Synthesis and structure-activity relationship of disubstituted benzamides as a novel class of antimalarial agents

Article abstract of DOI:10.1016/j.bmcl.2012.05.124

Malaria is a devastating world health problem. Using a compound library screening approach, we identified a novel series of disubstituted benzamide compounds with significant activity against malaria strains 3D7 and K1. These compounds represent a new antimalarial molecular scaffold exemplified by compound 1, which demonstrated EC₅₀ values of 60 and 430 nM against strains 3D7 and K1, respectively. Herein we report our findings on the efficient synthesis, structure-activity relationships, and biological activity of this new class of antimalarial agents.

Full text of DOI:10.1016/j.bmcl.2012.05.124

Bioorganic & Medicinal Chemistry Letters 22 (2012) 4536–4539
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and structure–activity relationship of disubstituted benzamides
as a novel class of antimalarial agents
⇑
Katsuhiko Mitachi, Yandira G. Salinas, Michele Connelly, Nicholas Jensen, Taotao Ling, Fatima Rivas
Department of Chemical Biology and Therapeutics, St. Jude Children’s Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105-3678, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Malaria is a devastating world health problem. Using a compound library screening approach, we iden-
tified a novel series of disubstituted benzamide compounds with significant activity against malaria
strains 3D7 and K1. These compounds represent a new antimalarial molecular scaffold exemplified by
compound 1, which demonstrated EC50 values of 60 and 430 nM against strains 3D7 and K1, respectively.
Herein we report our findings on the efficient synthesis, structure–activity relationships, and biological
activity of this new class of antimalarial agents.
Received 10 April 2012
Revised 31 May 2012
Accepted 31 May 2012
Available online 7 June 2012
Keywords:
Ó 2012 Elsevier Ltd. All rights reserved.
Plasmodium falciparum
Disubstituted benzamides
Antimalarial agents
Malaria is the world’s most deadly parasitic disease. It is caused
by a variety of plasmodial species, but only four speciesP(lasmodium
falciparum, Plasmodium vivax, Plasmodium ovale, and Plasmodium
malariae) are implicated in human disease,1 the most virulent
being P. falciparum. Malaria is endemic to 109 nations, most lo-
cated in Africa. This infectious disease disproportionately affects
poor communities that have inadequate vector control; it remains
a leading cause of death of young children.2
Despite the great world health impact of malaria, only a limited
number of therapeutic agents are available. Furthermore, the lack
of scaffold diversity poses a threat, should multi-drug–resistant
strains emerge. The current antimalarial arsenal comprises five
classes of compounds, the first two of which are chemically re-
lated: (1) 4-aminoquinolines (chloroquine, quinine), (2) 8-amino-
quinolines (primaquine), (3) proguanil, (4) atovaquone, and (5)
artemisinin derivatives.2–4 The only combination regimen cur-
rently available is artemisinin-based.1 P. falciparum is reported to
be susceptible only to artemisinin-based drugs, and resistance
can be expected to emerge with the increased use of artemisinin
derivatives.3–14 Therefore, new antimalarial agents are urgently
needed. Here we report our studies on the efficient synthesis,
structure–activity relationships, and biological activity of a new
class of disubstituted benzamide analogs (analogs 1–3, Fig. 1)
which show promising antimalarial properties.
antimalarial scaffold class whose mechanism of action may differ
from those of current agents, as it occupies a different chemical
space. All compounds were tested against both chloroquine-sensi-
tive (3D7) and chloroquine-resistant (K1) parasites by following a
known protocol.5–8 The heat map corresponding to our antimalar-
ial results and the cytotoxicity of these compounds to Raji, HepG2,
HEK293, and BJ cells can be found in Figure 4 and Supplementary
Table 1. As depicted in Fig. 4, there was a pattern of antimalarial
activity in the order of compounds 1 > 2 > 3, with compound 1
showing the greatest potency. The overall biological results ob-
tained for the isoquinoline and phenyl acetamides were not opti-
mal since they showed poor selectivity (cytotoxicity vs parasite).
However, compound 34 stood out from the group, with EC50 values
of 0.664 lM for 3D7and 0.822 lM for K1. Compound 34 showed
low cytotoxicity, but it was not further derivatized, as its isomers
31, 32, and 33 showed weaker antimalarial properties. The inhibi-
tion potency of compounds 1 and 4 did not differ substantially be-
tween 3D7 (EC50: 0.0604 and 0.1738
(EC50: 0.4377 and 0.9953 M, respectively). Compounds 2 and 5
showed some promising results (EC50 <1.8 M), but inhibited only
lM, respectively) and K1
l
l
the K1 parasite and displayed a modest therapeutic index (Fig. 4).
With this information in hand, we proceeded to develop a focus li-
brary from hit compound 1, the optimal lead compound for analy-
sis of structure–activity relationship (SAR).
A recent screen of a small library of two distinctive classes of
compounds, isoquinoline/phenethyl acetamide—type com-
pounds15 and disubstituted benzamide/aryl ether—type com-
pounds (Fig. 1), revealed compounds 1 and 4 to represent a novel
The benzamide scaffold of compound 1 is composed of an alkyl
substituent and a benzylic substituent with an ether group. We se-
lected the mono-ether linker carrying a carboxylic acid to serve as
a handle for future pull-down experiments and to enhance the sol-
ubility properties of these disubstituted benzamides. When we as-
sessed the length of the ether linker, it was found that either a
short ether linker (2–3 carbons) or a longer linker (>5 carbons)
⇑
Tel.: +1 901 595 6504; fax: +1 901 595 5715.
E-mail address: Fatima.rivas@stjude.org (F. Rivas).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.05.124

