, Py–H), 7.62 (s, 2H, Ar–H), 7.77 (s, 2H,
Ar–H), 8.10–8.19 (m, 3H, 3× –NHCH2), 9.94 (s, 1H, –NH–),
9.98 (s, 1H, –NH–). MS m/z Calcd. for C58H79N11O15S4
1297.4640 found 1298.4673 (M + 1).
1
182 °C. H NMR (DMSO-d6, 300 MHz): d 1.20 (s, 6H,
–(CH3)2), 1.55–1.67 (m, 4H, –CH2CH–, –CH2–), 1.95–2.18
(m, 4H, –CH2CO–, –CH2–), 2.48 (m, 4H, 2× –CH2CH3), 3.34
(m, 2H, –NCH2), 3.63 (s, 3H, –NCH3), 3.65 (s, 3H, –NCH3),
3.69 (s, 3H, –NCH3), 3.83 (s, 3H, –OCH3), 3.89–3.99 (m,
5H, –NCH–, –NHCH2, –OCH2–), 4.19 (m, 1H, –CHS2–),
7.28 (s, 1H, Im–H), 7.38 (s, 1H, Im–H), 7.56 (s, 1H, Im–H),
7.64 (s, 1H,Ar–H), 7.78 (s, 1H,Ar–H), 8.19 (t, 1H, J = 6.5 Hz,
–NHCH2–), 9.95 (s, 1H, –NH–), 9.97 (s, 1H, –NH–), 10.08
(s, 1H, –NH–), 10.80 (s, 1H, –COOH). HR-MS m/z Calcd.
for C38H48N12O11S2 912.3007; found 913.3040 (M + 1).
2.6.3. Compound-34
This compound was prepared starting from PBD-nitro
dithioacetal amine 3 (0.70 g, 1.32 mmol) and the acid 28
(1.10 g, 1.20 mmol) according to the general procedure
described for compound 32 as a yellow solid (1.25 g, 73%
yield), after developing in 5% methanol/dichloromethane, it
2.6. General procedure C
To a solution of PBD-nitro dithioacetal pyrrole or imida-
zole polyamide acids (20–22 or 23–25) in dry DMF (20 ml)
were added EDCI (2.5 equiv.), HOBt (1.0 equiv.), and N-(2-
amino-ethyl)-4-{4-[2-(bis-ethylsulfanyl-methyl)-pyrrolidine-
1-carbonyl]-2-methoxy-5-nitro-phenoxy}-butyramide (3)
(1.1 equiv.) under a nitrogen atmosphere and the mixture was
stirred for 12 h. When TLC indicated the absence of starting
material, DMF was removed under reduced pressure. The dark
residue was purified by column chromatography on silica gel
using MeOH/dichloromethane as eluent to afford the coupled
Bis-PBD-nitro dithioacetal pyrrole or imidazole polyamide
conjugates (32–34 or 35–37) as yellow solids in good yields.
1
showed a single spot on TLC with an Rf of 0.32. H NMR
(DMSO-d6, 300 MHz): d 1.20 (s, 12H, 4× –CH3), 1.55–1.61
(m, 8H, 2× –CH2CH–, 2× –CH2–), 1.95–2.18 (m, 8H, 2×
–CH2CO–, 2× –CH2–), 2.48 (m, 8H, 4× –CH2CH3), 3.35
(m, 4H, 2× –NCH2), 3.46 (m, 4H, 2× –NHCH2), 3.62 (s, 3H,
–NCH3), 3.64 (s, 3H, –NCH3), 3.68 (s, 3H, –NCH3), 3.73 (s,
3H, –OCH3), 3.75 (s, 3H, –OCH3), 3.85–3.99 (m, 8H,
–NHCH2, 2× –NCH–, 2× –OCH2–), 4.19 (m, 2H, 2×
–CHS2–), 6.81 (d, 1H, J = 1.8 Hz, Py–H), 6.86 (d, 1H,
J = 1.8 Hz, Py–H), 6.96 (d, 1H, J = 1.8 Hz, Py–H), 7.10 (d,
1H, J = 1.8 Hz, Py–H), 7.28 (d, 1H, J = 1.8 Hz, Py–H), 7.55
(d, 1H, J = 1.8 Hz, Py–H), 7.63 (s, 2H, Ar–H), 7.76 (s, 2H,
Ar–H), 8.12–8.19 (m, 3H, 3× –NHCH2), 9.94 (s, 1H, –NH–),
9.96 (s, 1H, –NH–), 9.98 (s, 1H, –NH–). MS m/z Calcd. for
C64H85N13O16S4 1419.5120; found 1420.5152 (M + 1).
