Antitrypanosomal and Antiplasmodial Activity of
Bis(2-aminoimidazoline) and Bisguanidine Derivatives
Christophe Dardonvillea (dardonville@iqm.csic.es), Lidia Nietoa, Fernando Rodríguezb, Isabel Rozasb,
Marcel Kaiserc, Reto Brunc, Binh Nguyend, W. David Wilsond, Rory Nelson Garcíae
Instituto de Química Médica, CSIC, Juan de la Cierva 3, E–28006 Madrid, Spain; b Centre for Synthesis and Chemical Biology, School of Chemistry, Trinity College
Dublin, Dublin 2, Ireland; c Swiss Tropical Institute, Socinstrasse, 57, CH-4002 Basel, Switzerland; d Department of Chemistry, Georgia State University, Atlanta, GA
30303-3083, USA; e Departamento de Parasitología, Facultad de Farmacia, Universidad Complutense de Madrid, Av. complutense s/n, E-28040 Madrid, Spain.
a
Antecedents: Recent findings by our group have shown that bisguanidine and especially bis(2-aminoimidazoline)diphenyl compounds displayed potent antitrypanosomal activity in
vitro1 and vivo2 against T. b. rhodesiense, the causative agent of acute human African trypanosomiasis (HAT). In addition, a correlation between antitrypanosomal activity and DNA
binding affinity was observed, suggesting a possible mechanism of action for these compounds.2 We also showed that this kind of molecules entered into trypanosomes via different
transporters in addition to P2, indicating that parasites that have lost the P2 transporter in selection of resistance to other drugs will not show cross-resistance to this class of molecules.
Others have described excellent antiplasmodial activity of related aromatic dicationic structures such as pentamidine or DB289.3 This prompted us to test our compounds against P.
falciparum as well.
1) In vitro antiprotozoal activity [Table 1]:
Aim of the study:
1) In vitro evaluation of new series of bis(2-aminoimidazolines) and their bisguanidine
counterparts4 against T. b. rhodesiense (STIB900) and P. falciparum (K1).
2) Evaluation of the selectivity index (SI = IC50 mammalian L6-cells/ IC50 parasite).
3) Study of the mechanism of action:
- DNA binding at AT-sites (Tm).
- Ferriprotoporphyrin IX binding inhibition test (FBIT).5






Antitrypanosomal activity: 9 new compounds with IC50 < 100 nM.
Best anti-T. brucei agent in vitro: bis(2-aminoimidazoline)fluorene 13b.
Antiplasmodial activity: 20 dicationic compounds with IC50 < 50 nM.
Best antimalarial compound in vitro: 1b and 13b.
Two cations are essential for good activity (data not shown)
Boc-protected derivatives (1d, 1e, 6e) show excellent in vitro antimalarial activity.
2) Selectivity index (safety profile) [Table 1]:
 Higher selectivity for the 2-aminoimidazolinium cation vs guanidinium.
X
X
6
1
3) Possible mechanism of action [Table 2]:
5
R
R
R
R
4'
1- 12
Y
2
 DNA binding: -Significant binding to A-T sites for several compounds: 1a, 1b, 5a,
12a, 12b, 9a, 9b, 13b.
- Tm (4,4’-Imi) > Tm (4,4’-Gua) in all cases.
 FBIT: - Some compounds show FPIX biomineralization inhibition similar to quinine
- There is no correlation between FPIX inhibition and antimalarial activity.
4
3
13 -14
Table 1. In vitro antitrypanosomal, antiplasmodial and DNA binding activity
of diphenyl dicationic compounds.
Table 2. Inhibition of FPIX biomineralisation
Compound
Cpd
1a
1b
1c
1d
1e
2a
2b
2c
3a
3b
4a
4b
5a
5b
5f
6a
6b
6e
7a
7b
Ra
Gua
Imi
(EtO)2CHCH2-Gua
(Boc)Gua
(Boc)Imi
Gua
Imi
(EtO)2CHCH2-Gua
Gua
Imi
Gua
Imi
Gua
Imi
CH2P+(n-pentyl)3
Gua
Imi
(Boc)Imi
Gua
Imi
X
Y
NH
CH2
O
S
O
O
O
S
O
N
H
N
H
-
IC50 (mM)
T.b.r.b
SI
30
0.022d
3072
0.069d
>767
0.228d
8
0.470d
204
0.048d
17
0.161d
71
0.897d
>553
0.316d
8
0.196
88
0.467
26
0.102
60
0.386
13
0.206d
>104
2.05d
29
0.414d
> 51
4.3d
32.4d
22
2.6d
> 1256
0.187
852
0.122
IC50 (mM)
P.f. c
SI
36
0.018
24091
0.0088
> 1548
0.113
48
0.077
166
0.059
88
0.032
4051
0.0157
> 4861
0.036
36
0.046
1079
0.038
76
0.035
932
0.025
40
0.068
> 1658
0.129
223
0.053
> 500
0.444
5.6
1025
0.055
> 2547
0.096
3714
0.028
Tm (ºC)
poly(dA•dT)2
not soluble
not soluble
2.0
4.3
2.2
3.3
a
The percentage of inhibition at 1 mg/mL (highest concentration tested)
is given when the IC50 could not be determined. b Taken from ref. 4.
