Comparison of the antiplasmodial and falcipain-2 inhibitory activity of β-amino alcohol thiolactone-chalcone and isatin-chalcone hybrids

Article abstract of DOI:10.1016/j.bmcl.2010.02.017

The synthesis and biological evaluation of two novel series of natural-product-like hybrids based on the chalcone, thiolactone and isatin scaffolds is herein described. Results for a 36-member β-amino alcohol triazole library showed that the thiolactone-chalcones, with IC₅₀s ranging from 0.68 to 6.08 μM, were more active against W2 strain Plasmodium falciparum than the isatin-chalcones with IC₅₀s of 14.9 μM or less. Also of interest is falcipain-2 inhibitory activity displayed by the latter, whereas the thiolactone-chalcones lacked enzyme inhibitory activity.

Full text of DOI:10.1016/j.bmcl.2010.02.017

Bioorganic & Medicinal Chemistry Letters 20 (2010) 2234–2237
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Comparison of the antiplasmodial and falcipain-2 inhibitory activity
of b-amino alcohol thiolactone-chalcone and isatin-chalcone hybrids
Renate H. Hans a, Jiri Gut b, Philip J. Rosenthal b, Kelly Chibale a,c,
*
a Department of Chemistry, University of Cape Town, Rondebosch 7701, South Africa
b Department of Medicine, San Francisco General Hospital, University of California at San Francisco, CA 94143, USA
c Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Rondebosch 7701, South Africa
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis and biological evaluation of two novel series of natural-product-like hybrids based on the
Received 18 December 2009
Revised 2 February 2010
Accepted 3 February 2010
Available online 10 February 2010
chalcone, thiolactone and isatin scaffolds is herein described. Results for a 36-member b-amino alcohol
triazole library showed that the thiolactone-chalcones, with IC50s ranging from 0.68 to 6.08
lM, were
more active against W2 strain Plasmodium falciparum than the isatin-chalcones with IC50s of 14.9
l
M
or less. Also of interest is falcipain-2 inhibitory activity displayed by the latter, whereas the thiolac-
tone-chalcones lacked enzyme inhibitory activity.
Keywords:
Thiolactone
Isatin
Ó 2010 Elsevier Ltd. All rights reserved.
Chalcones
Hybrids
1,2,3-Triazole
The reported decrease in the number of New Chemical Entities
(NCEs) introduced annually coupled with the increasing pace at
which resistance against established anti-infective drugs develops
presents a precarious situation especially for resource-poor coun-
tries with high endemicity for infectious diseases. Thus, there is
an increasing need for new, potent anti-infective agents.
One focus of our research programme is on the design, synthesis
and biological evaluation of hybrid compounds to identify novel
anti-infective agents. The rationale for this approach derives from
the expeditious SAR study offered through pharmacophore-rich
compound libraries.1 Further appeal stems from the reduced pro-
pensity for resistance development, the improved toxicity and
pharmacokinetic profile and enhanced and/or new properties
displayed by the hybrids so obtained.2 Another important,
although not often referred to, advantage offered is the potential
avoidance of unfavourable drug–drug interactions which may
occur in combination therapy or in the treatment of HIV, malaria
and TB co-infections.
tion reaction and functions as linker in the envisaged hybrids. To
our knowledge hybrids of this nature have not been reported pre-
viously. In an attempt to elucidate their mode of action the hybrids
were evaluated for inhibitory activity against the cysteine protease
falcipain-2, a validated target for antimalarial chemotherapy.
For guidance in hybrid design, we relied on previous SAR
studies and the thiolactomycin (1)-FabB model. One of the key
drug-receptor interactions revealed by this model is that the C-4
hydroxyl group of 1 (Fig. 1) extends into an incompletely filled
pantetheine pocket.4 Furthermore, a study revealed that modifica-
tion of the latter group, on C-5 derivatized analogues of 1, afforded
O
3
S
5
O
S
5
3
OH
-Thiolactomycin
O
OH
1 5
R
2 Thiolactone
This Letter reports on the synthesis and antiplasmodial activity
of two novel series of natural-product-like hybrids. Incorporated in
their design is the b-amino alcohol moiety, a known antimalarial
pharmacophore,3 and the solubility enhancing 1,2,3-triazole ring
system which is accessible via the Huisgen 1,3-dipolar cycloaddi-
O
5
3
O
R'
R
A
B
N
H
7
4 Chalcone
3 Isatin
* Corresponding author. Tel.: +27 21 650 2553; fax: +27 21 689 7499.
