Structure-activity relationships amongst 4-position quinoline methanol antimalarials that inhibit the growth of drug sensitive and resistant strains of Plasmodium falciparum

Article abstract of DOI:10.1016/j.bmcl.2010.01.001

Utilizing mefloquine as a scaffold, a next generation quinoline methanol (NGQM) library was constructed to identify early lead compounds that possess biological properties consistent with the target product profile for malaria chemoprophylaxis while reducing permeability across the blood-brain barrier. The library of 200 analogs resulted in compounds that inhibit the growth of drug sensitive and resistant strains of Plasmodium falciparum. Herein we report selected chemotypes and the emerging structure-activity relationship for this library of quinoline methanols.

Full text of DOI:10.1016/j.bmcl.2010.01.001

Bioorganic & Medicinal Chemistry Letters 20 (2010) 1347–1351
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Structure–activity relationships amongst 4-position quinoline methanol
antimalarials that inhibit the growth of drug sensitive and resistant strains
of Plasmodium falciparum
a
a
a
a
a
Erin Milner a, , William McCalmont , Jayendra Bhonsle , Diana Caridha , Dustin Carroll , Sean Gardner ,
Lucia Gerena a, Montip Gettayacamin b, Charlotte Lanteri a, ThuLan Luong a, Victor Melendez a, Jay Moon a,
Norma Roncal a, Jason Sousa a, Anchalee Tungtaeng b, Peter Wipf c, Geoffrey Dow a
*
a Walter Reed Army Institute of Research, Silver Spring, MD, United States
b United States Army Medical Component, Armed Forces Research Institute of Medical Sciences, Bangkok, Thailand
c Department of Chemistry, University of Pittsburgh, Pittsburgh, PA, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
Utilizing mefloquine as a scaffold, a next generation quinoline methanol (NGQM) library was constructed
to identify early lead compounds that possess biological properties consistent with the target product
profile for malaria chemoprophylaxis while reducing permeability across the blood–brain barrier. The
library of 200 analogs resulted in compounds that inhibit the growth of drug sensitive and resistant
strains of Plasmodium falciparum. Herein we report selected chemotypes and the emerging structure–
activity relationship for this library of quinoline methanols.
Received 17 December 2009
Revised 29 December 2009
Accepted 4 January 2010
Available online 7 January 2010
Keywords:
Antimalarials
Published by Elsevier Ltd.
Plasmodium falciparum
Plasmodium berghei
Mefloquine
Quinoline methanol
Structure–activity relationship
Malaria continues to be a major health challenge in most trop-
ical and many subtropical regions.1 Of the 1–3 million annual fatal-
ities due to malaria, the majority are pregnant women and children
below age five. Our broad objective is to satisfy a projected future
unmet medical need for cost-effective, intermittent preventative
treatment (IPT) of malaria and prophylaxis in which a blood
schizonticidal drug is given periodically as a single therapeutic
dose to otherwise healthy, non-immune individuals.2 We opted
for the quinoline methanol scaffold due to the relative safety of
mefloquine (MQ, WR142490) in pregnancy and its well character-
ized clinical profile.3
Mefloquine would be the US Army’s drug of choice for prophy-
laxis but for its association with adverse central nervous system
(CNS) effects.4 Current justification for use is the long half-life
(weekly dosing) and activity against chloroquine-resistant malar-
ia.5 The ability of mefloquine to accumulate in the CNS at micro-
molar concentrations and target multiple receptors has been well
documented, but the relevant targets have not been identified.6
We seek to prevent CNS penetrability from the outset by reengi-
neering the quinoline methanol scaffold 5 (Fig. 1) to yield deriva-
tives that exhibit fewer adverse CNS effects while retaining
antimalarial efficacy. Research within the Walter Reed Army Insti-
tute of Research has shown that acyclic N-alkylaminoquinoline
methanols display a selectivity index superior to mefloquine7
and further evidence suggests the neurotoxicity of mefloquine ap-
pears to be associated with the piperidine ring.
Generally, the structure–activity relationships (SAR) were not
adequately studied during the 1960s development of mefloquine,
which relied upon in vivo models.8 We chose to increase potency
and reduce absorption through the blood–brain barrier by optimiz-
ing the 4-position aminoalcohol moiety and probing the quinoline
scaffold by adding novel pentafluorosulfanyl (–SF5) substituents.9
In order to optimize the 4-position moiety, we studied the effect
of alterations of physiochemical properties on activity while con-
structing over two hundred analogs. Due to the breadth and diver-
sity of this endeavor, it was decided to report these data in two
parts: (1) the structure–activity relationship that emerged and
(2) the analysis of the relationship between physiochemical prop-
erties and biological data.10 Herein we examine the 4-position SAR
and emerging chemotypes of interest, which we are currently opti-
mizing as part of a structure-based rational drug design program.
In order to efficiently prepare analogs with the 2,8-trifluoro-
methyl quinoline core, the vast majority of derivatives were
* Corresponding author. Tel.: +1 301 319 9461; fax: +1 301 319 9449.
E-mail address: erin.milner@us.army.mil (E. Milner).
0960-894X/$ - see front matter Published by Elsevier Ltd.
doi:10.1016/j.bmcl.2010.01.001

