Development of Biarylalkyl Carboxylic Acid Amides with Improved Anti-schistosomal Activity

Article abstract of DOI:10.1002/cmdc.201900423

The parasitic disease schistosomiasis is the cause of more than 200 000 human deaths per year. Although the disease is treatable, there is one major shortcoming: praziquantel has been the only drug used to combat these parasites since 1977. The risk of the emergence of resistant schistosomes is known to be increasing, as a reduced sensitivity of these parasites toward praziquantel has been observed. We developed a new class of substances, which are derived from inhibitors of human aldose reductase, and which showed promising activity against Schistosoma mansoni couples in vitro. Further optimisation of the compounds led to an increase in anti-schistosomal activity with observed phenotypes such as reduced egg production, vitality, and motility as well as tegumental damage and gut dilatation. Here, we performed structure–activity relationship studies on the carboxylic acid moiety of biarylalkyl carboxylic acids. Out of 82 carboxylic acid amides, we identified 10 compounds that are active against S. mansoni at 25 μm. The best five compounds showed an anti-schistosomal activity up to 10 μm and induced severe phenotypes. Cytotoxicity tests in human cell lines showed that two derivatives had no cytotoxicity at 50 or 100 μm. These compounds are promising candidates for further optimisation toward the new anti-schistosomal agents.

Full text of DOI:10.1002/cmdc.201900423

DOI: 10.1002/cmdc.201900423
Full Papers
Development of Biarylalkyl Carboxylic Acid Amides with
Improved Anti-schistosomal Activity
Alejandra M. Peter Ventura,[a] Simone Haeberlein,[b] Kerstin Lange-Grꢀnweller,[a]
Arnold Grꢀnweller,[a] Roland K. Hartmann,[a] Christoph G. Grevelding,*[b] and
Martin Schlitzer*[a]
The parasitic disease schistosomiasis is the cause of more than
200000 human deaths per year. Although the disease is treata-
ble, there is one major shortcoming: praziquantel has been
the only drug used to combat these parasites since 1977. The
risk of the emergence of resistant schistosomes is known to be
increasing, as a reduced sensitivity of these parasites toward
praziquantel has been observed. We developed a new class of
substances, which are derived from inhibitors of human aldose
reductase, and which showed promising activity against Schis-
tosoma mansoni couples in vitro. Further optimisation of the
compounds led to an increase in anti-schistosomal activity
with observed phenotypes such as reduced egg production, vi-
tality, and motility as well as tegumental damage and gut dila-
tation. Here, we performed structure–activity relationship stud-
ies on the carboxylic acid moiety of biarylalkyl carboxylic acids.
Out of 82 carboxylic acid amides, we identified 10 compounds
that are active against S. mansoni at 25 mm. The best five com-
pounds showed an anti-schistosomal activity up to 10 mm and
induced severe phenotypes. Cytotoxicity tests in human cell
lines showed that two derivatives had no cytotoxicity at 50 or
100 mm. These compounds are promising candidates for fur-
ther optimisation toward the new anti-schistosomal agents.
Introduction
In recent years, the neglected tropical disease schistosomiasis
has gained attention as more than 206 million people were in-
fected in 2016, and 800 million people are at risk of being in-
fected.[1–3] This disease is caused by parasitic flatworms of the
genus Schistosoma, which are characterised by their sexual di-
morphism. Female and male worms pair once they are fully de-
veloped. Depending on the species, each pair produces be-
tween 300 and approximately 3000 eggs per day.[4] These eggs
either remain within the host and cause the symptoms of the
disease, or are excreted into the environment. Upon exposure
to fresh water, miracidia hatch from the eggs and seek an in-
termediate host (fresh water snails), in which they develop into
cercariae, the infectious stage of the parasite’s life cycle. After
penetrating the final host, cercariae develop into schistosomu-
la (juvenile stage) and later into adult worms.[2,5] With respect
to their relevance for human health, three species are most
common: S. mansoni, S. japonicum and S. haematobium. All
three species localise in different parts of the body but cause
similar symptoms.[5] If not released in feces (S. mansoni, S. japo-
nicum) or urine (S. haematobium) into the environment, re-
maining eggs get trapped in host tissues, such as liver and
spleen, causing severe inflammation, periportal fibrosis, anae-
mia, and/or haematuria.[6]
Praziquantel (PZQ) is the only drug that is widely used
against all schistosome species (Figure 1).[2] An advantage of
this drug is its efficacy toward all schisto-
some species and its low cost for treat-
ment.[7] If 60 mgkgÀ1 of PZQ is adminis-
tered orally on three consecutive days, a
90% cure rate can be achieved.[8] Notably,
the drug mainly affects adult worms but
not the juvenile stage.[9] As this drug has
been used since the 1970s, the hazard of
Figure 1. Structure
of the anti-schisto-
emerging resistance is growing. Further-
somal drug prazi-
quantel (PZQ).