2.6.1. Compound-32
Prepared according to the general procedure C by using
compound 26 (0.75 g, 1.12 mmol) and PBD-nitro dithoacetal
amine 3 (0.655 g, 1.23 mmol) in 75% yield (1.0 g) as a yel-
low solid, after developing in 5% methanol/dichloromethane,
1
2.6.4. Compound-35
showed a single spot on TLC with an Rf of 0.40. H NMR
(DMSO-d6, 300 MHz): d 1.20 (s, 12H, 4× –CH3), 1.55–1.61
(m, 8H, 2× –CH2CH–, 2× –CH2–), 1.98–2.18 (m, 8H, 2×
–CH2CO–, 2× –CH2–), 2.48 (m, 8H, 4× –CH2CH3), 3.35
(m, 4H, 2× –NCH2), 3.46 (m, 4H, 2× –NHCH2), 3.60 (s, 3H,
–NCH3), 3.75 (s, 3H, –OCH3), 3.76 (s, 3H, –OCH3), 3.85–
3.95 (m, 8H, –NHCH2, 2× –NCH-, 2× –OCH2–), 4.19 (m,
2H, 2× –CHS2–), 6.82 (d, 1H, J = 1.8 Hz, Py–H), 7.08 (d,
1H, J = 1.8 Hz, Py–H), 7.63 (s, 2H,Ar–H), 7.76 (s, 2H,Ar–H),
8.12–8.20 (m, 3H, 3× –NHCH2), 9.95 (s, 1H, –NH–). MS
m/z Calcd. for C52H73N9O14 S4 1175.4160; found 1176.4195
(M + 1).
This compound was prepared starting from PBD-nitro
dithioacetal amine 3 (0.70 g, 1.32 mmol) and the PBD-nitro
dithioacetal imidazole polyamide acid 29 (0.80 g, 1.20 mmol)
according to the general procedure C (1.0 g, 71% yield) as a
yellow solid, after developing in 5% methanol/dichloro-
methane with few drops of aq. ammonia, it showed a single
1
spot on TLC with an Rf of 0.41. H NMR (DMSO-d6,
300 MHz): d 1.20 (s, 12H, 4× –CH3), 1.55–1.62 (m, 8H, 2×
–CH2CH–, 2× –CH2–), 1.96–2.18 (m, 8H, 2× –CH2CO–,
2× –CH2–), 2.47 (m, 8H, 4× –CH2CH3), 3.36 (m, 4H, 2×
–NCH2), 3.45 (m, 4H, 2× –NHCH2), 3.65 (s, 3H, –NCH3),
3.74 (s, 3H, –OCH3), 3.76 (s, 3H, –OCH3), 3.85–3.99 (m,
8H, –NHCH2, 2× –NCH–, 2× –OCH2–), 4.19 (m, 2H, 2×
–CHS2–), 7.45 (s, 1H, Im–H), 7.62 (s, 2H, Ar–H), 7.76 (s,
2H, Ar–H), 8.15–8.20 (m, 3H, 3× –NHCH2), 9.95 (s, 1H,
2.6.2. Compound-33
This compound was prepared according to the method
described for the compound 32, employing PBD-nitro dithio-

652
R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
–NH–). MS m/z Calcd. for C51H72N10O14S4 1176.4112; found
1177.4148 (M + 1).
umn and first eluted with ethyl acetate to remove HgCl2.After
complete removal of HgCl2, the column was eluted with
CHCl3/MeOH system by which all other impurities were
removed. Then the column was further eluted with
CHCl3/MeOH/ammonia and the ammonia concentration was
slowly increased from 0.5% through 2%. The imine com-
pounds were collected at different percentage of ammonia
from 0.5% to 2%. The imine and corresponding methyl ether
move together as they have close Rf values. Evaporation of
the solvents under high vacuum, at RT, afforded an insepa-
rable mixture of imines and methyl ethers 38–43 in almost
1:1 ratio in 40–45% yield.