1348
E. Milner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1347–1351
Figure 1. Synthesis of the next generation quinoline methanol scaffold.
constructed according to the procedures outlined in Figure 1.11
Bis(trifluoromethyl)quinolin-4-ol 1 was melted along with phos-
phorous oxybromide to provide 4-bromo-2,8-bis(trifluoromethyl)
quinoline 2,12 which was subjected to a Stille13 reaction in order
to yield vinyl intermediate 3. Simple oxidation14 led to epoxide
4, which was diversified at the 4-position through a regioselective
SN2 nucleophilic ring opening mechanism. Since this was an
exploratory library meant to provide early-lead compounds, we
chose to construct and screen racemic compounds 5.
Table 2
P. falciparum IC90 (ng/mL) values and selectivity of phenyl, benzyl, and phenethyla-
mino QMs
Initially, the significance of the homobenzylic amine moiety
was explored. As shown in Table 1, while n-butyl amine
(WR177000) was quite efficacious, chains with heteroatoms such
as oxygen (WR308633), sulfur (WR308632), and carbon
(WR308653) were devoid of activity. Based upon these results,
only nitrogen-based moieties were further explored at the 4-posi-
tion in the remainder of the exploratory library.
a
b
Compound
n
LC50 (ng/mL)
IC90 (ng/mL)
WR308251
WR308252
WR308253
0
1
2
22,298
20,427
9638
484, 377, 437, 500
19, 67, 86, 151
19, 23, 43, 23
a
Cytotoxicity: macrophage IC50 (ng/mL).
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235,
b
and C2A.
In regards to nitrogen functionalization, the data suggests that
the nature of the 4-position amine and the degree of electron den-
sity around the nitrogen are most strongly associated with activity.
The remainder of the discussion explores the relationship between
4-position amine substituents and antimalarial activity.
Antimalarial and selectivity data for a subset of compounds
with aromatic side chains are presented in Table 2. The data sug-
gests a reduction of electron density with respect to the amine,
due to the adjacent benzene ring, diminishes potency. For example,
phenylamino QM (WR308251) has a much lower activity then ben-
zylamino QM (WR308252) and phenethylamino QM (WR308253).
All phenyl amines tested were nearly devoid of activity, and the
addition of electron-withdrawing groups further decreased
potency.
The imidazole class of compounds is interesting because the
IC90 values for many of them are similar across the four drug-resis-
tant strains with WR308437 and WR308623 in particular illustrat-
ing this reduction in cross-susceptibility to related drugs (Table 3).
This is encouraging, since in vitro susceptibility of clinical isolates
of Plasmodium falciparum to related aminoalcohol antimalarials
such as halofantrine correlates strongly to that of mefloquine.15
On the other hand, metabolic stability was poor, suggesting the
compounds may have short half-lives in vivo. Consequently, efforts
are underway to synthesize more metabolically stable compounds,
since their cross-susceptibility profiles are of great interest.
In regards to alkyl amino quinoline methanols, an intricate rela-
tionship between steric bulk and lipophilicity of the alkyl groups
has emerged and merits further study in order to determine the
discrete contributions to efficacy. Although the mechanism of ac-
tion for mefloquine is not fully understood, the lipophilic nature
of MQ and quinine is known to correlate with the delivery of drug
to the parasite.16 MQ is also known for high-affinity binding to
erythrocytes and other cells, which may provide a reservoir of drug
and contribute to the long half-life.17
Table 1
P. falciparum IC90 (ng/mL) values resulting from probing the homobenzylic site of QMs
Compound
R
IC90 (ng/mL)a
WR177000
WR308633
WR308632
WR308653
NH
0
S
2, 1, 9, 12
>500b
>500b
C
>500b
a
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235,
and C2A.
b
>500 ng/mL for each of four strains.