more, a lower sensitivity of schistosomes
toward this drug has been reported.[10,11]
Therefore, the development of new poten-
tial anti-schistosomal agents is urgently needed.[2]
The schistosomal aldose reductase (AR) is thought to play
an important role in antioxidant pathways and protection of
the worms from attack by its host’s reactive oxygen spe-
cies.[12,13] This makes schistosomal AR an interesting target for
novel anti-schistosomal inhibitors. Furthermore, one ortho-
logue of AR has been identified in the genome of S. mansoni
(Smp_053220).[14,15] Therefore, inhibitors of human AR were
tested against S. mansoni couples in our previous studies
(Figure 2).[16,17]
[a] A. M. Peter Ventura, Dr. K. Lange-Grꢀnweller, Prof. Dr. A. Grꢀnweller,
Prof. Dr. R. K. Hartmann, Prof. Dr. M. Schlitzer
Department of Pharmaceutical Chemistry, Philipps Universitꢁt Marburg,
Marbacher Weg 6, 35032 Marburg (Germany)
E-mail: schlitzer@staff.uni-marburg.de
[b] Dr. S. Haeberlein, Prof. Dr. C. G. Grevelding
BFS, Institute of Parasitology, Justus-Liebig-Universitꢁt Gießen, Schubert-
strasse 81, 35392 Gießen (Germany)
E-mail: christoph.grevelding@vetmed.uni-giessen.de
Supporting information and the ORCID identification number(s) for the
author(s) of this article can be found under:
https://doi.org/10.1002/cmdc.201900423.
Variation of the R substituent of the terminal phenyl moiety
showed that structures without electron-withdrawing substitu-
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sequence. Consequently, a higher yield was obtained for the
previously described biarylalkyl carboxylic acid 2a. First, a Frie-
del–Crafts acylation with 2-bromothiophene (4) and glutaric
anhydride was performed, affording precursor 5. This was sub-
sequently used in a Suzuki reaction, which afforded the previ-
ously described carboxylic acid 2a in a total yield of 77% over
both steps. The final step was coupling of 2a with the appro-
priate amines to yield the desired carboxylic amides 6a–46a.
In addition, the carboxylic acid amides were synthesised
without the carbonyl group in the linker to compare their bio-
logical activity. This was achieved by one additional step: the
reduction of 2a using a Wolff–Kishner protocol leading to the
carboxylic acid 2b. Subsequent coupling with the appropriate
amine afforded the desired products 6b–46b.
Figure 2. Inhibitors of human aldose reductase that were initially tested on
adult S. mansoni couples.[17]
ents had anti-schistosomal activity.[17] This is in contrast to in-
hibitors of human AR for which a NO2 group conferred best ac-
tivity.[16] Furthermore, good anti-schistosomal activity was ob-
served with the diethylamido compounds 3a and 3b
(Figure 3). Because most AR inhibitors have an acidic function-
ality, AR appears unlikely as a potential target structure.[17–19]
Nevertheless, we chose structures 3a and 3b (Figure 3) as our
starting point for further structure–activity relationship (SAR)
studies toward novel anti-schistosomal agents, in which the N-
substituents were varied.
The synthesis of the hydroxamic acid 48b was achieved by
in situ activation of the carboxylic acid moiety as an acid chlo-
ride and coupling with O-benzylhydroxylamine to afford 47b
(Scheme 2). After the removal of the benzyl protecting group,
the desired product 48b was obtained.