2.6.5. Compound-36
This compound was prepared according to the method
described for the compound 35, employing PBD-nitro dithio-
acetal imidazole polyamide acid 30 (0.85 g, 1.07 mmol) and
the PBD-nitro dithioacetal amine 3 (0.625 g, 1.18 mmol) in
72% yield (1.0 g) as a yellow solid, showed a single spot
on TLC with an Rf of 0.38, after developing in 5%
methanol/dichloromethane with few drops of aq. ammonia.
1H NMR (DMSO-d6, 300 MHz): d 1.21 (s, 12H, 4× –CH3),
1.55–1.64 (m, 8H, 2× –CH2CH–, 2× –CH2–), 1.95–2.19 (m,
8H, 2× –CH2CO–, 2× –CH2–), 2.48 (m, 8H, 4× –CH2CH3),
3.35 (m, 4H, 2× –NCH2), 3.46 (m, 4H, 2× –NHCH2), 3.64
(s, 3H, –NCH3), 3.67 (s, 3H, –NCH3), 3.73 (s, 3H, –OCH3),
3.75 (s, 3H, –OCH3), 3.85–3.98 (m, 8H, –NHCH2, 2×
–NCH–, 2× –OCH2–), 4.18 (m, 2H, 2× –CHS2–), 7.37 (s,
1H, Im–H), 7.48 (s, 1H, Im–H), 7.64 (s, 2H, Ar–H), 7.75 (s,
2H, Ar–H), 8.15–8.20 (m, 3H, 3× –NHCH2), 9.96 (s, 1H,
–NH–), 9.98 (s, 1H, –NH–). MS m/z Calcd. for
C56H77N13O15S4 1299.4545; found 1300.4578 (M + 1).
2.7.1. Compound-38
This compound was prepared according to the general pro-
cedure D by using compound 32 (0.9 g, 0.76 mmol) in 45%
1
yield (0.30 g) as a yellow solid after purification. H NMR
(DMSO-d6, 300 MHz): d 1.56–1.61 (m, 8H, 2× –CH2CH–,
2× –CH2–), 1.98–2.20 (m, 8H, 2× –CH2–, 2× –CH2CO–),
3.30–3.37 (m, 6H, 2× –NCH, 2× –NCH2), 3.45 (m, 4H, 2×
–NHCH2), 3.64 (s, 3H, –NCH3), 3.73 (s, 3H, –OCH3), 3.75
(s, 3H, –OCH3), 3.84 (m, 6H, –OCH3 for its methyl ether),
3.89–3.98 (m, 6H, –NHCH2, 2× –OCH2–), 6.80 (s, 2H,
Ar–H), 6.92 (d, 1H, J = 1.8 Hz, Py–H), 7.30 (s, 2H, Ar–H),
7.40 (d, 1H, J = 1.8 Hz, Py–H), 7.56 (m, 2H, imine proton),
8.12–8.18 (m, 3H, 3× –NHCH2), 9.96 (s, 1H, –NH–). MS
m/z Calcd. for C44H53N9O10 867.3915; found 868.3940 (M +
1) for its imine compound and 932.4460 (M + 1) for its methyl
ether compound. Anal. Calcd. for C44H53N9O10: C, 60.89;
H, 6.15; N, 14.52; found: C, 60.82; H, 6.19; N, 14.58.
2.6.6. Compound-37
Prepared according to the general procedure C using com-
pound 31 (1.0 g, 1.09 mmol) and PBD-nitro dithioacetal
amine 3 (0.636 g, 1.20 mmol) in 77% yield (1.2 g) as a yel-
low solid, after developing in 5% methanol/dichloromethane
with few drops aq. ammonia, showed a single spot on TLC
with an Rf of 0.32. 1H NMR (DMSO-d6, 300 MHz): d 1.20
(s, 12H, 4× –CH3), 1.56–1.66 (m, 8H, 2× –CH2CH–, 2×
–CH2–), 1.96-2.20 (m, 8H, 2× –CH2CO–, 2× –CH2–), 2.47
(m, 8H, 4× –CH2CH3), 3.36 (m, 4H, 2× –NCH2), 3.45 (m,
4H, 2× –NHCH2), 3.63 (s, 3H, –NCH3), 3.65 (s, 3H, –NCH3),
3.67 (s, 3H, –NCH3), 3.75 (s, 3H, –OCH3), 3.77 (s, 3H,
–OCH3), 3.86–3.98 (m, 8H, –NHCH2, 2× –NCH–, 2×
–OCH2–), 4.20 (m, 2H, 2× –CHS2–), 7.28 (s, 1H, Im–H),
7.35 (s, 1H, Im–H), 7.46 (s, 1H, Im–H), 7.62 (s, 2H, Ar–H),
7.77 (s, 2H, Ar–H), 8.15-8.20 (m, 3H, 3× –NHCH2), 9.95 (s,
1H, –NH–), 9.97 (s, 1H, –NH–), 9.99 (s, 1H, –NH–). MS m/z
Calcd. for C61H82N16O16S4 1422.4978; found 1423.5010 (M
+ 1).
