Synthesis and DNA-binding affinity of A-C8/C-C2 alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers

Article abstract of DOI:10.1016/j.bmcl.2003.08.071

The synthesis of new A-C8/C-C2 alkoxyamido-linked pyrrolo[2,1-c][1,4]- benzodiazepine dimers have been described in this report. These dimers exhibit significant DNA-binding ability with moderate anticancer activity.

Full text of DOI:10.1016/j.bmcl.2003.08.071

Bioorganic & Medicinal Chemistry Letters 13 (2003) 3955–3958
Synthesis and DNA-Binding Affinity of A-C8/C-C2 Alkoxyamido-
Linked Pyrrolo[2,1-c][1,4]benzodiazepine Dimers
Ahmed Kamal,* P. Ramulu, O. Srinivas and G. Ramesh
Division of Organic Chemistry, Indian Institute of Chemical Technology, Hyderabad 500007, India
Received 16 July 2003; accepted 28 August 2003
Abstract—The synthesis of new A-C8/C-C2 alkoxyamido-linked pyrrolo[2,1-c][1,4]-benzodiazepine dimers have been described in
this report. These dimers exhibit significant DNA-binding ability with moderate anticancer activity.
# 2003 Elsevier Ltd. All rights reserved.
DNA interstrand cross-linking agents that interact
within the minor groove constitute an important class of
antitumour drugs.1 There has been considerable interest
in the past few years in the design and synthesis of
symmetrical cross-linking agents, particularly based on
pyrrolobenzodiazepines (PBDs).2 In the literature a
number of PBD dimers have been designed and synthe-
sized that have exhibited varying degrees of cytotoxicity
and DNA cross-linking activity.3 These PBD dimers
have been joined through their different positions such
as A-C7/A-C70, A-C8/A-C80 and C-C2/C-C20, among
these A-C8/A-C8- linked PBD dimers have shown pro-
mising cytotoxicity and efficient cross-linking property.
Recently, Thurston and co-workers4 have reported the
first examples of A-C8/C-C2 amido-linked PBD dimers
with marginal cross-linking ability. In continuation of
our efforts in the design and synthesis of PBD
hybrids,5 we have been interested in exploring anti-
cancer potential and DNA-binding ability of A-C8/C-
C2 alkoxyamido-linked PBD dimers. Further, we have
also synthesized these PBD dimers with an amide
functionality at N10-C11 of C-C2 component to unra-
vel the aspect of non-covalent interactions in such
dimers.
the precursor (2S)-N-(4-hydroxy-5-methoxy-2-nitro-
benzoyl)pyrrolidine-2-carboxaldehyde diethyl thioacetal
3 has been prepared by employing the literature method,6
which upon etherification with methyl bromoalkanoates
provides 4a–c upon basic hydrolysis of these esters to give
the desired intermediate acids 5a–c (Scheme 1).
However, the C-C2 amino component has been prepared
as described below: trans-4-hydroxy-l-proline methyl-
ester hydrochloride has been coupled to the compound
6 to give the nitro ester 7. The C2 hydroxy groupis
protected with TBDMS-Cl followed by reduction with
DIBAL-H to produce the corresponding aldehyde,
which is protected with EtSH/TMS-Cl. Surprisingly, in
this reaction protection of aldehyde to diethyl thioacetal
and deprotection of TBDMS takes place in the same
stepto afford the compound 10. Mesylation upon C2
hydroxy groupgives compound
11. Azidation at C2
position proceeds through SN2 reaction mechanism to
afford compound 12, followed by reduction to give
compound 13.
The key intermediates 14a–c have been prepared by
coupling of compound 5a–c and compound 13. Further,
.
these upon reduction by SnCl2 2H2O in methanol and
These PBD dimers have been synthesized by preparing
the A-C8 acid component (5) and the C-C2 amino sub-
stituted component (13) individually and followed by
the coupling of these two precursors. The C8 alkoxy
acid has been synthesized in the following manner;
followed by deprotection of diaminothioacetal pre-
cursors 15a–c using HgCl2/CaCO3 affords the PBD
dimers 1a–c7 (Scheme 2).
The other key intermediates 20a–c have been prepared
by coupling compounds 5a–c with compound 19. Fur-
ther, these upon reduction gives aminothioacetal pre-
cursors 21a–c followed by deprotection to afford the
desired PBD dimers 2a–c8 (Scheme 3).
*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.2003.08.071