Biological evaluation and structure–activity relationships
The anti-schistosomal activity was determined using an estab-
lished assay.[17,20,21] Ten adult S. mansoni couples were exposed
to the respective compound in the medium at an initial con-
centration of 25 mm and analysed by bright-field microscopy
every 24 h for a total time of 72 h. Phenotypes such as worm
motility, pairing stability, egg production, tegumental damage,
and morphological aberrations including gut dilatation were
observed. For negative and positive controls, worms were
treated either with DMSO or PZQ, respectively. Furthermore,
the cytotoxicity of the most active compounds was assessed
by measuring the proliferation of HepG2 and LS174T cells ac-
cording to the instructions of the manufacturer (Roche, Mann-
heim, Germany, WST assay).[22]
Figure 3. The best biarylalkyl carboxylic acid derivatives from previous stud-
ies.[17]
Results and Discussion
Chemistry
The compounds used as references in previous studies,[17]
2a/b and 3a/b (Figure 3), were selected in order to compare
the extent of the phenotypes to those of the newly synthes-
ised compounds. Unexpectedly, the previous results with these
The biarylalkyl carboxylic acid amides were synthesised in
three steps (Scheme 1). In contrast to our previous proce-
dures,[16,17] we modified the synthesis by changing the reaction
Scheme 1. Modified synthesis of the biarylalkyl carboxylic acid derivatives: a) glutaric anhydride, AlCl3 in CH2Cl2, 08C to RT, overnight; b) 3-hydroxyphenylbor-
onic acid, K3PO4, Pd(OAc)2, EtOH/H2O, RT, overnight; c) amine, HOBt, EDC·HCl, NEt3, CH2Cl2, 08C to RT, overnight; d) hydrazine monohydrate, KOH, diethylene
glycol, 1808C, overnight.
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Derivatisation of the original compounds began
by varying the alkyl groups at the amide nitrogen
(Table 1). Lengthening the dialkyl chains of the
amide led to a significant reduction of egg produc-
tion (by 90% to 98%) after 72 h at 25 mm (Table 1).
Microscopic analysis of the worms revealed no effect
on motility or morphological phenotypes. Interest-
ingly, we observed a higher decrease in egg produc-
tion with compounds lacking the keto group except
for the dibutylamides 8a and 8b. The best result
was obtained with 9b, leading to a 98% reduction
in egg numbers after 72 h. These results led to the
assumption that the keto group within the alkyl
chain is not necessary to reduce the numbers of
eggs produced. In contrast to the N,N-disubstituted
Scheme 2. Synthesis of the hydroxamic acid derivative 48b: a) 1. oxalylchloride, DMF,
CH2Cl2, RT, 3 h; 2. O-benzylhydroxylamine, NEt3, CH2Cl2, RT, overnight; b) Pd/C, H2 atmos-
phere, MeOH, overnight.
compounds could not be completely reproduced. Phenotypes
obtained with 2a/b and 3a/b did not occur to the same
extent or at similar concentrations as before. Whereas a de-
crease in egg production was again found for 3b, on this occa-
sion, other effects were not observed with these compounds.
Because different sources of the added in vitro culture supple-
ments were used compared to the previous study (newborn
calf serum, HEPES and antibiotic–antimycotic; see the
Experimental Section), we tested their influence. A
slight influence was observed on egg production,
which was more affected by these compounds if the
supplements were missing (data not shown). Other
parameters such as vitality, motility, and pairing sta-
bility were unaffected. Thus, it is unclear why the re-
sults with 2a/b and 3a/b differed from the former
amides, N-monosubstituted amides showed no significant ac-
tivity on egg production. In this respect, hydroxamic acid de-
rivatives 47b and 48b were also inactive.
Glucose transporters and the amino acid transporter
SPRM1lc have been discovered in the tegument of schisto-
somes.[23–26] This indicates that nutrients such as glucose and
amino acids can be taken up by the schistosomes not only
Figure 4. General structures of the amino acid derivatives 13a–26a and 13b–26b.
study.
through their mouth but also through their tegument.[23–26]
Therefore, we coupled amino acids (Figure 4, 13a–26a and
13b–26b) with the carboxyl moiety to use them as “trojan
horses”, possibly favouring the uptake of the compounds by
schistosomes. As summarised in Table S1 (Supporting Informa-
tion), this approach failed to provide the desired results. None
of the amino acid derivatives showed anti-schistosomal activity
at a concentration of 25 mm after 72 h.