2.7.2. Compound-39
This compound was prepared starting from compound 33
(0.95 g, 0.732 mmol) according to the general procedure D
1
(0.3 g, 41% yield) as a yellow solid. H NMR (DMSO-d6,
300 MHz): d 1.55–1.60 (m, 8H, 2× –CH2CH–, 2× –CH2–),
1.99–2.18 (m, 8H, 2× –CH2–, 2× –CH2CO–), 3.31–3.35 (m,
6H, 2× –NCH, 2× –NCH2), 3.46 (m, 4H, 2× –NHCH2), 3.63
(s, 3H, –NCH3), 3.66 (s, 3H, –NCH3), 3.74 (s, 3H, –OCH3),
3.75 (s, 3H, –OCH3), 3.85 (m, 6H, –OCH3 for its methyl
ether), 3.89–3.98 (m, 6H, –NHCH2, 2× –OCH2–), 6.81 (s,
2H, Ar–H), 6.89 (d, 1H, J = 1.8 Hz, Py–H), 6.96 (d, 1H,
J = 1.8 Hz, Py–H), 7.10 (d, 1H, J = 1.8 Hz, Py–H), 7.30 (s,
2H,Ar–H), 7.48 (d, 1H, J = 1.8 Hz, Py–H), 7.56 (m, 2H, imine
proton), 8.12–8.20 (m, 3H, 3× –NHCH2), 9.94 (s, 1H, –NH–),
9.96 (s, 1H, –NH–). MS m/z Calcd. for C50H59N11O11
989.4396; found 990.4430 (M + 1) for its imine compound
and 1054.4953 (M + 1) for its methyl ether compound. Anal.
Calcd. for C50H59N11O11: C, 60.66; H, 6.01; N, 15.56; found:
C, 60.70; H, 6.05; N, 15.51.
2.7. General procedure D
Nitrodithioacetal pyrrole and imidazole polyamides 32–37
were dissolved in methanol (50.0 ml), respectively, and hydro-
genated over 10% Pd–C at 50 psi pressure for 2 h and then
the catalyst was removed by filtration through a Celite pad.
The filtrate was concentrated to dryness under reduced pres-
sure. The resultant amino compounds were dissolved in
CH3CN/H2O (4:1) and HgCl2 (1.5 equiv.) and HgO
(1.5 equiv.) were added and the mixture was stirred slowly at
RT for 12 h. When TLC (CHCl3/MeOH/ammonia) indicated
the complete disappearance of starting materials, the reac-
tion mixtures were charged directly on to a short silica col-
2.7.3. Compound-40
This compound was prepared according to the general pro-
cedure D by using compound 34 (1.2 g, 0.845 mmol) in 42%
1
yield (0.40 g) as a yellow solid. H NMR (DMSO-d6,
300 MHz): d 1.59–1.62 (m, 8H, 2× –CH2CH–, 2× –CH2–),

R. Kumar, J.W. Lown / European Journal of Medicinal Chemistry 40 (2005) 641–654
653
1.98–2.19 (m, 8H, 2× –CH2–, 2× –CH2CO–), 3.30–3.36 (m,
6H, 2× –NCH, 2× –NCH2), 3.45 (m, 4H, 2× –NHCH2), 3.63
(s, 3H, –NCH3), 3.65 (s, 3H, –NCH3), 3.67 (s, 3H, –NCH3),
3.74 (s, 3H, –OCH3), 3.76 (s, 3H, –OCH3), 3.84 (m, 6H,
–OCH3 for its methyl ether), 3.89–3.95 (m, 6H, –NHCH2,
2× –OCH2–), 6.80 (s, 2H, Ar–H), 6.84 (d, 1H, J = 1.8 Hz,
Py–H), 6.89 (d, 1H, J = 1.8 Hz, Py–H), 6.96 (d, 1H,
J = 1.8 Hz, Py–H), 7.10 (d, 1H, J = 1.8 Hz, Py–H), 7.28 (d,
1H, J = 1.8 Hz, Py–H), 7.30 (s, 2H, Ar–H), 7.50 (d, 1H,
J = 1.8 Hz, Py–H), 7.55 (m, 2H, imine proton), 8.15–8.20
(m, 3H, 3× –NHCH2), 9.95 (s, 1H, –NH–), 9.97 (s, 1H,
–NH–), 9.99 (s, 1H, –NH–). MS m/z Calcd. for C56H65N13O12
1111.4876; found 1112.4910 (M + 1) for its imine com-
pound and 1176.5430 (M + 1) for its methyl ether com-
pound. Anal. Calcd. for C56H65N13O12: C, 60.47; H, 5.89; N,
16.37; found: C, 60.40; H, 5.92; N, 16.32.
