The synthesis and in vitro cytotoxic studies of novel bus-naphthalimidopropyl polyamine derivatives

Article abstract of DOI:10.1016/S0960-894X(00)00293-6

Bis-naphthalimidopropyl putrescine (BNIPPut), spermidine (BNISpd), spermine (BNISpm) and oxa-putrescine (BNIPOPut) were synthesised and their growth-inhibitory properties characterised. All these compounds except for BNIPOPut, showed high in vitro cytotox

Full text of DOI:10.1016/S0960-894X(00)00293-6

Bioorganic & Medicinal Chemistry Letters 10 (2000) 1609±1612
The Synthesis and In Vitro Cytotoxic Studies of Novel
Bis-naphthalimidopropyl Polyamine Derivatives
Paul Kong Thoo Lina,* and Valentin A. Pavlovb
aThe Robert Gordon University, School of Applied Sciences, St. Andrew Street, Aberdeen AB25 1HG, Scotland, UK
bDepartment of Human and Animal Physiology, Faculty of Biology, University of So®a ``St. Kliment Ohridski'',
Dr. Tzankov Blvd. 8, So®a, 1421, Bulgaria
Received 31 March 2000; accepted 18 May 2000
AbstractÐBis-naphthalimidopropyl putrescine (BNIPPut), spermidine (BNIPSpd), spermine (BNIPSpm) and oxa-putrescine
(BNIPOPut) were synthesised and their growth-inhibitory properties characterised. All these compounds except for BNIPOPut,
showed high in vitro cytotoxic activity (with mean GI50 values between 0.5 and 8.45 mM) and selectivity against cancer cells derived
from nine di€erent human tumours. The increased content of nitrogen atoms in the linker chain of BNIPSpd and BNIPSpm sig-
ni®cantly improved their aqueous dissolution properties with a marginal decrease in their cytotoxic activity. # 2000 Elsevier
Science Ltd. All rights reserved.
In the 1970s, naphthalimides were synthesised as
potential antitumour agents,1 which have resulted in the
development of two compounds, namely mitona®de and
amona®de.2,3 In the early 1990s bis-naphthalimido com-
pounds which contain two naphthalimido rings, cova-
lently attached at the end of a linker chain containing two
nitrogen atoms were synthesised and their anticancer
properties evaluated.4 Among the numerous compounds
synthesised in this series, DMP 84085 and LU 7955396
(Fig. 1) have shown promise for cancer therapy.
chain. Our approach to increase the aqueous solubility
of bis-naphthalimido compounds was to introduce
more heteroatoms in the linker chain. The cytotoxic
properties of the newly synthesised bis-naphthalimido-
propyl putrescine (BNIPPut), spermidine BNIPSpd),
spermine (BNIPSpm) and oxa-putrescine (BNIPOPut)
against di€erent panels of human cancer cell lines are
also presented.
Bis-naphthalimido compounds with linker chain contain-
ing 2 nitrogens were previously synthesised by simply
reacting the corresponding alkyltetraamine with 1,8-
naphthalic anhydride.8 In order to introduce more het-
eroatoms in the linker chain, N-alkylation reaction was
chosen according to a modi®ed method which we recently
reported.9 The common intermediate for the synthesis of
BNIPPut, BNIPSpd, BNIPSpm and BNIPOPut is tolue-
nesulfonyloxypropylnaphthalimide 2. This was prepared
by ®rst reacting 1,8-naphthalic anhydride with aminopro-
panol to give N-(3-hydroxypropyl)naphthalimide which
upon reaction with tosyl chloride gave 2 in 60% yield
(Scheme 1). It is important to note that during the tosy-
lation reaction we found that the best condition to obtain
a maximum yield for 2 is to use four times excess of tosyl
chloride in a small volume of solvent.10 Under other
conditions (e.g., with equimolar or 2 molar quantities of
tosyl chloride), a mixture of 2 and 3, N-(3-chloropro-
pyl)naphthalimide is always formed thus reducing the
overall yield of the reaction and also, renders puri®ca-
tion very dicult.
With regard to their mode of action, it was suggested
that bis-naphthalimido compounds bind to DNA double
helix via the major groove.7 Although most of the bis-
naphthalimides have shown higher cytotoxic activity
than the monomeric compounds, they are however very
insoluble in aqueous solutions,8 making their testing and
potential development into chemotherapeutics dicult.
There have been a number of modi®cations undertaken
on the napthalimido rings of these compounds in an
attempt to improve their solubility and activity.8 Bis-
naphthalimido compounds which have been synthesised
and tested so far have only one or two nitrogen atoms in
the linker chain.8 We report in this paper the synthesis
of bis-naphthalimidopropyl polyamine derivatives,
which contain 2, 3 and 4 nitrogen atoms in the linker
*Corresponding author. Tel.: +44-1224-262818; fax: +44-1224-
262828; e-mail: p.kong@rgu.ac.uk
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00293-6