E. Milner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1347–1351
1349
Table 3
P. falciparum IC90 (ng/mL) values, selectivity, and metabolic stability of heterocyclic amino quinoline methanols (HAQMs)
R1
R2
R3
LC50 (ng/mL)
IC90 (ng/mL)
Moused (mm)
a
b
Compound
WR308437
WR308623
WR308626
WR308763
WR308764
Et
Me
H
H
H
H
H
25,707
50,082
50,082
NDe
69, 43, 56, 57
22
8
10
9
n-Pr
i-Pr
Me
H
360, 320, 250, 240
>500c
–Benzene–
–Benzene–
110, 220, 310, 320
120, 410, 480, 450
NDe
4
a
b
c
Cytotoxicity: macrophage IC50 (ng/mL).
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235, and C2A.
Selectivity index = LC50 against macrophages/IC90 against PF W2.
Half life in human liver micosomes. The half-life of mefloquine in this test system is >60 min.
ND = no data.
d
e
In general, these compounds were more potent than MQ and
der of magnitude. Thioether WR308278, ether WR308622, and
benzyl amine WR308396 were more potent and demonstrated a
superior selectivity index compared to mefloquine. In this series,
the ethers, thioethers, and secondary amines proved most promis-
ing, and second generation libraries based on these lead com-
pounds are currently being constructed.
After constructing a small series of cyclic diamines, we observed
promising activity emerging within the series as illustrated by
WR308621 and WR319535, which correlate with WR308396
(Table 6). Likewise, a second generation library based upon cyclic
motifs is underway.
Representative compounds from this library were evaluated for
their in vivo efficacy against P. berghei in a mouse model. The bidi-
rectional permeability of most of the compounds was measured
across MDR1-transformed MDCK cell monolayers in the presence
of the PgP inhibitor cyclosporin A. This assay is normally used as
an indicator for permeability properties across the blood–brain
barrier.
displayed a one-log increase in selectivity. The primary amine
WR308314 is nearly devoid of activity, while the addition of
methyl, ethyl, propyl, and butyl groups substantially increases effi-
cacy (Table 4). Interestingly, branched alkyl substituents such as i-
Pr and i-Bu prove quite active, while t-Bu displays moderate activ-
ity. Chain length also appears to affect activity since n-Bu
(WR177000) and n-hexyl (WR308442) derivatives have different
levels of potency, presumably resulting from the addition of the
two methylene units. N-Methyl (WR308245), i-Pr (WR308257),
and t-Bu (WR183545) derivatives all displayed favorable metabolic
stability profiles, presumably due to inhibition of N-dealkylation.18
A variety of alkyl amino quinoline methanols containing side
chains with additional heteroatoms were also constructed (Table
5). In this series, it became apparent that efficacy is reduced by
the addition of an alcohol or fluorine within the side chain. In par-
ticular, when the hydroxyl group of WR308258 is transformed into
the methyl ether (WR308412), potency increases by nearly an or-
Table 4
P. falciparum IC90 (ng/mL) values and selectivity of alkyl amino quinoline methanols (AAQMs)
R1
R2
LC50 (ng/mL)
IC90 (ng/mL)
Mousec 1/2 life (mm)
a
b
Compound
WR308314
WR308245
WR308246
WR308254
WR183544
WR308257
WR308277
WR177000
WR176990
WR308607
WR183545
WR308442
H
H
Me
Et
n-Pr
H
n-Pr
H
n-Bu
H
H
H
H
Me
Me
Et
n-Pr
i-Pr
n-Pr
n-Bu
n-Bu
i-Bu
i-Bu
n-hex
8074
470, 400, >500, >500 ND
17, 45, 58, 80
20, 69,79, 173
9, 26, 37, 67
1,7, 14, 16
5,23,23,39
5, 20, 20, 30
2, 7, 9, 12
2, 11,27, 19
2, 13, 15,21
13,667
24,626
2092
>60
ND
ND
ND
>60
ND
ND
ND
27
NDd
5201
33,160
2965
20,075
27,117
10,459
NDd
19, 101, 137, 154
124, 372, 409, 481
>60
ND
a
b
c
Cytotoxicity: macrophage IC50 (ng/mL).
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235, and C2A.
Half life in mouse liver micosomes (min).
ND = no data.
d