E-mail address: Kelly.Chibale@uct.ac.za (K. Chibale).
Figure 1. Natural product and natural product-like scaffolds used in hybrid
construction.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.02.017

R. H. Hans et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2234–2237
2235
S
ing the 5-chloroisatin derivative in target hybrids 7 and 8 stems
from reports on the modulating effect of the chlorine atom on the
chemical and biological properties of compounds.8 Its presence in
bioactive natural and synthetic drugs, for example vancomycin, gris-
eofulvin, clindamycin and especially chloroquine, is noteworthy.
As shown in Figure 2, the chalcone scaffold is a common mono-
mer in the envisaged hybrids 5–8. Chalcones 4 are of considerable
interest in drug discovery because of the diverse biological activi-
ties displayed by derivatives9 and the ease and simplicity of their
synthesis. As antimalarials, they may exert their effect via inhibi-
tion of cysteine proteases10 or new permeability pathways.11
Moreover, this scaffold allows for the systematic variation of sub-
stituents and/or substitution patterns on the aromatic rings A
and B for SAR investigation. Selection of the methoxy substituent
is on the basis of previous SAR which revealed that alkoxylated
chalcones are important for antimalarial activity.10,11
The envisaged 18-member thiolactone-chalcone hybrid library
is organized into two series 5 and 6, as depicted in Figure 2. The
aromatic ring A of 5 has the triazole moiety as a substituent at-
tached at either the ortho-, meta- or para-position, whereas ring
B is methoxylated to varying degrees. A similar arrangement holds
for 6 with the only difference being that the substituents on rings A
and B are interchanged. SAR studies set out to investigate the effect
on antimalarial activity of (i) the number of methoxy substituents
on ring B and ring A for 5 and 6, respectively, (ii) the position of
attachment of the triazolic moiety on ring A and ring B for 5 and
6, respectively, and (iii) the importance of substitution on ring A/
B for activity. The isatin-chalcone hybrids 7 and 8 were subjected
to a similar SAR study.
O
S
O
O
N
O
N
OH
OH
N
N
N
N
O
O
O
O
R
R
O
O
5
Cl
6
Cl
O
OH
O
N
N
OH
N
N
N
N
N
N
O
O
O
O
R
R
7
8
R = 4-methoxy; 2,4-dimethoxy; 2,3,4-trimethoxy
Figure 2. b-Amino alcohol hybrids 5–8.
S
S
O
O
S
O
iii
i
ii
5 + 6
O
O
75%
O-K+
83%
OH
9
O
10
N3
11
O
O
R
O
O
R
13
R = 4-methoxy; 2,4-dimethoxy; 2,3,4-trimethoxy
12
Synthesis of the thiolactone hybrids 5–6 is outlined in Scheme 1.
The intermediate 10 was obtained by reacting the potassium salt of
thiolactone 9 with a large excess of ( )-epichlorohydrin at 130 °C for
7 h under a nitrogen atmosphere.
Scheme 1. Reagents and conditions: (i) ( )-Epichlorohydrin (excess), 130 °C, 7 h;
(ii) NaN3 (5 equiv), NH4Cl (3 equiv), MeOH/H2O (8:1), 25 °C, 48 h; (iii) 12 (for 5,
1.1 equiv) or 13 (for 6, 1.1 equiv), CuSO4Á5H2O (5 mol %), sodium ascorbate
(10 mol %), CH2Cl2/H2O (1:1), 25 °C, 16 h.
For the azidolysis of 10 we employed a modification of the clas-
sical, non-chelating conditions.12 It involved the reaction of 1 equiv
of 10, 5 equiv of NaN3 and 3 equiv of NH4Cl—which acts as both
proton source and buffer—in aqueous alcohol at room temperature.
The latter method afforded 11 regioselectively in 83% yield.
