Synthesis and antitumour activity of pyrene-linked pyrrolo [2,1-c][1,4]benzodiazepine hybrids

Article abstract of DOI:10.1016/j.bmcl.2003.10.050

Pyrene-linked pyrrolobenzodiazepine hybrids have been synthesized that exhibit potential anticancer activity in a number of human tumour cell lines. These hybrids also exhibit much enhanced DNA-binding ability in comparison to the parent pyrrolobenzodiazepine ring system (DC-81).

Full text of DOI:10.1016/j.bmcl.2003.10.050

Bioorganic & Medicinal Chemistry Letters 14 (2004) 471–474
Synthesis and antitumour activity of pyrene-linked pyrrolo
[2,1-c][1,4]benzodiazepine hybrids
Ahmed Kamal,* G. Ramesh, O. Srinivas and P. Ramulu
Biotransformation Laboratory, Division of Organic Chemistry, Indian Institute of Chemical Technology, Hyderabad 500007, India
Received 7 August 2003; accepted 23 October 2003
Abstract—Pyrene-linked pyrrolobenzodiazepine hybrids have been synthesized that exhibit potential anticancer activity in a num-
ber of human tumour cell lines. These hybrids also exhibit much enhanced DNA-binding ability in comparison to the parent pyr-
rolobenzodiazepine ring system (DC-81).
# 2003 Elsevier Ltd. All rights reserved.
The pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) are
naturally occurring antitumour antibiotics isolated from
Streptomyces species.1 These compounds are known to
exhibit antitumour activity based on their covalent
binding ability tothe N2 of the guanine residues in the
minor groove of DNA. These properties have been
investigated in detail for such compounds that form
covalent adducts in the minor groove of DNA and show
sequence specificity for GC-rich DNA regions, in parti-
cular for Pu-G-pu triplets.2 In recent years, a number of
PBD based symmetrical and unsymmetrical DNA
cross-linking agents have been designed and synthesized
that exhibit significant DNA binding and anticancer
activity.3 In our studies, we have investigated the non-
cross linking PBD dimers that exhibit remarkable DNA
binding in the absence of cross linking.4 In this context
many PBD conjugates have been synthesized and
investigated for their anticancer activity.5 A number of
polyaromatic hydrocarbons (PAH) and their derivatives
in view of their planar ring system are known to inter-
calate with DNA resulting in the anticancer activity.6
Some of the polyaromatic hydrocarbons (PAH) based
on pyrene and chrysene derivatives like (1-pyr-
enylmethyl)amino alcohol derivatives 1, crisnatol 2
(770U82) have undergone different stages of clinical
trials.7 Recently, we have synthesized some hybrid
molecules 3 wherein chrysene is linked toPBD at C8
position through an alkylamide spacer.8 It has been
envisaged that such hybrids of PBD that are linked to
polyaromatic hydrocarbons (PAH) may produce
enhanced anticancer activity. Our interest in the struc-
tural modifications and development of new synthetic
strategies on the PBD ring system9 has lead tothe
design and synthesis of pyrene-linked PBD hybrids as
potential DNA binding anticancer agents.10
Synthesis of these pyrene-linked PBD hybrids has been
carried out by employing aminopyrene 6 as one of the
starting materials and this has been obtained by the
.
reduction of the nitropyrene 5 by SnCl2 2H2O in EtOAc
(Scheme 1). Whereas, (2S)-N-(4-benzyloxy-5-methoxy-
2-nitro-benzoyl)pyrrolidine-2-carboxaldehyde diethyl-
thioacetal 7 has been prepared by literature method,11
which upon debenzylation gives 8. Etherification of (2S)
-N-(4-hydroxy-5-methoxy-2-nitrobenzoyl)pyrrolidine-2-
carboxaldehyde diethylthioacetal 8 by methyl bromo-
alkanoates provides 9a–b. Basic hydrolysis of these
esters 9a–b give the desired precursor acids 10a–b. The
key intermediates 11a–b have been prepared by amida-
tion of 6 with the acid precursors 10a–b. Further, these
upon reduction and followed by deprotection of ami-
nothioacetal precursors 11a–b afford the target pyrene-
linked PBD hybrids 4a–b (Scheme 2).12
Amongst these hybrids 4a has shown promising anti-
cancer activity in 60-cell line panel. The LC50 values of
compound 4a against leukemia (SR), non-small cell
lung cancer (NCI-H226), colon (HCC-2298), CNS (SF-
539 and SNB-19), melanoma (M14 and A498) and
prostate (DU-145) have been illustrated in Table 1. The
in vitro cytotoxicity (IC50) for the naturally occurring
Keywords: Pyrrolobenzodiazepines; Pyrene amine; Cytotoxity; DNA-
binding affinity.
* Corresponding author. Tel.: +91-40-2719-3157; fax: +91-40-2719-
3189; e-mail: ahmedkamal@iict.ap.nic.in
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.10.050