1610
P. K. T. Lin, V. A. Pavlov / Bioorg. Med. Chem. Lett. 10 (2000) 1609±1612
To obtain bis-naphthalimides BNIPPut, BNIPSpd,
BNIPSpm and BNIPOPut (Scheme 2), polyamines 4±7
were ®rst protected with 2,4,5-trimethylsulphonyl chloride
(Mts-Cl) in pyridine followed by their N-alkylation with
compound 2 to yield the fully protected bis-naphthali-
mido compounds which, on subsequent deprotection with
hydrobromic acid/glacial acetic acid in dichloromethane
a€orded BNIPPut, BNIPSpd, BNIPSpm and BNIPO-
Put as their hydrobromide salts.
The growth-inhibitory e€ects of BNIPPut, BNIPSpd,
BNIPSpm and BNIPOPut on cancer cells derived from
di€erent human tumours were carried out at the
National Cancer Institute (NCI) in the USA. Data from
the in vitro testing by the standard procedures of the
NCI Drug Discovery and Developmental Therapeutic
Programme11 are shown in Table 1 and Table 2. Each
compound was routinely tested at ®ve 10-fold dilutions
with a highest concentration of 100 mM. Cell growth and
Figure 1. Chemical structure of bis-naphthalimides DMP 8408 and LU 795553.
Scheme 1.
Scheme 2.

