Cryptolepine analogues containing basic aminoalkyl side-chains at C-11: Synthesis, antiplasmodial activity, and cytotoxicity

Article abstract of DOI:10.1016/j.bmcl.2008.01.015

A series of cryptolepine derivatives has been synthesized through the incorporation of short basic side-chains in the C-11 position of the 10H-indolo[3,2-b]quinoline scaffold. Their antiplasmodial activity was evaluated in vitro against the chloroquine resistant Plasmodium falciparum W2 strain, showing IC₅₀ values between 22 and 184 nM, while their cytotoxicity was assessed using HUVEC cells, revealing three compounds with a selectivity ratio higher than 10. The most selective of these derivatives, 4d, with a selectivity ratio of 46, was also the least cytotoxic of the series.

Full text of DOI:10.1016/j.bmcl.2008.01.015

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 1378–1381
Cryptolepine analogues containing basic aminoalkyl side-chains
at C-11: Synthesis, antiplasmodial activity, and cytotoxicity
a
a
b
b
a,
*
Jo a˜ o Lavrado, Alexandra Paulo, Jiri Gut, Philip J. Rosenthal and Rui Moreira
a
iMed, CECF, Faculty of Pharmacy, University of Lisbon, Av Prof. Gama Pinto, 1649-003 Lisbon, Portugal
Department of Medicine, San Francisco General Hospital, University of California, San Francisco, Box 0811,
San Francisco, CA 94143, USA
b
Received 13 November 2007; revised 3 January 2008; accepted 3 January 2008
Available online 9 January 2008
Abstract—A series of cryptolepine derivatives has been synthesized through the incorporation of short basic side-chains in the C-11
position of the 10H-indolo[3,2-b]quinoline scaffold. Their antiplasmodial activity was evaluated in vitro against the chloroquine
resistant Plasmodium falciparum W2 strain, showing IC50 values between 22 and 184 nM, while their cytotoxicity was assessed using
HUVEC cells, revealing three compounds with a selectivity ratio higher than 10. The most selective of these derivatives, 4d, with a
selectivity ratio of 46, was also the least cytotoxic of the series.
Ó 2008 Elsevier Ltd. All rights reserved.
Malaria is a major infectious disease in tropical and sub-
tropical regions, caused by parasites of Plasmodium spe-
cies, the most lethal of which is P. falciparum.
Me
NH
CH3
1
Et N
2
6
7
4
1
N5
8
Chloroquine (CQ) 1 (Fig. 1) has been the mainstay of
malaria chemotherapy for 50 years, but its use is now
limited by the spread of CQ-resistant P. falciparum
3
2
9
N
Cl
Y
N
11
10
2
strains. Drug resistance results from the loss of activity
1
2
on erythrocyte-stage parasites, which are responsible for
the clinical symptoms of the disease. Thus, it is urgent to
find new blood schizontocides to combat the spread of
CH3
CH3
Cl
N
N
3
multi-drug resistant P. falciparum strains.
N
N
H
Cryptolepine (5-methyl-10H-indolo[3,2-b]quinoline) 2
(Fig. 1) is the major alkaloid from the roots of the west
African climbing shrub Cryptolepis sanguinolenta, an
H
X
3
4
1
2
X = NH-alkyl-NR R
herbal drug used in traditional medicine for the treat-
4
,5
Figure 1. Structures of chloroquine, 1, cryptolepine, 2, cryptolepine
hydrochloride (3, Y = Cl), cryptolepine triflate (3, Y = OTf), and
cryptolepine derivatives with basic side-chains at C-11, 4.
ment of malaria. Cryptolepine and its hydrochloride,
(Fig. 1), display potent in vitro antiplasmodial activ-
3
ity but also present cytotoxic properties that preclude
6
7
their clinical use. The cytotoxicity can be ascribed to
the ability of cryptolepine to intercalate into DNA and
However, studies on the possible mode of antimalarial
action suggest that cryptolepine is able to inhibit
hemozoin formation (i.e., the heme detoxification pro-
8
,9
inhibit topoisomerase II as well as DNA synthesis.
1
0,11
cess),
aminoquinolines (e.g., 1 and amodiaquine).
which is also the mechanism reported for 4-
1
2,13
A basic
amino side-chain is a major chemical feature required
for 4-aminoquinoline accumulation in the acid (pH
Keywords: Antiplasmodial; Cryptolepine; Quinolines; Malaria; 4-
Aminoquinolines.
5
hemoglobin digestion.
.5) food vacuole of the parasite, the site of the host
1
4
Based on this structural
requirement, we designed a novel group of cryptolepine
*
Corresponding author. Tel.: +35 1217946400; fax: +35
217946470; e-mail: rmoreira@ff.ul.pt
1
0
960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.01.015