Table 1. Alkyl amides and phenotype observation.
By variation of the amide dialkyl chain length, different com-
pounds with an inhibitory effect on egg production were
found. However, no effects on vitality, motility and pairing sta-
bility of the adult S. mansoni couples were found using this
series. Mono-substituted amides and amino acids showed no
activity at 25 mm.
To increase rigidity of the amide, piperidine derivatives
(Figure 5, 27a–30a and 27b–30b) were synthesised. This var-
iation also provided no apparent anti-schistosomal activity at
25 mm (Table S2).
In summary, dialkylamines showed negligible activity, as did
mono-substituted alkylamines, amino acids, and piperidine de-
rivatives. Next, we examined whether an additional heteroa-
tom in the piperidine moiety would increase activity (Table 2).
After treating the worms with such compounds for 72 h at
25 mm, we observed remarkable reduction in egg production.
In addition, we observed separation of worm couples, reduced
Compd
R
Activity [mm][a]
Reduction in egg number
6a
7a
7b
8a
8b
N(CH3)2
N(C3H7)2
n.a.
n.a.
25
25
25
–
–
90%
96%
90%
90%
98%
–
–
–
–
–
N(C4H9)2
N(C5H11)2
9a
9b
25
25
47b
48b
10a
10b
11 a
11 b
12a
12b
NHOBzl
NHOH
n.a.
n.a.
n.a.
n.a.
n.a.
25
NHC2H5
NHC4H9
NCH3Bzl
81%
–
–
n.a.
n.a.
[a] Activity measured at 25 mm, values were determined in single experi-
ments, and are the lowest concentration at which an effect on egg pro-
duction was observed; n.a.=not active. Bzl=benzyl. n=1.
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Table 2. Phenotype observation for piperazine derivatives.
Figure 5. General structures of the piperidine derivatives 27a–30a and
27b–30b.
Compd
R
Activity
[mm][a]
Phenotypes
31a
31b
n.a.
25
–
100% e.r., 100% c.s., teg. dam.
!
motility, and tegumental damage (Figure 6A) and/or severe
gut dilatation (Figure 6B) for most derivatives.
32a
32b
n.a.
n.a.
–
–
33a
33b
n.a.
n.a.
–
–
34a
34b
n.a.
n.a.
–
–
35a
35b
n.a.
n.a.
–
–
36a
36b
n.a.
n.a.
–
–
37a
37b
n.a.
n.a.
–
–
38a
38b
n.a.
n.a.
–
–
Figure 6. Photographs of S. mansoni worms after 72 h treatment with the
following compounds (25 mm): A) 46b, arrows show tegumental damage;
B) 39b, arrows show gut dilatation; C) control, untreated S. mansoni couple.
99% e.r., 90% c.s., gut dil. !
and ?
85% e.r.
100% e.r., 80% c.s., gut dil. !
97% e.r., gut dil. !
39a
25
39b
40a
40b
25
25
25
Treating worms with the phenylsulfonyl piperazine com-
pound 44b provoked gut dilatation in the female and tegu-
mental damage of both female and male worms. The Boc-pro-
tected piperazine derivative 41b and the deprotected pipera-
zine analogue 42b showed similar phenotypes at 25 mm. Inter-
estingly, these phenotypes were observed with the Boc-pro-
tected 41b after prolonged treatment. This might be
explained by the Boc-protected piperazine acting as a prodrug
being slowly transformed into its active amine form.
41a
41b
n.a.
25
–
100% e.r., 70% c.s., gut dil. !
42a
42b
n.a.
25
–
100% e.r, 90% c.s, gut dil. ! and
?
–
–
43a
43b
n.a.
n.a.