1350
E. Milner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1347–1351
Table 5
consideration, only WR308396 and WR319535/WR309581 exhib-
ited efficacy in vivo after oral dosing, lower or equivalent perme-
ability across MDCK cell monolayers relative to mefloquine, and
reduced permeability across MDCK cell monolayers relative to
lipophilic alkyl compounds such as WR177000.
P. falciparum IC90 (ng/mL) values and selectivity of alkyl amino quinoline methanols
containing additional heteroatoms within the side-chain)
Structure–activity relationships were studied amongst a library
of 200 4-position analogs of mefloquine. The 4-position alcohol and
at least one diamine met the minimum requirement for equivalent
potency to mefloquine. Decreased electron density around the
homobenzylic side chain amine greatly diminished potency. Mod-
erately enhanced potency, increased selectivity, and altered cross-
susceptibility patterns were achieved with imidazole-containing
analogs. Ten-fold greater potency and selectivity than mefloquine
without altered cross-susceptibility patterns were achievable in
analogs with alkyl side chains. Introduction of heteroatoms into
these latter analogs generally reduced potency, although select
analogs exhibited equivalent potency to mefloquine.
Taken together, these observations suggest a hypothesis for fur-
ther rational lead optimization: Diamino-QMs should retain the
potency of mefloquine but exhibit significantly reduced permeabil-
ity across the blood–brain barrier. Second-generation libraries that
test this hypothesis are currently underway. Due to the breadth of
this library, the compounds were synthesized as racemates. We are
currently optimizing a cost-effective, enantiopure synthetic route
to epoxide 4. The active lead compounds will be tested in vitro
and in vivo as pure enantiomers or diastereomers, respectfully.
a
b
Compound
n
R1
R2
LC50 (ng/mL)
IC90 (ng/mL)
WR308258
WR308412
WR308622
WR308278
WR308247
WR308411
WR308396
1
1
2
2
1
0
1
H
H
H
H
H
H
H
OH
NDd
260, 490, 485, 388
15, 59, 93, 100
12, 51, 84, 86
10, 36, 39, 54
>500c
OMe
OMe
SMe
F
CF3
NHBn
38,230
25,167
8618
NDd
32,053
8279
>500c
6, 29, 40, 48
a
Cytotoxicity: macrophage IC50 (ng/mL).
b
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235,
and C2A.
c
>500 ng/mL for each of four strains.
ND = no data.
d
Table 6
P. falciparum IC90 (ng/mL) values and selectivity of selected diamines
Acknowledgments
We thank the Military Infectious Diseases Research Program
(MIDRP) for financial support.
This manuscript was reviewed by the Walter Reed Army Insti-
tute of Research and the US Army Medical Research and Materiel
Command, and there is no objection to its publication or dissemi-
nation. The opinions expressed herein are those of the authors and
do not necessarily reflect the views or opinions of the Department
of the Army and the Department of Defense. Animal care and use
was conducted under IACUC protocol # 06-05 approved by the
Institutional Animal Care and Use Committee in accordance with
national guidelines. The in vivo studies were conducted at
USAMC-AFRIMS in an Association for Assessment and Accredita-
tion of Laboratory Animal Care (AAALAC) international accredited
animal facility. Research was conducted in compliance with the
Animal Welfare Act, other federal statutes and regulations that re-
late to animals and experiments involving animals, and principles
stated in the ‘Guide for the Care and Use of Laboratory Animals
(Guide for Care and Use of Laboratory Animals. National Academy
Press, 1996).
a
b
Compound
WR308621
R
LC50 (ng/mL)
IC90 (ng/mL)
21,790
4110
46, 92, 100, 198
275, 75, 260, 251
WR319535
a
Cytotoxicity: macrophage IC50 (ng/mL).
b
In vitro IC90 values against four different strains of P. falciparum, W2, D6, C235,
and C2A.
As illustrated in Table 7, WR308278, WR177000, and
WR308245 were active in vivo after oral dosing but exhibited
greater permeability relative to mefloquine across MDCK cell mon-
olayers. The latter property suggests greater propensity for accu-
mulation into the central nervous system. Taking all data into
Table 7
Permeability across MDR1-transfected MDCK Cell monolayersa and in vivo efficacy of
selected quinoline methanols in the blood schizonticidal P. berghei–ICR mouse modelb
Supplementary data
Compound
Permeability efficacy
In Miceb
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.01.001.
A–Ba
Racemic mefloquine
WR177000
WR308245
WR308278
WR308396
WR308412
WR308413
WR308622
WR308446
9.4
25
24
38
8.4
32
17
8
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25
17
10
15
2
9
3
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