Used in the final step of the synthesis of racemic 5 and 6, is a
procedure reported by Lee et al.13 Following this procedure 1 equiv
of the azide 11, 1.1 equiv of the acetylene 12 (13 for target mole-
cule 6), 5 mol % of CuSO4Á5H2O and 10 mol % of sodium ascorbate
was stirred in CH2Cl2/H2O (1:1) at 25 °C. Scheme 2 outlines the
synthesis of isatin-chalcone hybrids 7–8. The direct alkylation of
5-chloroisatin 14 with ( )-epichlorohydrin did not yield the
desired product. Synthesis of 7–8 therefore commenced with the
acetalization of 14. The protocol employed involved heating under
analogues which displayed a 143-fold increase in antimalarial
activity relative to the parent compound 1—a selective type II
FAS inhibitor.5 It was therefore anticipated that modification at
the C-4 hydroxyl group of
2 could provide analogues with
enhanced antimalarial activity. Derivatives of isatin 3 reportedly
display a wide spectrum of activity which includes the inhibition
of cysteine and serine proteases.6 Other attractive features of 3 in-
clude its non-peptidic nature and the ease with which it can be
synthetically manipulated, making it the ideal template onto
which bioactiphores can be appended.7 The rationale behind utilis-
O
MeO
MeO
OMe
O
OMe
O
Cl
Cl
Cl
ii
i
O
N
H
85%
N
N
H
O
14
15
16
iii
93%
MeO
O
N
OMe
Cl
Cl
v
iv
O
O
7 + 8
N
90%
OH
OH
N3
N3
17
18
Scheme 2. Reagents and conditions: (i) CH(OMe)3 (30 equiv), p-TsOHcat, MeOH, reflux, 48 h; (ii) ( )-epichlorohydrin (5 equiv), KF/Al2O3 (5 equiv), DCM, 25 °C, 20 h; (iii) NaN3
(5 equiv), NH4Cl (3 equiv), MeOH/H2O (8:1), 25 °C, 16 h; (iv) 10% HCl, acetone, 25 °C, 3 h; (v) 12 (for 7, 1.1 equiv) or 13 (for 8, 1.1 equiv), CuSO4Á5H2O (5 mol %), sodium
ascorbate (10 mol %), CH2Cl2/H2O (1:1), 25 °C, 3 h.

2236
R. H. Hans et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2234–2237
Table 1
of CuSO4ÁH2O and 10 mol % of sodium ascorbate in 4 mL DCM/
H2O (1:1) at room temperature. Synthesis of 8 proceeded in a sim-
ilar fashion with the acetylenic chalcone 13.
In vitro antimalarial activity against chloroquine-resistant (W2) strains of P.
falciparum and recombinant falcipain-2 inhibitory activity for 5–8
rec-FP-2,a IC50
M)
W2, IC50 (lM)
All the hybrids 5–8 gave 1H, 13C NMR, IR, HRMS and elemental
analysis data consistent with the proposed structures.
Compd
Position
R
(
l
1
2
3
5a
5b
5c
>100
>100
>100
>100
>100
>100
>20
>20
>20
2.31
2.08
1.14
Hybrids 5–8 were evaluated for growth inhibitory activity
against the W2 chloroquine-resistant strain of Plasmodium falcipa-
rum14 as well as for inhibitory activity against recombinant falci-
pain-2.14 The results obtained are summarised in Table 1.
The b-amino alcohol thiolactone-chalcone hybrids 5–6 showed
ortho
ortho
ortho
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
promising antiplasmodial activity with IC50s 6 6.08 lM against the
5d
5e
5f
meta
meta
meta
>100
>100
>100
2.43
1.64
2.00
W2 strain. However, relative to the control drug chloroquine, these
activities were still poor. The most interesting compound identi-
fied is the meta-substituted, trimethoxylated derivative 6f
5g
5h
5i
para
para
para
>100
>100
>100
3.12
1.54
5.04
[IC50 = 0.68 lM]. Molecules 5–6, with the exception of 6h, did not
show falcipain-2 inhibitory activity at the maximum concentration
tested. This result suggests that impairment of other parasite-path-
ways or targets is responsible for antimalarial activity. Results for
compounds 5–6 also indicate that antiplasmodial activity against
the W2 strain increases with an increase in the number of methoxy
substituents on the chalcone moiety, as exemplified by 5a–c and
6a–i and is consistent with earlier reports.10,11 The enhanced par-
asite inhibitory activity displayed by 6f and 6i indicates a prefer-
ence for methoxy substituents on ring A, whereas the activities
of compounds 5f and 6f suggest a preference for meta substitution
as far as the attachment of the triazole moiety to the chalcone scaf-
fold is concerned.