1.5
4.1
S
8a
Gua
9a
9b
10a
10b
11a
11b
12a
12b
13a
13b
14a
14b
Gua
Imi
Gua
Imi
Gua
Imi
Gua
4,4’-Imi
4,4’-Gua
4,4’-Imi
5,4’-Gua
5,4’-Imi
N
H
N
H
O
N
H
N
N
N
nil
nil
CH2
CH2
-
0.538
29
0.607
26
1.4
CH2
CH2
CH2
CH2
0.036
0.025
0.045
0.054
0.217
0.038
0.270d
0.118d
0.024e
0.0049
0.050
0.060
319
0.055
0.028
0.019
0.016
0.041
0.011
0.0152
0.0123
0.0023e
0.0115
0.0088
0.0186
209
15.0
18.2
4.1
6.1
3.1
4.8
7720
24
646
111
3474
172
881
196
17020
15
620
6893
57
2181
585
12000
3053
8455
2043
7252
83
2000
18.1
4.1
5.2
Gua = guanidinium, Imi = 2-aminoimidazolinium, (Boc)Gua = N,N’-di(tert-butoxycarbonyl)guanidine, (Boc)Imi = N,N’-di(tertbutoxycarbonyl)imidazolidin-2-yl; b T. brucei rhodesiense STIB900 strain. Control: melarsoprol, IC50 = 0.0055 mM; c P. falciparum K1
strain. Control: chloroquine, IC50 = 0.278 mM; d Data taken from ref. 1; e Data taken from: Arafa et al. J. Med. Chem. 2005, 48, 5480.
a
Quinine
Chloroquine
1a
1b
1c
1d
1e
2a
2c
5b
5f
6a
6e
12b
Inhibition FPIX
IC50 (mM)a
324b
17.9
0%
429.2
161.8
45%
170.7
0%
15%
117.2
219.2
42%
27%
127.5
Conclusions:
 Bis(2-aminoimidazoline) compounds are promising anti-T. brucei and
antimalarial agents with better safety profile vs bisguanidine counterparts.
 Binding to DNA minor groove possibly plays a role in the antiprotozoal activity.
 Inhibition of heme biosynthesis may be responsible to some extent for the
observed antiplasmodial activity.
 In vivo assays are ongoing.
Acknowledgements: This work was supported by the UNDP/World Bank/WHO Special Program for
Research and Training in Tropical Diseases (RB), PIE grant 200680I121 from the CSIC (CD, LN), the
“Programa Nacional de Biomedicina” SAF2006-04698 grant from the Spanish “Ministerio de Educación
y Ciencia” (RNG), Cycle III HEA PRTLI grant by means of the CSCB in Ireland, and by a grant of the
Consejeria de Educacion Cultura y Deporte de la Comunidad Autonoma de La Rioja, Spain (F.R.).
References: [1] Dardonville, C.; Brun, R. J Med Chem 2004, 47, 2296. [2] Dardonville, C.; Barrett, M. P.; Brun, R.; Kaiser,
M.; Tanious, F.; Wilson, W. D. J Med Chem 2006, 49, 3748. [3] Soeiro, M. N. C.; De Souza, E. M.; Stephens, C. E.; Boykin,
D. W. Expert Opin Inv Drug 2005, 14, 957. [4] Rodriguez, F.; Rozas, I.; Ortega, J.E.; Meana, J.J.; Callado, L.F. J. Med.
Chem., 2007 (submitted). [5] Deharo, E.; Garcia, R. N.; Oporto, P.; Gimenez, A.; Sauvain, M.; Jullian, V.; Ginsburg, H. Exp
Parasitol 2002, 100, 252.