Because the compound containing an unprotected pipera-
zine moiety (42b) showed promising results, we exchanged
the distal nitrogen atom with an oxygen or sulfur atom that is,
exchanging piperazine for morpholine and thiomorpholine
groups, respectively (Table 3). These modifications resulted in
active compounds. Both the morpholine (45b) and thiomor-
pholine (46b) derivatives without the carbonyl linker were
active at 25 mm. Treatment of worms with the morpholine de-
rivative 45b at 25 mm led to a 98% inhibition of egg produc-
tion and 40% decreased pairing stability. Also, at 10 mm, this
compound was also active although the phenotypes were of
lower intensity.
44a
44b
n.a.
25
–
100% e.r., 100% c.s., teg. dam.
! and ?, gut dil. !
[a] Activity measured at 25 mm; values are the lowest concentration at
which anti-schistosomal activity was observed. n.a.=not active at 25 mm;
e.r.=egg number reduction; c.s.=couple separation; gut dil.=gut dilata-
tion; teg. dam.=tegumental damage. n=3 for active compounds.
Compound 46b was active at a concentration of 25 mm. The
worms showed the same phenotypes as those treated with
45b. In addition, we observed tegumental damage (Figure 6A)
of both male and female worms. Notably, only the compounds
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promising anti-schistosomal compounds. Our results indicate
that the carboxylic acid amides possess higher activity than
the corresponding free carboxylic acid. Because the carboxylic
acid is an important moiety for inhibiting the human AR,[29] we
think that the schistosome AR might not be the target for
these compounds. Evidently, the keto carbonyl group is not es-
sential for anti-schistosomal activity, as its removal led to a
higher activity in most cases. Out of 82 carboxylic acid amides,
we identified the best six, which showed activity as low as
10 mm. These six compounds were tested for their cytotoxicity
in two cell lines (HepG2 and LS174T). The results showed that
two compounds were non-cytotoxic at concentrations of up to
100 mm in both cell lines. Therefore, these two compounds,
the morpholine derivative 45b and the methylsulfonyl pipera-
zine derivative 31b, are the most interesting starting points for
further optimisation. Selected compounds will be assayed
against other pathogens such as S. japonicum and Fasciola
hepatica. In addition, the most active compounds will be eval-
uated for their DMPK data to facilitate subsequent in vivo test-
ing. Furthermore, additional SAR studies will be performed, for
example, variation of the thiophene ring.
Table 3. Phenotype observation for morpholine and thiomorpholine de-
rivatives.
Compd
R
Activity [mm][a]
Phenotypes
45a
45b
n.a.
10
–
73% e.r., 50% c.s.
46a
46b
n.a.
25
–
100% e.r., 100% c.s., teg. dam.
[a] Activity measured at 25 mm; values are the lowest concentration at
which anti-schistosomal activity was observed. n.a.=not active. e.r.=egg
number reduction; c.s.=couple separation; gut dil.=gut dilatation; teg.
dam.=tegumental damage. n=3.
without the carbonyl group in the linker exhibited anti-schisto-
somal activity. This result is consistent with the other results in
this series. We assume that the carbonyl group in the linker
might not be required, and it even lowers the anti-schistoso-
mal effects.
Of all the prepared series, 10 compounds active at 10 and/or
25 mm were obtained. The six most active compounds (39a,
31b, 42b, 44b, 45b and 46b) were further tested for their cy-
totoxicity in HepG2 and LS174T cells (Table 4). Importantly, two
compounds (31b and 45b) showed no cytotoxicity at 100 mm
in either cell line. Compound 39a was cytotoxic only at
100 mm but not at 50 mm in both cell lines. Interestingly, 44b
had no cytotoxicity at 100 mm in LS174T cells, however it was
cytotoxic at 50 mm in HepG2 cells. Based on these results, 31b
and 45b were selected for further evaluation as the most
promising candidates.
Experimental Section
Biological methods
In vitro assay on S. mansoni couples: Animal experiments were
approved by the Regierungsprꢂsidium (regional council) Giessen
(V54-19 c20/15 cGI18/10) and were performed in accordance with
the European Convention for the Protection of Vertebrate Animals
guidelines on the use of animals for experimental and other scien-
tific purposes (ETS No. 123; revised Appendix A).