K. Mitachi et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4536–4539
4537
Figure 1. Focused library of disubstituted benzamides and isoquinoline/phenyl acetamides.
afforded unsatisfactory chemical yields so the linear 5-carbon
ether linker was selected for further SAR studies due to good phys-
ical properties such as solubility. The alkyl group on the amide was
also evaluated and the results showed that a bulky side chain was
optimal. Amongst the studied alkyl groups, n-butyl, n-propyl, and
isopropyl amine, the isopropyl group was the best candidate. As
observed in our initial screen, compound 1 was approximately
three times more potent against 3D7 malarial strain than its iso-
mer 4. The meta-linked ether position in these disubstituted benz-
amides also provided greater antimalarial activity than their
corresponding para/ortho isomers (1, 2, and 3). Together, these
disubstituted benzamide isomers provided an activity profile on
two variables; the ether linker size, and the substitution pattern
on the phenyl ring, but the role of the functional group on the
benzamide ring remained to be investigated. The main objective
was then to identify the steric and electronic factors of the aro-
matic ring that might be responsible for the observed antimalarial
activity. Compound 23 was selected as the core to be derivatized.
Efficient synthesis of the disubstituted benzamide scaffold ser-
ies was designed from commercially available meta-hydroxybenz-
aldehyde (the method could also be used with para/ortho isomers).
Figure 2 outlines our key synthetic disconnections, which consisted
of four synthetic maneuvers: Williamson etherification, reductive
alkylation, amide formation, and Suzuki coupling, utilizing
Figure 2. General synthetic disconnections.
minimal purification procedures. Parallel synthesis solution-phase
chemistry was utilized, as it offers simple and rapid synthesis of
this type of disubstituted benzamide.
Scheme 1 depicts our general synthetic approach. Synthesis
commenced with m-hydroxybenzaldehyde, which was etherified
using conventional Williamson etherification conditions16
(1.5 equiv BrCH2CO2Me, 1.3 equiv K2CO3, DMF, 90 °C, 4 h, 98%),
affording compound 23 almost quantitatively. The reaction mix-
ture was filtered, washed, and used without further purification.
The corresponding aldehyde 23 was treated with isopropyl amine
in the presence of catalytic acid (4 N HCl/EtOAc, 23 °C, 1.5 h), and
reducing agent (NaCNBH3 or NaBH4) was then directly added.17

4538
K. Mitachi et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4536–4539
Scheme 1. Efficient general synthesis of disubstituted benzamides 1–7.
The crude reaction mixture was treated with 4-bromobenzoyl
chloride, TEA, and catalytic DMAP in DCM to afford compound 13
in excellent yield. Compound 13 was dissolved in DMF, treated
with Na2CO3 and tetrakis(triphenylphosphine)palladium, and de-
gassed, and the corresponding boronic acid was added. The reac-
tion mixture was heated to 50 °C for 8 h to afford compound 7
(86% yield after silica gel purification). Saponification of compound
7 with NaOH produced compound 1 (97% yield). Compound 13 was
synthesized in multigram scale to allow development of its deriv-
atives (shown in Fig. 3).
In our follow-up focused library (Fig. 3), a broad range of both
electronic and steric factors affecting the benzamide aromatic ring
of lead compound 1 were evaluated. The chemical yields of the Su-
zuki coupling reaction and hydrolysis ranged from modest to good.
The Suzuki reactions that provided compounds 7c0–e0 and 7j0 were
less efficient, and large quantities of starting materials were recov-
ered; changes in the catalyst, solvent, and/or overall conditions did
not improve reactivity. Hydrolysis reactions of the heterocycles
(7f0–h0) were impeded by competing decomposition under basic
conditions.
Heat maps of the antimalarial activity of compounds 1–54 and
1a–7p0 are shown with their cytotoxicity profiles in Figure 4. The
results indicate that the nature of the benzamide aromatic ring
influences its antimalarial properties; for example, 1a inhibited
the K1, but not the 3D7, strain. Addition of moderately small
groups, halogens, or bulky groups to the benzamide ring, such as
phenyl, naphthyl, heterocycles (furan, thiophene, oxadiazole, in-
doles and their corresponding sulfonyl counterparts) did not sub-
stantially affect the antimalarial profile (EC50: >13 or 15
Some compounds, such as 1v, and 1l, showed good activity
(EC50: 0.0175 and 0.15015 M, respectively) against K1, but not
lM).
l
against 3D7. Interestingly, compound 1l differed from our lead
compound 1 in the size of the ether ring size (dioxole vs dioxine),
indicating the importance of spatial factors around the benzamide
Figure 3. Chemical yields for Suzuki and saponification reactions of disubstituted benzamides 1–7p0.

K. Mitachi et al. / Bioorg. Med. Chem. Lett. 22 (2012) 4536–4539
4539
Figure 4. Antimalarial activity and cytotoxicity of compounds 1–54 and 1a–7p0.
ring. Gratifyingly, this new compound library provided compound
References and notes
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Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012. 05.124.