Synthesis and anticancer evaluation of spermatinamine analogues

Article abstract of DOI:10.1016/j.bmcl.2016.01.083

Spermatinamine was isolated from an Australian marine sponge, Pseudoceratina sp. as an inhibitor of isoprenylcysteine carboxyl methyltransferase (Icmt), an attractive and novel anticancer target. Herein, we report the synthesis of spermatinamine analogues

Full text of DOI:10.1016/j.bmcl.2016.01.083

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and anticancer evaluation of spermatinamine analogues
c
Basem A. Moosa a,b, , Sunil Sagar , Song Li a,b, Luke Esau c, Mandeep Kaur c,d, Niveen M. Khashab a,b
⇑
a Controlled Release and Delivery (CRD) Lab, Chemical Life Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955-6900, Saudi Arabia
b Center for Advanced Membranes and Porous Materials, King Abdullah University of Science and Technology, Thuwal 23955-6900, Saudi Arabia
c Biomolecular Lab, Computational Bioscience Research Center, King Abdullah University of Science and Technology, Thuwal 23955-6900, Saudi Arabia
d School of Molecular and Cell Biology, University of the Witwatersrand, Private Bag 3, Wits, 2050, Johannesburg, South Africa
a r t i c l e i n f o
a b s t r a c t
Article history:
Spermatinamine was isolated from an Australian marine sponge, Pseudoceratina sp. as an inhibitor of iso-
prenylcysteine carboxyl methyltransferase (Icmt), an attractive and novel anticancer target. Herein, we
report the synthesis of spermatinamine analogues and their cytotoxic evaluation against three human
cancer cell lines, that is, cervix adenocarcinoma (HeLa), breast adenocarcinoma (MCF-7), and prostate
carcinoma (DU145). Analogues 12, 14 and 15 were found to be the most potent against one or more cell
Received 14 June 2015
Revised 25 January 2016
Accepted 29 January 2016
Available online xxxx
lines with the IC50 values in the range of 5–10 lM. The obtained results suggested that longer polyamine
linker along with aromatic oxime substitution provided the most potent analogue compounds against
cancer cell lines.
Keywords:
Natural products
Bromotyrosine
Spermatinamine
Cancer
Ó 2016 Elsevier Ltd. All rights reserved.
Cytotoxicity
Bromotyrosine secondary metabolites are marine invertebrates
derived natural products and have been described for their variety
of biological activities including: anticancer, antimicrobial,
antifouling, antiviral, ATPase regulator, calcium channel modulator,
etc.1 More than 300 bromotyrosine-derived alkaloids are currently
known and divided into six categories according to their chemical
structures: simple bromotyrosine derivatives (suberedamines A),2
oximes (spermatinamine),3 bastadins ((E,E)-Bastadin 19),4 spirocy-
clohexadienylisoxazolines (11-hydroxyaerothionin),5 and other
more complex structural classes. The anticancer activity of bromo-
tyrosine-derived natural products has also been investigated and a
significant number of compounds have been found to elicit anti-
cancer activity, both in vitro and in vivo.6–8
Spermatinamine (1), a polyamine alkaloid, containing a bromo-
tyrosyl-spermine-bromotyrosyl sequence was isolated from an
Australian marine sponge, Pseudoceratina sp. as an inhibitor of iso-
prenylcysteine carboxyl methyltransferase (Icmt).3 Icmt is an
attractive and novel anticancer target and the various studies have
provided strong evidence that tumorigenesis can be markedly
impaired in cells by blocking Icmt activity.9 Polyamines have been
described earlier for their variety of cellular functions and cancer
associations.10–12 Polyamines analogues are being developed as
anticancer drugs to target polyamines metabolic enzymes.10,12–16
The ability of different types of polyamines to recognise different
receptor systems provided the rationale for developing polyamines
as drugs selective for different biological targets.17 Polyamines
could be considered as a skeleton key in the drug-receptor recog-
nition process because it can assume different conformations in
order to enable interaction between the drug and the receptor.18
Recently, Hillgren et al.19 reported the isolation and characterisa-
tion of four novel bromotyrosine polyamine alkaloids, pseudoce-
ramines A–D (2–5) from marine sponge, Pseudoceratina sp. as
inhibitors of the type III secretion (T3S) system of Gram-negative
bacterium Yersinia pseudotuberculosis (see Fig. 1).
As minor structural variations can have considerable effect on
the biological activity of these natural compounds, we synthesized
a number of spermatinamine analogues by modifying the polya-
mine linker between the two-bromotyrosine rings and the substi-
tution on oxime group. The compounds were then evaluated
against three human cancer cell lines, that is, cervix adenocarci-
noma (HeLa), breast adenocarcinoma (MCF-7), and prostate carci-
noma (DU145) by using MTT assay. To the best of our knowledge
no such study on the anticancer activities of spermatinamine ana-
logues has been performed to date, which is necessary for an even-
tual lead optimisation within this class of modified natural
products.
Spermatinamine was synthesized by following the recently
reported synthetic route19 with an overall yield of ꢀ55% (over 4
steps). In our synthetic strategy, 3,5-dibromo-5-methoxybenzalde-
hyde was used as starting material to prepare dibromo-O-methylty-
rosine which was then converted to azalactone (6) by reacting it
⇑
Corresponding author. Tel.: +966 128082677.
E-mail address: basem.moosa@kaust.edu.sa (B.A. Moosa).
http://dx.doi.org/10.1016/j.bmcl.2016.01.083
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Moosa, B. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.01.083

