4
R. K. Amewu et al. / Bioorg. Med. Chem. xxx (2013) xxx–xxx
Table 3
Table 6
In vitro antimalarial and anticancer activity of tetraoxane dimers against chloroquine
In vitro anticancer activities against (HT29-AK) and HL-60 cells
sensitive strain 3D7a
a
a
Drug
IC50 mean ± SD (
l
M)
IC50 mean ± SD (
lM)
Compound
3D7 IC50 (nM)
Calculated logPb
HT29-AK
HL-60
6
7
8
9
16.9
57.2
58.4
4.0
3.5
4.7
11.2
38.6
47.1
21.3
28.9
937.7
76.5
908.2
305.2
40.9
415.5
47.5
101.9
1.6
6.5
8.1
8.1
7.9
6.9
6.9
7.3
6.7
5.6
5.7
6.0
4.1
4.4
4.8
5.7
5.6
5.4
4.5
3.0
2.7
2.5
4.6
7
8
9
92.5 ± 1.6
77.6 ± 5.5
90.2 ± 2.1
22.9 ± 4.4
37.2 ± 2.5
41.8 ± 3.3
13.6 ± 0.9
3.8 ± 0.8
32.8 ± 1.9
75.8 ± 0.8
38.8 ± 3.1
66.1 ± 2.9
>100.0
>100.0
18.8 ± 2.3
20.2 ± 2.4
7.0 ± 1.7
6.6 ± 1.8
7.1 ± 1.2
12.2 ± 2.0
34.3 ± 3.6
31.2 ± 1.8
2.0 ± 0.3
0.4 ± 0.1
11
20
21
23
24
25
27
ASN
DHA
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
0
1
2
3
4
5
6
0
1
2
3
4
5
6
7
ASN = artesunate, DHA = dihydroartemisinin.
IC50 values are averaged values determined from three independent
experiments.
a
equivalent to 10. Although the in vivo data is encouraging, none of
the dimers screened were as potent as artesunate.
Artesunate
Dihydroartemisinin
Chloroquine
3.5
10.2
In order to determine the anticancer potential of these dim-
mers, further experiments were conducted by screening a number
of the compounds against HT29-AK (colon) and HL-60 (leukemic)
cancer cell and the results are shown in Table 6 below. The cells
HT29-AK and HL-60 were treated with varying concentrations of
a
See Experimental section for the description of the assays. IC50 values are
averaged values determined from three independent experiments.
b
drugs (0–100 lM) for 24 h.
The results obtained showed that all the compounds tested pos-
sess activity against the colon cancer cell line HT29-AK. Com-
pounds 7, 8 and 9 displayed lower potency compared with DHA
while 11, 20, 21, 23, 24, and 25 displayed superior potency with
Table 4
In vitro antimalarial and anticancer activity of a selection of the tetraoxane dimers
against the chloroquine resistant strain K1 and multidrug resistant cancer cell line
a
KB
2
4 being the most potent. The dimers prepared from ethyl levuli-
Compound
IC50 (nM)
KB
nate generally displayed superior activity than the corresponding
adamantane derived analogues. For example the activities of the
K1
TI (KB/K1)
adamantanone fused dimers 7, 8,
90.2 M, respectively, while the activities of the ethyl levulinate
counterparts 23, 24, 25 are 13.6 M, 3.8 M, and 32.8 M, respec-
tively. These results show that while the adamantane fused dimers
displayed superior antimararial activity, the reverse is true against
the HT29-AK cancer cell line. On the contrary, the compounds were
not as potent as DHA against the leukemia cancer cell line HL-60.
Similar to the observation with the HT20-AK, the ethyl levulinate
analogues were, overall, more potent than the adamantane based
analogues.
9 are 92.5 lM, 77.6 lM,
6
7
9
1
1
2
2
2
2
2
8.9
101.5
2.6
24.6
2.7
308.5
266.6
101.5
136.4
611.8
250.0
ND
62696.9
133529.2
98577.8
62119.8
5041.0
39317.8
40709.0
7768.5
63129.9
24953.8
97725.5
11.7
7044.6
1315.6
377914.5
2525.2
1867.3
137.5
152.7
75.4
452.3
40.8
390.9
ND
l
l
l
l
1
2
0
2
3
5
6
Chloroquine
Podophylotoxin
To investigate whether the dimers have the potential to in-
crease apoptosis, a number of the compounds were tested for cas-
pase 3 activity against HL-60 cells. The results obtained showed
that all compounds increased apoptosis by increasing catalytically
active caspase-3 activity, with 9 and 11 displaying identical effects
on caspase-3 as DHA (Fig. 1).
ND = not determined.
See Experimental section for the description of the assays. IC50 values are
averaged values determined from three independent experiments.
a
Table 5
a
Oral efficacy of against P. berghei ANKA
%
Inhibition
10 mg/kg
4. Conclusions
1
mg/kg
3 mg/kg
30 mg/kg
ED50 (95% CI)
A selection of 1,2,4,5-tetraoxane dimers have been prepared
and their antimalarial activities evaluated. Most of the compounds
displayed activities in the low nanomolar range against chloro-
quine-sensitive and resistant strains, in vitro, versus P. falciparum
malaria parasite. Selected dimers had measureable oral activity
in the mouse model of malaria but were not as potent as artesu-
nate in this assay. The monospiro and dispiro tetraoxanes were
also shown to possess activity against HT29-AK (colon) and HL-
1
1
1
2
0
2
4
6
36.1
22.1
29.9
2.1
43.2
27.5
46.1
22.3
62.4
46.2
49.6
54.3
21.4
95.2
64.5
54.8
63.9
33.2
99.7
7.7
16.1
6.4
>30.0
1.6
ASN
38.5
ASN = artesunate.
a
See Experimental section for the description of the method used.
6
0 (leukaemic) cancer cells. The tetraoxane compounds with lower
>
30 mg/kg. In contrast to SAR performed on monomeric 1,2,4,5-
antimalarial activity were found to possess better anticancer activ-
ities and found to induce apoptosis by increasing caspase-3
activity.
tetraoxanes where the adamantane based systems performed best,
here the cyclohexyl based 1,2,4,5-tetraoxane dimer 14 proved to be