Forty-six days after infection of Syrian hamsters (Mesocricetus aura-
tus) with cercariae, S. mansoni couples were obtained by hepato-
portal perfusion and incubated in groups of 20 couples in 60 mm
diameter Petri dishes (Greiner Bio-One) with M199 medium (Gibco)
supplemented with 10% newborn calf serum (NCS, Sigma), 1%
HEPES (1m, Roth), and 1% antibiotic–antimycotic (ABAM, GE
Healthcare; 10000 units penicillin, 10 mg streptomycin, and 25 mg
amphotericin B per mL) at 378C with 5% CO2. Twenty-four hours
after the perfusion, in vitro culture experiments were started. Six-
well plates (Greiner Bio-One) were filled with 10 couples per well
and a total volume of 5 mL (medium+compound). All substances
were dissolved as 10 mm stock solutions in DMSO (Sigma–Aldrich).
The substances were initially tested at 25 mm. Therefore, the latter
were dissolved in DMSO. If a positive result was achieved, the ex-
periment was repeated at a lower concentration of that substance
(10 mm). DMSO (25 mm) was used in every experiment as the nega-
tive control, whereas PZQ (5 mm) served as a positive control. Every
24 h, new plates with medium and substance were prepared. After
evaluating the worms for their pairing stability and phenotypes
(such as gut peristalsis, vitality and motility) under bright-field mi-
croscopy, the worms were transferred to the newly prepared six-
well plates. Then, the eggs produced during the preceding 24 h
were counted. Compounds showing only a reduction in egg pro-
duction were not regarded as active. Compounds showing at least
one phenotype in addition to egg number reduction were regard-
ed as active and re-tested twice at different days. For active com-
pounds, three independent test experiments were performed.
Table 4. Cytotoxicity of the top six compounds in HepG2 and LS174T
cells.
Cytotoxicity
Compd
Activity [mm][a]
HepG2
LS174T
39a
31b
44b
42b
45b
46b
25
25
25
25
10
25
50<x<100
>100
<50
50ꢀx<100
ꢁ100
ꢁ100
n.d.
<50
>100
<50
ꢁ100
n.d.
[a] Activity measured at 25 mm; values are the lowest concentration at
which anti-schistosomal activity was observed. n.d.=not determined.
Conclusions
Schistosomiasis represents a worldwide threat for humans and
animals. Lowered PZQ sensitivity in treated patients has been
observed, provoking the fear of future resistance,[10,11,27,28]
therefore alternative medications are needed. A new approach
originating from potential inhibitors of human AR revealed
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Cytotoxicity measurements: The two cell lines tested, HepG2
(human hepatoma) and LS174T (colon carcinoma), were each cul-
tured in Iscove’s modified Dulbecco medium (IMDM), l-glutamine
(5 mm) and 10% fetal calf serum (FCS) at 378C and 5% CO2. A WST-
1 assay was performed for both cell lines. In brief, 1ꢃ104 cells in
IMDM (200 mL, containing 10% FCS) were seeded in 96-well micro-
plates and incubated for 24 h (378C, 5% CO2). Afterward, old
medium was replaced by fresh medium containing the respective
compound (50 or 100 mm), and the cells were incubated for up to
144 h. DMSO-treated cells served as controls. Samples were mea-
sured after 24, 72, 120, and 144 h (n=3 for each time point and
concentration). After the removal of the medium, 10% WST-1 re-
agent (110 mL in phosphate-buffered saline; Roche, Mannheim,
Germany) was added, and the absorbance at 450 nm was mea-
sured after 2 h by using a Tecan Safire II plate reader.