2
B. A. Moosa et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Br
OH
OR
Br
O
Me
N
N
H
N
Br
N
H
N
Me
N
N
O
O
MeO
HO
HO
Br
1
R = Me
2 R = H
Br
Me
OMe
Br
O
Me
N
N
H
N
Br
N
H
N
Me
MeO
Br
O
3
O
Br
Br
NHMe
N
NHMe
N
N
Me
H
H
N
N
MeO
HO
MeO
HO
Br
Br
5
4
Figure 1. Structures of spermatinamine (1) and pseudoceramines A–D (2–5).
yielded three types of substituted hydroxamic acid. The hydrox-
amic acid analogues were converted to methyl esters (7–9) by
reacting it with methyl iodide in presence of DBU as a catalyst as
shown in Scheme 1.21 It is noteworthy that most of reaction prod-
ucts were reasonably pure and column chromatography purifica-
tion was needed only in the final step. The oxime geometry was
determined as (E) based on NMR and as reported earlier.22,23
Methyl esters (7–9) were then coupled with various types of
polyamines20,24 as shown in Scheme 2. This efficient synthesis
afforded the products (10–21) with approx. 50–55% yield. The
polyamines chosen for the coupling were biogenic polyamines,
Scheme 1. Synthesis of substituted oxime methyl esters (2–4).
with N-acetylglycine.20 The synthesized azalactone (1) was then
hydrolysed by using barium hydroxide followed by addition of dif-
ferent types of hydroxylamine hydrochloride (R = H, Me, Bz) which
Scheme 2. Synthesis of spermatinamine analogues (10–21).
Please cite this article in press as: Moosa, B. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.01.083