P. K. T. Lin, V. A. Pavlov / Bioorg. Med. Chem. Lett. 10 (2000) 1609±1612
1611
Table 1. Growth-inhibitory activity of bis-naphthalimidopropyl
polyamines against di€erent panels of human cancer cell lines
OVCAR-4, OVCAR-5, OVCAR-8, SK-OV-3); renal can-
cer (786-0, A498, ACHN, CAKI-1, RXF 393, TK-10,
UO-31); prostate cancer (PC-3, DU-145); breast cancer
(MCF-7, NCI/ADR-RES, MDA-MB-231/ATCC, HS
578T, MDA-MB-435, MDA-N, BT-549).
Panel of cell linesa
Compound/cytotoxicity
(GI50 (mM))b,c,d
BNIPPut BNIPSpd BNIPSpm BNIPOPute
In our preliminary experiments BNIPPut and BNIPO-
Put were found to be insoluble in either water or 10%
dimethylsulfoxide (soluble in dimethylsulfoxide on
warming). However, BNIPSpd and BNIPSpm showed
good solubility especially in 10% dimethylsulfoxide at
100-fold higher concentration than the highest test con-
centration in the cytotoxic study. Thus, it was concluded
that the increase of the solubility was assisted by an
increase of the number of nitrogens in the molecule. The
results from the cytotoxic studies (Table 1) show that
BNIPPut having 2 nitrogen atoms is the most active
compound according to the mean GI50 values against
di€erent panels of cancer cell lines. However, the newly
synthesised BNIPSpd and BNIPSpm with 3 and 4 nitro-
gen in the linker chain atoms respectively retain high
cytotoxic activity (Table 1). Thus, the better solubility of
BNIPSpd and especially of BNIPSpm in aqueous media
was achieved without any signi®cant drop in their
anticancer activity. Apart from the bis-naphthalimido
groups, the polyamine linker appears to be important for
the cytotoxic properties of these compounds since the
introduction of oxygen atoms in the linker of the most
active BNIPPut led to a dramatic increase in the GI50
values of the corresponding compound BNIPOPut (Table
1). BNIPPut, BNIPSpd and BNIPSpm were demon-
strated to possess a high selectivity in their cytotoxic
e€ects against di€erent panels of cell lines (Table 1). The
data in Table 2 show high selectivity in the cytotoxic
action of bis-naphthalimidopropyl polyamines against
di€erent cell lines in each panel (Table 2). For example
BNIPPut, BNIPSpd and BNIPSpm were found to be
202-, 443- and 24-fold respectively more active against
HS 578T than NCI/ADR-RES cell lines in the panel of
Breast Cancer. Nevertheless, with a few exemptions
BNIPPut was the most active compound against each
cell line, followed by BNIPSpd and BNIPSpm.
Leukaemia (6)
Non small cell
lung cancer (9)
Colon cancer (7)
CNS cancer (6)
Melanoma (8)
Ovarian cancer (6)
Renal cancer (7)
Prostate cancer (2)
Breast cancer (7)
Mean valuee
0.08
0.53
0.25
0.58
1.43
3.51
25.72
72.65
0.59
0.29
0.44
0.34
0.57
0.13
2.26
0.58
2.35
1.16
1.17
0.79
2.49
0.46
3.49
1.42
4.23
4.07
4.04
6.12
7.43
3.46
4.49
4.31
73.19
65.88
77.99
81.03
65.39
77.55
77.57
68.55
aThe number of cell lines routinely tested is shown in parentheses.
bData obtained from NCI's in vitro tumour cells screen.
cData are mean values of the corresponding panel.
dThe value of each cell line (not shown) is an average of at least two
testings.
eMean values over all cell lines tested.
Table 2. Cytotoxic activity of BNIPPut, BNIPSpd and BNIPSpm
against di€erent human cancer cell lines
Panel of cell lines
Cell line
Compound/cytotoxicity
(GI50 (mM))a,b
BNIPPut BNIPSpd BNIPSpm
Leukemia
HL-60 TB
K-562
EKVX
NCI-H522
HCT-15
SW-620
SF-268
0.222
0.012
1.370
0.013
2.660
0.037
0.029
0.488
0.082
0.722
0.024
1.910
0.156
1.170
1.050
0.030
10.900
0.449
0.111
4.780
0.267
1.310
0.185
10.600
19.500
0.044
0.033
3.600
4.500
0.825
12.800
1.160
0.722
5.350
0.856
10.800
0.555
10.800
15.100
0.630
Non small
cell lung Cancer
Colon cancer
CNS cancer
SF-295
Ovarian Cancer
Renal cancer
Breast cancer
IGROV1
SK-OV-3
786-0
UO-31
NCI/ADR-RES 13.10
HS 578T 0.065
aData obtained from NCI's in vitro tumour cells screen.
bData are an average of at least two testings.
After repeat of the primary screen, BNIPPut, BNIPSpd
and BNIPSpm were recently selected by the NCI Biologi-
cal Evaluation Committee for further in vivo testing. The
mechanism of action of the bis-naphthalimidopropyl
polyamines is currently under investigation in our
laboratory. These compounds possess rapid cytotoxic
activity and strongly bind to DNA which may contribute
to their cytotoxic action (unpublished data).
viability were estimated by the sulforhodamine B method
after 48 h drug exposure.11 The growth-inhibitory proper-
ties of the bis-naphthalimidopropyl polyamines were
expressed as a GI50 value de®ned as the drug concentration
that causes 50% growth inhibition.
The increased solubility and promising cytotoxic e€ects of
the newly synthesised BNIPSpd and BNIPSpm are good
basis for their further development as anticancer drugs.
The cell lines used in the in vitro screen were the following:
Leukaemia (CCRF-CEM, HL-60 (TB), K-562, MOLT-4,
RPMI-8226, SR); 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, HT-29, KM-
12, SW-620); CNS cancer (SF-268, SF-295, SF-539, SNB-
19, SNB-75, U251); melanoma (LOXIMVI, MALME-
3M, M14, SK-MEL-2, SK-MEL-28, SK-MEL-5, UACC-
257, UACC-62); ovarian cancer (IGROV1, OVCAR-3,
Acknowledgements
V.A.P. is a recipient of a postdoctoral NATO/Royal
Society fellowship. The authors wish to thank The
Robert Gordon University and The Royal Society of
Chemistry for the ®nancial support, The Centre of Mass

1612
P. K. T. Lin, V. A. Pavlov / Bioorg. Med. Chem. Lett. 10 (2000) 1609±1612
Spectrometry at the University of Wales, Swansea, for
mass spectroscopic analyses and the National Cancer
Institute (USA) for screening the compounds.
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