A novel approach to the synthesis of cytotoxic C2-C3 unsaturated pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) with conjugated acrylyl C2-substituents

Article abstract of DOI:10.1016/j.bmcl.2003.12.094

A concise synthesis of three novel C2-C3 unsaturated pyrrolo[2,1-c][1,4] benzodiazepine analogues (18-20) containing conjugated acrylyl C2-substituents is reported that utilises Heck coupling to install the C2-acrylyl side chains. These analogues possess

Full text of DOI:10.1016/j.bmcl.2003.12.094

Bioorganic & Medicinal Chemistry Letters 14 (2004) 1547–1549
A novel approach to the synthesis of cytotoxic C2–C3 unsaturated
pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) with conjugated
acrylyl C2-substituents
Zhizhi Chen, Stephen J. Gregson, Philip W. Howard* and David E. Thurston*
Cancer Research UK Gene Targeted Drug Design Research Group, Department of Pharmaceutical and Biological Chemistry,
School of Pharmacy, 29–39 Brunswick Square, London, WC1N 1AX, UK
Received 25 November 2003; revised 22 December 2003; accepted 23 December 2003
Abstract—A concise synthesis of three novel C2–C3 unsaturated pyrrolo[2,1-c][1,4]benzodiazepine analogues (18–20) containing
conjugated acrylyl C2-substituents is reported that utilises Heck coupling to install the C2-acrylyl side chains. These analogues
possess significant cytotoxicity according to the NCI 60-cell line screen with 18 surpassing anthramycin (1) in potency.
# 2004 Elsevier Ltd. All rights reserved.
There is currently interest in the pyrrolobenzodiazepine
(PBD) group of antitumour antibiotics not only for
their inherent antitumour activity, but also because of their
ability to recognise GC base pairs and their potential as
components of larger molecules that can regulate gene
expression.1ꢀ4 SARstudies have revealed that the pre-
sence of a C2-substituent along with exo and/or endo
unsaturation in the C-ring are important for optimal
DNA binding and antitumour activity.5 Crucially, most
published synthetic pathways to C-ring endo/exo unsa-
turated PBDs rely on the presence of a 9-hydroxy
substituent in the A-ring (e.g. anthramycin, 1, Fig. 1).
This allows the formation of an O9–N10 benzal-pro-
tected dilactam intermediate of type 2 which can then be
reduced to the target PBD containing the DNA-inter-
active electrophilic N10–C11 imine moiety.6 However, it
is now accepted that the presence of a 9-hydroxy group
is therapeutically undesirable as it leads to acute cardio-
toxicity.7 Thus, a concise synthetic approach was required
to allow the introduction of unsaturated C2-substituents
to C2–C3 unsaturated des-9-hydroxy PBD targets.
More recently we demonstrated that C2-aryl-substituted
PBDs with C2–C3 endo-unsaturation could be prepared
from N10-SEM-protected dilactam intermediates of
type 4.9 However, in our hands, attempts to reduce C2-
N,N0-dimethylacrylamide dilactams of type 5 to endo/
exo-unsaturated PBDs with intact N10–C11 moieties have
been unsuccessful.10,11 Therefore, we have developed a
strategy whereby the desired unsaturated C2 substituents
are added by a Heck coupling reaction to an N10-pro-
tected PBD C11-carbinolamine, thus avoiding the need
to reduce a dilactam intermediate.12 This approach also
has the advantage of directly introducing the complete
C2-side chain thus avoiding the multiple functional
group interconversions employed by Fukuyama13 and
Langlois5 in their syntheses of porothramycin B (6),
and by Leimgruber6 in his synthesis of anthramycin (1).
The first task was to prepare a PBD C2–C3 enol triflate
capable of participating in a Heck coupling reaction.
The synthesis (Scheme 1) commenced with the coupling
of an o-nitrobenzoic acid (7) to a TBS-protected C-ring
fragment14 employing oxalyl chloride/DMF to give 8.
The nitro group was then reduced with Raney nickel/
hydrazine, and the resulting aniline (9) protected as a
Troc carbamate (10). The Troc group was chosen for its
compatibility with both palladium coupling chemistry
and PBD N10–C11 imine formation.15 The pro-C2-
hydroxy group in 10 was oxidized to the ketone 11
under Swern conditions in excellent yield (99%).
Removal of the TBS group in high yield (93%) provided
Mori and co-workers have reported the synthesis of C2-
exo-unsaturated PBD natural products such as tomay-
mycin via N10 -MOM-protected dilactams of type 3.8
Keywords: PBDs; DNA binding; Cytotoxic; SAR.
* Corresponding authors. Tel.: +44-(0)207-753-5932; e-mail: philip.
howard@ulsop.ac.uk; david.thurston@ulsop.ac.uk
0960-894X/$ - see front matter # 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.12.094