The isatin-chalcone hybrids 7–8 also displayed promising activ-
ity but were generally less active than the thiolactone-chalcone hy-
brids 5–6. Moreover, all compounds in the series 7–8 inhibited
falcipain-2 activity, although they were several 100-fold less active
than the cysteine protease inhibitor E64. However, E64 shows
6a
6b
6c
ortho
ortho
ortho
>100
>100
>100
2.64
2.14
1.72
6d
6e
6f
meta
meta
meta
>100
>100
>100
3.69
3.51
0.68
6g
6h
6i
para
para
para
>100
28.5
>100
6.08
3.10
0.81
7a
7b
7c
ortho
ortho
ortho
25.44
15.58
90.47
>20
>20
6.10
7d
7e
7f
meta
meta
meta
10.29
6.80
11.49
5.75
3.17
2.18
fairly weak antiplasmodial activity against W2 (IC50 ꢀ3
lM), pre-
sumably due to poor cell penetration. The results further show that
for falcipain-2 inhibitory activity meta substitution is preferred,
and that increased methoxylation does not generally enhance
activity (Table 1). Failure to correlate enzyme inhibitory activity
with growth inhibitory activity led us to conclude that cysteine
protease inhibition is not the primary mode by which these hy-
brids (7, 8) exert their antimalarial effect.
7g
7h
7i
para
para
para
10.84
ND
16.62
10.61
9.73
10.98
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
triOMe
4-OMe
2,4-diOMe
2,3,4-
8a
8b
8c
ortho
ortho
ortho
15.96
15.04
15.33
7.79
2.09
6.21
In conclusion a novel series of b-amino alcohol hybrids which
displayed encouraging antiplasmodial activity has been designed
and synthesized. The results demonstrate the potential for hybrid-
ization as an antimalarial drug discovery tool as evidenced by the
lack of activity of the advanced intermediates 11, 17, 12–1315 and
parent drugs (1 and 3) compared to the low micromolar activity of
the hybrids 5–8. This study also confirmed the interaction capabil-
ities of the isatin scaffold with the thiols of cysteine proteases.
8d
8e
8f
meta
meta
meta
9.91
10.61
16.73
3.57
2.95
2.57
8g
8h
8i
para
para
para
15.89
18.28
25.06
14.90
14.52
>20
triOMe
11
17
>100
>100
—
>20
>20
0.0694
ꢀ3
Chloroquine
E64
0.04733
Acknowledgements
a
Rec-FP-2 = recombinant falcipain-2.
ND = not determined.
b
Financial support by the African American Institute (RHH), the
Department of Science and Technology and the NRF through the
South African Research Chairs Initiative (KC), is gratefully
acknowledged.
reflux a mixture of 1 equiv of 14 and 30 equiv of anhydrous
trimethyl orthoformate in anhydrous MeOH in the presence of a
catalytic amount of para-toluene sulfonic acid for 48 h. With 15
in hand, the next step involved synthesis of the epoxide 16. This
was achieved by the reaction of 15 with 5 equiv of ( )-epichlorohy-
drin in the presence of 5 equiv of KF-Al2O3 in anhydrous CH2Cl2 at
25 °C.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.02.017.
Synthesis of the azide intermediate 17, proceeded as mentioned
earlier. Acid hydrolysis of 17 afforded 18 which participated in the
ligand-free Cu(I) catalyzed 1,3-cycloaddition protocol described
earlier. As shown in Scheme 2, hybrid 7 was obtained by reaction
of the azide 18 with the acetylene 12 in the presence of 5 mol %
References and notes
1. Biot, C.; Chibale, K. Infect. Disord. Drug Targets 2006, 6, 173.
2. (a) Barbachyn, M. R.. In Ann Rep Med Chem; Macor, J. E., Ed.; Elsevier Inc.: USA,
2008; Vol. 43, p 281; (b) Tietze, L. F.; Bell, H. P.; Chandrasekhar, S. Angew. Chem.,