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A. Kamal et al. / Bioorg. Med. Chem. Lett. 14 (2004) 471–474
DC-8113 is 0.38 and 0.33 mM in L1210 and PC6 cell
lines, respectively. It is interestingly toboserve that
compound 4a has promising anticancer activity in
number of cancer cell lines.
The DNA binding ability for these pyrene-linked
PBD hybrids has been examined by thermal dena-
turation studies using calf thymus (CT) DNA. Inter-
estingly, in this assay one of the pyrene-linked PBD
hybrid (4a) elevates the helix melting temperature of
CT-DNA by a 6.5 ꢀC after incubation for 18 h at
37 ꢀC. On the other hand, the naturally occurring
DC-81 exhibits a ÁTm of 0.7 ꢀC. This demonstrates
that hybrid 4a has significant DNA binding affinity as
illustrated in Table 2. This increase of melting tempera-
ture could be attributed to the additional effect of
intercalation with DNA by the pyrene subunit apart
from the covalent binding that may take place by the
PBD ring system.
.
Scheme 1. (i) SnCl2 2H2O, EtOAc, 3 h, 76%.
Table 1. In vitro cytotoxicity of compound 4a in selected human
cancer cell lines
Cancer panel/cell line
LC50 (mM)
Leukemia
SR
0.05
<0.01
0.01
Non-small cell lung
NCI-H226
Colon
HCC-2298
CNS
SF-539
SNB-19
0.18
0.42
In conclusion, one of the new pyrene-linked PBD
hybrid (4a) not only exhibits promising cytotoxic activ-
ity in a number of cancer cell lines but also shows good
DNA-binding ability. The detailed biological and
molecular modeling studies of chrysene and pyrene
linked PBD hybrids are in progress.
Melanoma
M14
A498
0.05
0.04
Prostate
DU-145
0.24
Table 2. Thermal denaturation data for pyrene-linked PBD hybrids with CT-DNA
PBD hybrids
[PBD]/[DNA] molar ratiob
ÁTm (ꢀC)a after incubation at 37 ꢀC fo r
18 h
0 h
4a
4b
DC-81
1:5
1:5
1:5
6.1
3.6
0.3
6.5
3.8
0.7
a For CT-DNA alone at pH 7.00Æ0.01, Tm=69.2 ꢀCÆ0.01 (mean value from 10 separate determinations), all ÁTm values are Æ0.1–0.2 ꢀC.
b For a 1:5 molar ratio of [PBD]/[DNA], where CT-DNA concentration=100 mM and ligand concentration=20 mM in aqueous sodium phosphate
buffer (10 mM sodium phosphate+1 mM EDTA, pH 7.00Æ0.01).

A. Kamal et al. / Bioorg. Med. Chem. Lett. 14 (2004) 471–474
473
Scheme 2. (i) EtSH–BF3OEt2, CH2Cl2, 12 h, rt, 75%; (ii) methyl bromoalkanoate, K2CO3, DMF, 24 h, rt, 89–91%; (iii) 1 N LiOH, THF–MeOH–
.
H2O, 12 h, rt, 74–78%; (iv) isobutyl chloroformate, Et3N, compound 6, 12 h, rt, 55–56%; (v) SnCl2 2H2O, EtOAc, reflux, 3 h, 70–72%; (vi) HgCl2–
CaCO3, CH3CN–H2O, 12 h, rt, 49–54%.
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Siim, B. G.; Wu, K.; Pullen, S. M.; Botting, K. J.; Wilson,
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vas, O.; Neelima, K.; Kondapi, A. K.; Srinu, V. B.;
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Lown, W. J. Anti-Cancer Drug Design 2000, 15, 225. (b)
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Acknowledgements
We thank the National Cancer Institute, Maryland,
USA for the in vitro anticancer assay in human cancer
cell lines. We are alsograteful toCSIR, New Delhi for
the award of research fellowships to G.R., O.S. and
P.R.
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