(1.1 g, 0.773 mmol) in 41% yield (0.35 g) as a yellow solid.
1H NMR (DMSO-d6, 300 MHz): d 1.55–1.62 (m, 8H, 2×
–CH2CH–, 2× –CH2–), 1.98–2.19 (m, 8H, 2× –CH2–, 2×
–CH2CO–), 3.30–3.36 (m, 6H, 2× –NCH, 2× –NCH2), 3.45
(m, 4H, 2× –NHCH2), 3.62 (s, 3H, –NCH3), 3.65 (s, 3H,
–NCH3), 3.68 (s, 3H, –NCH3), 3.74 (s, 3H, –OCH3), 3.76 (s,
3H, –OCH3), 3.85 (m, 6H, –OCH3 for its methyl ether), 3.89–
3.99 (m, 6H, –NHCH2, 2× –OCH2–), 6.80 (s, 2H, Ar–H),
7.22 (s, 1H, Im–H), 7.33 (s, 2H, Ar–H), 7.44 (s, 1H, Im–H),
7.51 (s, 1H, Im–H), 7.55 (m, 2H, imine proton), 8.15–8.22
(m, 3H, 3× –NHCH2), 9.94 (s, 1H, –NH–), 9.96 (s, 1H,
–NH–), 9.99 (s, 1H, –NH–). MS m/z Calcd. for C53H62N16O12
1114.4733; found 1115.4767 (M + 1) for its imine com-
pound and 1179.5290 (M + 1) for its methyl ether com-
pound. Anal. Calcd. for C53H62N16O12: C, 57.08; H, 5.60; N,
20.10; found: C, 57.12; H, 5.62; N, 20.08.
2.7.4. Compound-41
This compound was prepared according to the method
described for the compound 39, by employing compound 35
(0.95 g, 0.807 mmol) in 42% yield (0.30 g) as a yellow solid.
1H NMR (DMSO-d6, 300 MHz): d 1.55–1.60 (m, 8H, 2×
–CH2CH–, 2× –CH2–), 1.98–2.18 (m, 8H, 2× –CH2–, 2×
–CH2CO–), 3.31–3.35 (m, 6H, 2× –NCH, 2× –NCH2), 3.46
(m, 4H, 2× –NHCH2), 3.63 (s, 3H, –NCH3), 3.73 (s, 3H,
–OCH3), 3.75 (s, 3H, –OCH3), 3.85 (m, 6H, –OCH3 for its
methyl ether), 3.88–3.98 (m, 6H, –NHCH2, 2× –OCH2–),
6.81 (s, 2H, Ar–H), 7.32 (s, 2H, Ar–H), 7.40 (s, 1H, Ar–H),
7.56 (m, 2H, imine proton), 8.12–8.18 (m, 3H, 3× –NHCH2),
9.97 (s, 1H, –NH–). MS m/z Calcd. for C43H52N10O10
868.3868; found 869.3904 (M + 1) for its imine compound
and 933.4428 (M + 1) for its methyl ether compound. Anal.
Calcd. for C43H52N10O10: C, 59.44; H, 6.03; N, 16.12; found:
C, 59.40; H, 6.08; N, 16.16.
Acknowledgements
We thank the National Cancer Institute, Maryland for the
anticancer assay in human cell lines. This research was sup-
ported by a grant (to J.W.L.) from the Natural Sciences and
Engineering Research Council of Canada.
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654
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