concentrated HCl. The precipitate was filtered, washed with H2O
and EtOH and dried to afford 2a as a light-brown solid (5.00 g,
1
17.2 mmol, 88%). H NMR ([D6]DMSO, 400 MHz): d=12.03 (s, 1H),
3
3
9.73 (s, 1H), 7.91 (d, J=4.0 Hz, 1H), 7.55 (d, J=3.9 Hz, 1H), 7.26
3
3
4
(t, J=7.9 Hz, 1H), 7.19 (ddd, J=7.7 Hz, J=1.7, 1.0 Hz, 1H), 7.11
3
3
4
(t, J=2.0 Hz, 1H), 6.81 (ddd, J=8.1 Hz, J=2.4, 1.0 Hz, 1H), 2.99
(t, 3J=7.3 Hz, 2H), 2.31 (t, 3J=7.3 Hz, 2H), 1.85 (quin, 3J=7.5 Hz,
2H); 13C NMR ([D6]DMSO, 100 MHz): d=192.5, 174.1, 157.9, 151.3,
142.1, 134.2, 133.8, 130.4, 124.8, 116.8, 116.3, 112.6, 37.2, 32.8,
19.6 ppm; MS (ESI+) m/z (%): 291 (100) [M+H]+; HRMS (ESI+): m/
z: calcd for C15H15O4S: 291.0686 [M+H]+; found: 291.0677; calcd
for C15H14O4SNa: 313.0505 [M+Na]+; found: 313.0497.
5-[5-(3-Hydroxyphenyl)thiophene-2-yl]pentanoic acid (2b):[17] The
keto carboxylic acid 2a (2.50 g, 8.61 mmol, 1.00 equiv), hydrazine
Chemistry
5-(5-Bromothiophene-2-yl)-5-oxopentanoic acid (5):[17] Under an
argon atmosphere, 2-bromothiophene (4, 1.00 g, 6.13 mmol,
monohydrate (1.68 mL, 34.4 mmol, 4.00 equiv) and KOH (1.93 g,
34.4 mmol, 4.00 equiv) were suspended in diethylene glycol
(35.0 mL) and heated at reflux for 21 h. The reaction was quenched
with H2O (200 mL) and the aqueous phase was extracted with
EtOAc. The aqueous phase was acidified with 10% aq HCl solution
and extracted with EtOAc. The combined organic layers were
washed with 1m NaOH and the aqueous phase was acidified with
concentrated HCl. The precipitate was filtered, washed with H2O
and dried to afford 2b as a brown solid (2.10 g, 7.59 mmol, 88%).
1H NMR ([D6]DMSO, 400 MHz): d=11.98 (s, 1H), 9.49 (s, 1H), 7.22
1.0 equiv) and glutaric anhydride (0.70 g, 6.13 mmol,1.0 equiv)
were dissolved in anhydrous CH2Cl2 (40 mL) and cooled to 08C.
After the addition of AlCl3 (1.80 g, 13.5 mmol, 2.2 equiv), the reac-
tion mixture was stirred at 08C for 5 h and at RT overnight. The re-
action was quenched by the addition of 10% aq HCl (60 mL) and
the mixture was stirred at RT for 1 h. The aqueous phase was ex-
tracted with CH2Cl2 and the combined organic layers were washed
with 1m NaOH. The aqueous phase was acidified with concentrat-
ed HCl, and the precipitate was filtered, washed with demineral-
ised H2O and dried to afford 5 as an orange solid (1.50 g,
5.44 mmol, 89%), which was used without further purification.
3
3
3
(d, J=3.6 Hz, 1H), 7.17 (t, J=7.9 Hz, 1H), 7.01 (d, J=7.8 Hz, 1H),
6.95 (s, 1H), 6.82 (d, 3J=3.6 Hz, 1H), 6.67 (dd, 3J=8.5 Hz, 4J=
3
3
1.8 Hz, 1H), 2.79 (t, J=7.3 Hz, 2H), 2.25 (t, J=7.0 Hz, 2H), 1.67–
1.53 ppm (m, 4H); 13C NMR ([D6]DMSO, 100 MHz): d=174.3, 157.8,
144.3, 140.9, 135.1, 130.0, 125.6, 123.0, 115.8, 114.3, 111.7, 33.3,
30.5, 29.1, 23.9 ppm; MS (ESI+): m/z (%): 277 (75) [M+H]+, 294
(100) M+NH4]+, 299 (20) [M+Na]+; HRMS (ESI+): m/z: calcd for
C15H16O3SNa: 299.0712 [M+Na]+; found: 299.0709.