J. Lavrado et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1378–1381
1379
analogues, 4 (Fig. 1), that incorporate an alkyldiamino
NH2
O
side-chain at C-11, aiming to increase accumulation in
the parasite food vacuole. We now report the synthesis
and evaluation of the antiplasmodial activity and cyto-
toxicity of compounds 4.
i
N
N
2
5
4%
1
3
14
O
O
ii
Cryptolepine hydrochloride, 3 (Y = Cl), and its C-11
alkyldiamine derivatives, 4, were synthesized via 11-
N
Br
N
N
1%
1
5
O
O
16
chloroquindoline intermediate, 9, according to the
route depicted in Scheme 1. Anthranilic acid, 5, was
treated with bromoacetyl bromide to afford the bromo-
acetyl derivative 6, which was treated with aniline to
give compound 7. Acid-catalyzed cyclization of 7 with
polyphosphoric acid (PPA) gave the quindolone 8,
15
iii 71%
H N
N
2
which, by reaction with POCl , gave the corresponding
3
1
7
1
5
1
1-chloroquindoline 9. Hydrogenation of 9 with 10%
1
6
Pd-C at 60 psi provided the quindoline 10. Reaction
of 9 and 10 with methyl triflate formed the correspond-
ing triflate 11 and cryptolepine triflate (3, Y = OTf).
Scheme 2. Synthesis of alkyldiamines required for compounds 4c and
4i. Reagents and conditions: (i) NH OAc, NaBH CN, anhydrous
MgSO , dry methanol, reflux; (ii) piperidine, TEA, CH
hydrazine, EtOH, reflux.
4
3
1
7
4
2
2
Cl , reflux; (iii)
Initial attempts to prepare compounds 4 involved the
reaction of triflate 11 with alkyldiamines, but the final
product contained always triflate and chloride anions.
Alternatively, compound 11 was first treated with so-
dium carbonate and then with HCl to give the key inter-
1
1
to the route depicted in Scheme 2. Thus, N ,N -dimeth-
ylpropane-1,2-diamine, 14, used for synthesis of 4c, was
prepared by reductive amination of N,N-dimethylami-
nopropanone, 13, with ammonium acetate and
1
7
mediate 11-chlorocryptolepine hydrochloride, 12.
Finally, cryptolepine derivatives containing an alkyldi-
NaBH CN. 3-Piperidin-1-ylpropan-1-amine, 17, used
3
amine side-chain at C-11, 4, were obtained by reacting
1
for synthesis of 4i, was prepared using a Gabriel synthe-
sis starting with the phthalimide 15 (Scheme 2).
8
1
2 with the appropriate amine, in 29–82% yield. Most
of the alkyldiamines used in this final step were pur-
chased and used without further purification. However,
for 4c and 4i, the corresponding amines required for the
substitution reaction with 12 were synthesized according
Compounds 4a–j were tested in vitro against CQ-resis-
1
9
tant W2 P. falciparum strain. The corresponding
IC50 values, together with their cytotoxicity determined
using human umbilical vein endothelial cells (HUVEC),
are presented in Table 1. As it appears from the data,
IC50 values for 4a–j range from 22 to 184 nM, which
represent a significant improvement in antiplasmodial
activity over the cryptolepine, 3, (755 nM) and chloro-
H
N
NH2
i
Br
O
9
6%
CO H
2
CO H
2
2
0
quine (249 nM). Inspection of the data in Table 1 al-
lows the following conclusions to be drawn.
5
6
ii 70%
H
N
H
N
First, compounds containing side-chains with three car-
bon atoms (e.g., 4e, f, h, i) generally present better anti-
plasmodial activity than those with two carbon atoms
N
H
iii
6%
O
6
N
CO2H
(e.g., 4a–c) or four carbon atoms (4j).
H
O
7
8
Second, branched side-chains reduce significantly the
activity independently of side-chain length (two carbon
atoms: 4c vs 4a, b; three carbons atoms: 4g vs 4e, f, h, i);
the exception to this trend is observed for the analogue
with the less flexible piperidine side-chain, that is, 4d.
iv 60%
Me
N
Y
N
vi
N
H
N
H
X
X
Third, terminal secondary and tertiary alkylamine
groups do not affect antiplasmodial activity as shown
by the IC50 values for 4h when compared with those
of its N,N-dimethyl and N,N-diethyl counterparts, 4e
and 4f, respectively.
9
X = Cl
11 Y = OTf; X = Cl
5
1%
v
10 X = H 85%
3 Y = OTf; X = H 96%
vii
1
2 Y = Cl; X = Cl
90%
viii
4
29-82%
Fourth, the cytotoxicity/antiplasmodial ratio for most
of the compounds 4 is >1, indicating some selectivity
against the parasite. Compound 4d emerged as the most
selective (selectivity ratio of ca. 46) as well as the less
cytotoxic (ca. 2 times less cytotoxic than cryptolepine)
of the present series. In contrast, compounds with
Scheme 1. Synthesis of cryptolepine derivatives 4a–j. Reagents and
conditions: (i) Bromoacetyl bromide, DMF/Dioxane, rt; (ii) aniline,
DMF, 120 °C; (iii) PPA, 130 °C; (iv) POCl
NaOAc, AcOH, 60 psi; (vi) 9 or 10, methyl triflate, anhydrous toluene,
rt; (vii) (a) 5% Na CO ; (b) HCl–Et O; (viii) RNH , AcOEt, reflux.
3
2
, 120 °C; (v) H Pd-C 10%,
2
3
2
2