B. A. Moosa et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
Table 1
Cytotoxicity (IC50 in
and DU145)
to make hybrid drug molecules in order to improve cellular import,
to increase affinity for chromatin, or to serve as carriers. The 1H
NMR analyses of the purified compounds in Scheme 2 revealed
the symmetric character of the final products (10–12 and 16–18).
While compounds (13–15) showed the unsymmetrical characteris-
tics (see Supporting information). The configuration of the oxime
was confirmed for the final products to be (E) based on NMR and
as reported in the literature for spermatinamine and other similar
natural product like psammapline A.23
The synthesized spermatinamine analogues were evaluated for
their cytotoxicity against three human cancer cell lines represent-
ing breast, cervical and prostate. MTT (3-(4,5-dimethylthiazol-z-
yl)-2,5-diphenyltetrazolium bromide) assay was used to assess
the IC50 (compound concentration required to inhibit 50% cell via-
bility) and growth inhibition of the cells after treatment with test
compounds. This assay targets the activity of succinate dehydroge-
nase enzyme in mitochondria, which reduces the tetrazolium salt
into formazan crystals. The intensity of the colour of formazan
dye correlates to the number of viable cells.28 The IC50 values were
calculated and are summarised in Table 1. Out of the 12 test com-
pounds, five compounds (11, 12, 14, 15 and 20) were found to be
active at lower concentrations while other compounds showed
lM) of test compounds against cancer cell lines (HeLa, MCF-7
Analogue No.
cLogPa
HeLa
DU145
MCF-7
24 h
48 h
24 h
48 h
24 h
48 h
10
11
12
13
14
15
16
17
18
19
20
21
6.29
8.50
11.25
6.73
9.01
11.74
6.64
9.01
11.70
5.84
8.00
10.80
7.45
>100
26.3
4.0
>100
8.9
>100
25.0
2.9
>100
8.4
>100
26.2
4.0
>100
8.9
>100
27.7
4.6
>100
8.5
>100
24.8
4.5
>100
11.1
5.4
>100
>100
>100
>100
27.1
>100
1.60
>100
19.1
3.4
>100
7.5
7.3
3.3
7.3
5.8
5.7
>100
>100
>100
>100
25.8
>100
—
>100
>100
>100
>100
10.8
>100
—
>100
>100
>100
>100
25.8
>100
1.60
>100
>100
>100
>100
27.9
>100
—
>100
>100
>100
>100
30.9
>100
—
Spermitinamine29
The bold values are to stress on the good biological activity.
a
cLogP was calculated using Chem draw software.
that is, naturally available such as spermine, spermidine and
cadaverine. Its well-known that biogenic polyamines such as
putrescine, spermidine, and spermine, are ubiquitous cellular
cations and exert multiple biological functions.25 Nayvelt et al.26
described that several polyamine analogues down-regulate polya-
mine levels, inhibit cellular uptake, and activate catabolism. Casero
and Woster27 also indicated that polyamine chains have been used
IC50 values greater than 100 lM. Comparison of cytotoxicities of
active analogues based on the type of linker suggested that sper-
mine, spermidine and bis(2-aminoehtyl)amine linkers were able
to induce significant cytotoxicity in all the cell lines, however ana-
logues with cadaverine linker were found to be inactive. It is also
Figure 2. Percentage growth inhibition of (a) DU145, (b) HeLa and (c) MCF-7 cancer cells after treatment with five most potent test compounds for 24 h. Data were
normalised to untreated controls, and error bars represent the standard deviation for triplicate experiments.
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4
B. A. Moosa et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
noteworthy that the only difference between spermatinamine and
spermine tethered compound (R = H) is the presence of methyl
group, but their cytotoxic effects differ widely. Activity profiles of
these five compounds were also suggestive of the fact that methyl
and aromatic substitution of oxime group is useful for their potent
activity.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.01.
083.
The effect of test compounds on cell growth/proliferation was
also analysed. Figure 2 represents the growth inhibition of cancer
cells after treatment with five (11, 12, 14, 15 and 20) active com-
pounds selected based on IC50 values (Table 1). Based on cell
growth inhibition, compounds 12, 14 and 15 were found to be
most active compounds (below 10 lM). Compound 12 was found
to be the most potent as it inhibited growth of approximately
70% of DU145 cells (Fig. 2a), >90% of HeLa cells (Fig. 2b) and
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approximately 60% of MCF-7 cells (Fig. 2c) at <5
None of the other test compound was found to be active below
M within 24 h of treatment. The anticancer activity of the com-
lM concentration.
5
l
pound 10 was found to be time dependent as the growth inhibition
was observed to be significantly increased from approximately 60%
at 24 h to 90% at 48 h in HeLa cells at 5 lM concentration (data not
shown). Both 12 and 15 were most active against HeLa cell line as
compared to DU145 and MCF-7 and this shows that these com-
pounds have selective activities against cervical cancer. Comparing
the structure activity relationship of spermatinamine analogues, it
has been understood that the longer chains linkers with the aro-
matic substitution of oxime group are essential for their cytotoxi-
city profiles.
In conclusion, present manuscript describes the synthesis of
spermatinamine analogues by using different biogenic polyamine
linkers and oxime substitutions and their cytotoxicity/growth inhi-
bition against three human cancer cell lines namely HeLa, DU145
and MCF-7. Syntheses of spermatinamine analogues have been
achieved in an overall yield of ꢀ34%. Based on IC50 and growth
inhibition results, 12, 14 and 15 were found to be the most potent
compounds. Structure activity relationship of these compounds
suggested that the longer chains linkers with the aromatic substi-
tution of oxime group are essential for their cytotoxicity profiles.
The study provided significantly important results for the lead
optimisation process and needs further studies in order to improve
the activity profiles of scaffolds.
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Acknowledgements
We gratefully acknowledge King Abdullah University of Science
and Technology (KAUST) – KSA and SEDCO grant for the financial
support. Also we would like to acknowledge Dr. Maan Hamad for
his help in getting the HRMS data.
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Please cite this article in press as: Moosa, B. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.01.083