3
1H NMR ([D6]DMSO, 400 MHz): d=12.03 (s, 1H), 7.78 (d, J=4.1 Hz,
1H), 7.39 (d, 3J=4.0 Hz, 1H), 2.95 (t, 3J=7.2 Hz, 2H), 2.28 (t, 3J=
7.6 Hz, 2H), 1.81 ppm (quin, 3J=7.3 Hz, 2H); 13C NMR ([D6]DMSO,
100 MHz): d=192.2, 174.2, 145.4, 134.0, 132.4, 121.6, 36.9, 32.7,
19.4 ppm; MS (ESI+): m/z (%): 277 (10) [M+H]+, 294 (100) [M+
NH4]+; HRMS (ESIÀ): m/z: calcd for C9H8O3SBr: 274.9383 [MÀH]À;
found: 274.9374.
General procedure 1A: synthesis of carboxylic acid amides:[17]
The carboxylic acid (1.0 equiv), amine (1.0 equiv) and NEt3
(3.0 equiv) were dissolved in CH2Cl2 and stirred at RT for 5 min. The
mixture was cooled to 08C and EDC·HCl (1.5 equiv) and HOBt
(1.5 equiv) were added. Then, the mixture was stirred at RT over-
night. The solvent was removed under reduced pressure and the
crude product purified by column chromatography.
5-[5-(3-Hydroxyphenyl)thiophene-2-yl]-5-oxopentanoic
acid
(2a):[17] Under an argon atmosphere,
5
(5.38 g, 19.5 mmol,
General procedure 1B: synthesis of carboxylic acid amides: The
carboxylic acid (1 equiv) was dissolved in CH2Cl2. Oxalyl chloride
(1.5 equiv) and DMF (cat. amount) were added. The reaction mix-
ture was stirred for 2 h at RT. The volatile compounds were re-
moved under reduced pressure and a mixture of NEt3 (1.5 equiv)
and amine (1.5 equiv) in CH2Cl2 was added. The reaction was
stirred at RT overnight, diluted with CH2Cl2, washed with aq HCl
(1m) and saturated NaHCO3 solution, and dried over MgSO4. The
solvent was removed under reduced pressure and the crude prod-
uct purified by column chromatography.
1.2 equiv), 3-hydroxyphenylboronic acid (3.23 g, 23.4 mmol,
1.2 equiv) and K3PO4 (8.28 g, 39.0 mmol 2.0 equiv) were suspended
in EtOH/H2O (2:1, 100 mL). After 5 min of stirring, Pd(OAc)2
(87.6 mg, 0.39 mmol, 2 mol%) was added and the mixture was
stirred at RT overnight. Aqueous HCl (1m) was added and the mix-
ture extracted with EtOAc. The combined organic layers were
washed with NaOH (1m) and the aqueous phase was acidified with
General procedure 2A: alkaline hydrolysis:[30] The ester
(1.00 equiv) was dissolved or suspended in MeOH and KOH
(3.0 equiv) was added. The reaction mixture was stirred overnight
at 358C. The reaction was quenched by the addition of H2O and
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Full Papers
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Acknowledgements
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Conflict of interest
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Keywords: biaryls
·
carboxamides
·
inhibitors
·
schistosomiasis · structure–activity relationships
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749–760.
[2] P. Mꢂder, G. Rennar, A. M. Peter Ventura, C. G. Grevelding, M. Schlitzer,
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Manuscript received: July 16, 2019
Revised manuscript received: August 9, 2019
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Accepted manuscript online: August 27, 2019
Version of record online: && &&, 0000
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FULL PAPERS
A. M. Peter Ventura, S. Haeberlein,
K. Lange-Grꢀnweller, A. Grꢀnweller,
R. K. Hartmann, C. G. Grevelding,*
M. Schlitzer*
No flukes with these amides! Schisto-
somiasis causes more than 200000
deaths per year and is treated with the
single drug praziquantel. We have previ-
ously reported biarylalkyl carboxylic acid
derivatives as novel chemical entities
with in vitro anti-schistosomal activity.
In continuation of these studies, we
report here a series of amide deriva-
tives. The best results were obtained
with morpholine, thiomorpholine and
substituted piperazine derivatives.
&& – &&
Development of Biarylalkyl Carboxylic
Acid Amides with Improved Anti-
schistosomal Activity
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ꢁ 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
ÝÝ These are not the final page numbers!