1380
J. Lavrado et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1378–1381
Table 1. Antiplasmodial activity against P. falciparum W2 strain and cytotoxicity in HUVEC cells of cryptolepine analogues 4
a,b
a,c
Compound
X
H
Antiplasmodial IC50
b
W2
Cytotoxicity IC50
HUVEC
Cyto./antipl. ratio
20
CQ,1
249
—
3
755 (±1)
50 (±8)
82 (±23)
132 (±7)
44 (±1)
30 (±6)
32 (±5)
184 (±21)
22 (±2)
36 (±2)
127 (±2)
1180 (±1)
1.6
3.2
4
4
4
4
4
4
4
4
4
4
a
b
c
d
e
f
N
161 (±1)
674 (±1)
617 (±1)
2042 (±1)
155 (±2)
114 (±1)
82 (±1)
HN
HN
8.2
N
N
4.7
HN
HN
NH
46.4
5.2
HN
HN
N
N
3.6
g
h
i
HN
HN
N
0.5
794 (±1)
1161 (±1)
35 (±1)
36.1
32.3
0.3
N
H
HN
HN
N
j
N
IC50 values are in nM.
a
IC50 values are means of at least three determinations.
Ref. 20.
Ref. 23.
b
c
branched side-chains seem to be particularly cytotoxic in
the HUVEC cell line (i.e., 4g and 4j are 14 and 34 times
more cytotoxic than cryptolepine), with selectivity ratios
of ca. 0.4. Worthy of note, the most cytotoxic com-
pound, 4j, contains the chloroquine alkylamino substi-
tuent. Cytoxicity displayed by compounds 4 may
reflect electrostatic or hydrogen-bonding interactions
between the amine side-chain and the phosphate back-
References and notes
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2
3
. Hyde, J. E. Trends Parasitol. 2005, 21, 494.
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1,22
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In conclusion, a number of cryptolepine analogues, 4,
containing an alkyldiamine side-chain at C-11 have been
synthesized using a single key intermediate, 11-chloro-
cryptolepine, 12. Six of these cryptolepine analogues dis-
play potent antiplasmodial activity against the P.
falciparum W2 CQ-resistant strain, with IC50 values
ranging from 22 to 82 nM. The highest cytotoxicity/anti-
plasmodial ratio was observed for the piperidine deriva-
tive 4d. These results indicate that introducing an
alkyldiamino side-chain at position C-11 of cryptolepine
is a promising approach for further optimization against
drug-resistant malaria parasites.
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Acknowledgments
This work was supported by Funda c¸ a˜ o para a Ci eˆ ncia e
Tecnologia (FCT, Portugal); J.L. acknowledges FCT
for the PhD Grant SFRH/BD/29202/2006.
1