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A. Kamal et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3955–3958
Scheme 1. (i) Methyl bromoalkanoate, K2CO3, DMF, rt, 24 h, 90%; (ii) 1 N LiOH, THF–MeOH–H2O, 12 h, rt, 78%.
Scheme 2. (i) SOCl2, C6H6, trans-4-hydroxyprolinemethylesterhydrochloride, H2O, 2 h, rt, 75%; (ii) TBDMSCl, imidazole, CH2Cl2, 8 h, rt, 80%;
(iii) DIBAL-H, CH2Cl2, 1 h, À78 ꢀC, 70%; (iv) EtSH, TMSCl, CH2Cl2, 8 h, rt, 62%; (v) TEA, MsCl, CH2Cl2, 5 h, 0 ꢀC, 78%; (vi) NaN3, DMF,
60 ꢀC, 16 h, 76%; (vii) TPP, THF–NH4OH, 8 h, rt, 72%; (viii) HOBt, EDCl, compound 5, CH2Cl2–H2O, 8 h, rt, 55–59%; (ix) SnCl2–2H2O, MeOH,
6 h, reflux, 70–72%; (x) HgCl2, CaCO3, MeCN–H2O, 12 h, rt, 56–60%.

A. Kamal et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3955–3958
3957
Scheme 3. (i) TEA, MsCl, CH2Cl2, 5 h, 0 ꢀC, 78%; (ii) SnCl2–2H2O, MeOH, 2 h, reflux, 68%; (iii) NaN3, DMF, 16 h, 60 ꢀC, 76%; (iv) TPP, THF–
H2O, 8 h, rt, 72%; (v) HOBt, EDCl, compound 5, CH2Cl2–H2O, 8 h, rt, 63–65%; (vi) SnCl2–2H2O, MeOH, 2 h, reflux, 68–73%; (vii) HgCl2, CaCO3,
CH3CN–H2O, 12 h, 51–55%.
Compounds 1a–c and 2a–b have been evaluated for the
primary anticancer activity in the standard three-cell
line panel comprising of the MCF7 (breast), NCI-H460
(lung) and SF-268 (CNS), and none of these compounds
showed significant cytotoxicity. The DNA binding abil-
ity for these novel A-C8/C-C2 alkoxyamido-linked PBD
dimers has been examined by thermal denaturation
studies using calf thymus (CT) DNA. Melting studies
show (Table 1) that these compounds stabilize the ther-
mal helix ! coil or melting stabilization (ÁTm) for the
CT–DNA duplex at pH 7.0, incubated at 37 ꢀC, where
PBD/DNA molar ratio is 1:5. The compounds 1a–c
exhibited a large value of ÁTm particularly after 18 h
incubation in comparison to DC-81 and for compounds
2a–c these values are insignificant.
DNA binding ability. On the contrary, the imine-amide
dimers (2a–c) have not exhibited any significant DNA
binding ability. Unlike for the previously reported PBD-
dimers a correlation between the DNA-binding affinity
and cytotoxictiy could not be derived in this class of
head to tail PBD dimers. Therefore, preparation of
structurally modified analogues of such dimers particu-
larly by incorporating exo- and endo-unsaturation at C2
and variation of linker length could probably address
the factors responsible for the insignificant cytotoxicity
of such efficient DNA-binding PBD dimers. The
detailed cross-linking ability, anticancer activity and
molecular modelling studies for these compounds is in
progress and will be published in due course.
It is observed from this preliminary data that imine-
imine PBD dimers (1a–c) have interesting profile of
Acknowledgements
We thank the National Cancer Institute, Maryland for
the primary anticancer assay in human cancer cell lines.
We are also grateful to CSIR, New Delhi for the award
of research fellowships to P.R., O.S. and G.R.
Table 1. Thermal denaturation data for A-C8/C-C2 alkoxyamido-
linked PBD dimers with CT–DNA
PBD
dimers
[PBD]/[DNA]
molar ratiob
ÁTm (ꢀC)a after
incubation at 37 ꢀC for
References and Notes
0 h
18 h
1a
1b
1c
DC-81
DSB-120
1:5
1:5
1:5
1:5
1:5
0.9
3.7
6.1
0.3
4.7
9.3
10.9
0.7
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.
1
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7.24–7.42 (m, 7H), 7.70 (d, 1H, J=4.6 Hz), 9.15 (s, 1H); MS
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.