1548
Z. Chen et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1547–1549
Figure 1. Some PBD natural products (1,6) and their synthetic precursors (2–5).
Scheme 1. (a) (COCl)2, DMF, TEA, CH2Cl2, 55%; (b) Raney nickel, hydrazine, MeOH, 98%; (c) Troc–Cl, pyridine, DCM, 86%; (d) (COCl)2,
DMSO, TEA, DCM, ꢀ60 ꢁC, 99%; (e) AcOH, H2O, THF, 93%; (f) (COCl)2, DMSO, TEA, DCM, ꢀ60 ꢁC, 55%; (g) Tf2O, pyridine, DCM, 30%;
(h) CH2=CHCONMe2, DABCO, (CH3CN)2Pd(II)Cl2, MeOH, 45–55 ꢁC, 59%; (i) CH2=CHCOOMe, (PPh3)2Pd(II)Cl2, TEA, DMF, 75–78 ꢁC,
10%; (j) CH2=CHCONH2, DABCO, (CH3CN)2Pd(II)Cl2, MeOH, 45 ꢁC, 48%; (k) 10% Cd–Pb, THF, NH4OAc, 34% (18), 90% (19), 36% (20).
12 which could be cyclized to form the PBD B-ring (13)
through Swern oxidation. The cyclized ketone 13 was
treated with trifluoromethanesulphonic anhydride to
afford the C2–C3 enol triflate 14 in 30% yield. This dis-
appointing yield was due in part to a competing aldol
reaction which lead to the formation of a C–C linked
PBD dimer by-product. The triflate 14 was coupled to
dimethyl acrylamide, methyl acrylate and acrylamide
under Heck conditions to afford three N10-Troc-pro-
tected PBDs with unoptimised yields of 59% (15), 10%
(16) and 48% (17), respectively. Removal of the Troc
protecting group with Cd/Pb-couple afforded the target
PBDs (18–20) in modest to good yields (34–90%).
Compounds 18–20 were evaluated in the NCI 60-cell
line screen. Compared to the known cytotoxic agent
anthramycin (1) which has an average GI50 value of
29.2 nM across the 60 cell lines, these analogues pos-
sessed potent cytotoxicity with average GI50 values of
12.0 nM (18), 12.6 nM (19) and 52.5 nM (20). Table 1
provides a summary of GI50, TGI and LC50 data. In
particular, compound 18 which possesses a N,N-dimethyl
acrylamide C2-side chain is significantly more potent
than anthramycin across all of these endpoints. It is
remarkably cytotoxic towards certain cell lines (e.g.,
sub-nanomolar in CCRF-CEM and UACC-257), and a
selection of these results in comparison to anthramycin

Z. Chen et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1547–1549
1549
Table 1. Average GI50, TGI and LC50 values for compounds 18, 19
and 20 across the 60 cell lines of the NCI screen. Data for anthramycin
methyl ether (1) are provided for comparison
currently underway to improve the efficiency of this
critical step. Compounds 18–20 are presently under-
going in vivo evaluation and the results of these studies
will be reported elsewhere.
Compd
GI50 (mM)a
TGI (mM)b
LC50 (mM)c
Anthramycin
0.029
0.012
0.013
0.053
0.61
0.096
1.32
3.72
12.7
0.65
55.0
69.2
18
19
20
Acknowledgements
This work was supported by Cancer Research UK
(Grant number: C180/A1060 to D.E.T. and P.W.H.).
The NCI is also thanked for carrying out the cytotoxi-
city evaluations, and The School of Pharmacy (Uni-
versity of London) is acknowledged for providing an
ORS-supported bursary to Z.C.
a GI50: Dose that inhibits 50% of cell growth.
b TGI: Dose that inhibits 100% of cell growth.
c LC50: Dose that kills 50% of cells.
Table 2. Cytotoxicity data for 18 in selected sensitive cell lines from
the NCI 60-cell line screen. Data for anthramycin methyl ether (1) are
included for comparison
References and notes
Tumour type
Cell Lines
GI50 (nM)a
Anthramycin
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CCRF-CEM
HOP-92
HCT-116
U251
UACC-257
OVCAR-5
RXF 393
PC-3
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is shown in Table 2. It is noteworthy that the des-9-
hydroxy analogue of anthramycin (20) retains a sig-
nificant degree of the potency of the parent compound,
thus indicating that cytotoxicity is essentially indepen-
dent of the problematic 9-OH moiety.
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Matthews, C. S.; Howard, P. W.; Thurston, D. E.
Chemical Communications 2002, 1764.
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Tetrahedron Lett. 1984, 25, 2649.
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H. Tetrahedron Lett. 1993, 34, 2577.
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In summary, the successful synthesis of the three novel
PBDs 18–20 demonstrates the versatility of this new
convergent route which should be applicable to a range
of des-9-hydroxy C2-endo/exo-unsaturated PBDs other-
wise difficult to access. The route is short compared to
those previously reported for C-ring endo-exo-unsatu-
rated PBDs. For example, Leimgruber employed six
synthetic steps to elaborate the C2 side chain in his
pioneering synthesis of anthramycin.6 A further advan-
tage is that the Heck approach avoids stereoselectivity
issues potentially arising with alternative methods
employing olefination reactions. The low yield of the
triflation reaction was disappointing and efforts are
15. Guiotto, A.; Howard, P. W.; Baraldi, P. G.; Thurston,
D. E. Bioorg. Med. Chem. Lett. 1998, 8, 3017.