J. Lavrado et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1378–1381
1381
1
1
1
3. Kumar, S.; Guha, M.; Choubey, V.; Maity, P.; Bandyo-
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130.89; 130.32; 129.94; 126.08; 124.86; 123.15; 119.41;
119.00; 55.70; 52.11; 44.08; 35.22. Anal. (C, H, N): Calcd
C
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4 2
for C21H23ClN Æ2HClÆ0.4H O: C, 56.42; H, 5.82; N, 12.53;
Found: C, 56.52 H, 6.01 N, 13.80. ESI-TOF/MS (m/z):
Calcd for C21H23N : 331.1923; Found: 331.1917 Yields
þ
4
for remaining compounds: 4a 62%; 4b 35%; 4c 29%; 4e
34%; 4f 82%; 4g 78%; 4h 56%; 4i 38% and 4j 30%.
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23. The cytotoxicity evaluation of the series of compounds
studied was made using human umbilical vein endothelial
cells (HUVEC). The degree of cell injury was evaluated by
the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide (MTT) assay, in which cell viability is assessed
by mitochondrial-dependent reduction of MTT to forma-
zan. IC50 values were determined from graphics of degree
of cell injury over inhibitor concentrations with GraphPad
Prism software. Experimental details are provided as
Supporting Information.
1
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Hufford, C. D. Bioorg. Med. Chem. 2002, 10, 1337.
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E.; Litvak, J.; Lu, Q.; Zhang, P. S.; Reed, M. J.; Waldeck,
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8. 5-Methyl-11-(piperidin-4-ylamino)quindolinium hydro-
chloride (4d). To a suspension of 12 (50.0 mg, 0.165 mmol,
1
1
1
equiv) in AcOEt (5 mL) was added commercial piperi-
din-4-amine (29.7 mg, 0.296 mmol, 31.1 lL, 1.8 equiv).
Reaction mixture was refluxed for 24 h and after this
period, the formed precipitate was collected, washed with
diethyl ether, and dried, to give 4d, 66% (40.3 mg) yield, as
1
an orange solid, mp 325–327 °C. H NMR (400 MHz,
H
DMSO) d (ppm) 8.66 (d, J = 8.3 Hz, 1H), 8.56 (d,
J = 8.6 Hz, 1H), 8.29 (d, J = 6.2 Hz, 1H), 8.09 (dd, J = 8.6,
5
5
.6 Hz, 1H), 7.97 (d, J = 6.3 Hz, 1H), 7.84 (dd, J = 6.2,
.6 Hz, 1H), 7.81 (dd, J = 6.3, 5.6 Hz, 1H) 7.44 (dd,
J = 8.3, 5.6 Hz, 1H), 4.80 (s, 3H), 4.14 (d, J = 12.8 Hz,
H), 3.85 (t, J = 11.6 Hz, 2H), 3.55–3.45 (m, 1H), 2.28 (t,
J = 11.6 Hz, 2H), 2.10–1.97 (m, 2H). C NMR (100 MHz,
2
1
3