R. H. Hans et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2234–2237
8. Naumann, K. J. Prakt. Chem. 1999, 341, 417.
2237
Int. Ed. 2003, 42, 3996; (c) Vennerstrom, J. L.; Arbe-Barnes, S.; Brun, R.;
Charman, S. A.; Chiu, F. C. K.; Chollet, J.; Dong, Y.; Dorn, A.; Hunziker, D.; Matile,
H.; McIntosh, K.; Padmanilayman, M.; Santo, T. J.; Scheurer, C.; Scorneaux, B.;
Tang, Y.; Urwyler, H.; Wittlin, S.; Charman, W. N. Nature 2004, 430, 900.
3. Pérez-Silanes, S.; Berrade, L.; García-Sánchez, R. N.; Mendoza, A.; Galiano, S.;
Pérez-Solórzano, B. M.; Nogal-Ruiz, J. J.; Martínez-Fernández, A. R.; Aldana, I.;
Monge, A. Molecules 2009, 14, 4120.
4. Price, A. C.; Choi, K. H.; Heath, R. J.; Li, Z.; White, S. W.; Rock, C. O. J. Biol. Chem.
2001, 276, 6551.
5. Jones, S. M.; Urch, J. E.; Brun, R.; Harwood, J. L.; Berry, C.; Gilbert, I. H. Bioorg.
Med. Chem. 2004, 12, 683.
9. (a) Nielsen, S. F.; Bosen, T.; Larsen, M.; Schonning, K.; Kromann, H. Bioorg. Med.
Chem. 2004, 12, 3047; (b) Go, M. L.; Wu, X.; Liu, X. L. Curr. Med. Chem. 2005, 12,
483; (c) Chen, M.; Theander, T. G.; Christensen, B. S.; Hviid, L.; Zhai, L.; Kharazmi,
A. Antimicrob. Agents Chemother. 1994, 38, 1470; (d) Lin, Y.-M.; Zhou, Y.; Flavin, M.
T.; Zhou, L.-M.; Nie, W.; Chen, F.-C. Bioorg. Med. Chem. 2002, 10, 2795.
10. Ring, C. S.; Sun, E.; McKerrow, J. H.; Lee, G. K.; Rosenthal, P. J.; Kuntz, I. D.;
Cohen, F. E. Proc. Natl. Acad. Sci. U.S.A. 1993, 90, 3583.
11. Go, M. L.; Liu, M.; Wilairat, P.; Rosenthal, P. J.; Saliba, K. J. Antimicrob. Agents
Chemother. 2004, 48, 3241.
12. Swift, G. A.; Swern, D. J. Org. Chem. 1967, 32, 511.
6. (a) Lyer, R. A.; Hanna, P. E. Bioorg. Med. Chem. Lett. 1995, 5, 89; (b) Chiyanzu, I.;
Hansell, E.; Gut, J.; Rosenthal, P. J.; McKerrow, J. H.; Chibale, K. Bioorg. Med.
Chem. Lett. 2003, 13, 3527; (c) Daisley, R. W.; Shah, V. K. J. Pharm. Sci. 1984, 73,
407; (d) Pandeya, S. N.; Sriram, D.; De Clerq, E.; Panne-Couque, C.; Witurouw,
M. Indian J. Pharm. Sci. 1998, 60, 207.
13. Lee, B.-Y.; Park, S. R.; Jeon, H. B.; Kim, K. S. Tetrahedron Lett. 2006, 47, 5105.
14. Greenbaum, D. C.; Mackey, Z.; Hansell, E.; Doyle, P.; Gut, J.; Caffrey, C. R.;
Lehman, J.; Rosenthal, P. J.; McKerrow, J. H.; Chibale, K. J. Med. Chem. 2004, 47,
3212.
15. Hans, R. H.; Guantai, E. M.; Lategan, C.; Smith, P. J.; Wan, B.; Franzblau, S. G.;
Gut, J.; Rosenthal, P. J.; Chibale, K. Bioorg. Med. Chem. Lett. 2010, 20, 942.
7. Chiyanzu, I.; Clarkson, C.; Smith, P. J.; Lehmann, J.; Gut, J.; Rosenthal, P. J.;
Chibale, K. Bioorg. Med. Chem. 2005, 13, 3249.