Expanding the SAR of Nontoxic Antiplasmodial Indolyl-3-ethanone Ethers and Thioethers

Article abstract of DOI:10.1002/cmdc.201800235

Despite major strides in reducing Plasmodium falciparum infections, this parasite still accounts for roughly half a million annual deaths. This problem is compounded by the decreased efficacy of artemisinin combination therapies. Therefore, the development and optimisation of novel antimalarial chemotypes is critical. In this study, we describe our strategic approach to optimise a class of previously reported antimalarials, resulting in the discovery of 1-(5-chloro-1H-indol-3-yl)-2-[(4-cyanophenyl)thio]ethanone (13) and 1-(5-chloro-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone (14), whose activity was equipotent to that of chloroquine against the P. falciparum 3D7 strain. Furthermore, these compounds were found to be nontoxic to HeLa cells as well as being non-haemolytic to uninfected red blood cells. Intriguingly, several of our most promising compounds were found to be less active against the isogenic NF54 strain, highlighting possible issues with long-term dependability of malarial strains. Finally compound 14 displayed similar activity against both the NF54 and K1 strains, suggesting that it inhibits a pathway that is uncompromised by K1 resistance.

Full text of DOI:10.1002/cmdc.201800235

Accepted Article
Title: Expanding the SAR of non-toxic antiplasmodial indolyl-3-
ethanone ethers and thioethers
Authors: Mayibongwe J Lunga, Ruramai L Chisango, Carli Weyers,
Michelle Isaacs, Dale Taylor, Adrienne L Edkins, Setshaba D
Khanye, Heinrich C Hoppe, and Clinton Veale
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To be cited as: ChemMedChem 10.1002/cmdc.201800235
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ChemMedChem
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Expanding the SAR of non-toxic antiplasmodial indolyl-3-
ethanone ethers and thioethers
Mayibongwe J. Lunga,[a] Ruramai L. Chisango,[a] Carli Weyers,[a] Michelle Isaacs,[b] Dale Taylor,[c]
Adrienne L. Edkins,[b] Setshaba D. Khanye,[d] Heinrich C. Hoppe,[b] and Clinton G. L. Veale*[a,e]
[a]
[b]
[c]
[d]
[e]
Mr, M.J. Lunga, Ms R.L. Chisango, Ms C.W. Weyers, Dr. C.G.L. Veale
Faculty of Pharmacy
Rhodes University
Grahamstown, 6140, South Africa
Ms. M. Isaacs, Prof. A. L. Edkins, Prof. H. Hoppe
Department of Biochemistry and Microbiology
Rhodes University
Grahamstown, 6140, South Africa
Dr. D Taylor
Division of Clinical Pharmacology, Department of Medicine
University of Cape Town
Groote Schuur Hospital, Observatory, 7925, South Africa,
Dr. S.D. Khanye orcid.org/0000-0003-0725-5738
Department of Chemistry
Rhodes University
Grahamstown, 6140, South Africa
Dr. C.G.L. Veale orcid.org/0000-0002-4043-7106
Current address: School of Chemistry and Physics, Pietermaritzburg Campus
University of KwaZulu-Natal, Private Bag X01
Scottsville, 3209, South Africa
Email: VealeC@ukzn.ac.za
Supporting information for this article is given via a link at the end of the document.
of resistance to first-line treatments.[5–8] The rapid, efficacious
nature of artemisinin combination therapies (ACT) against multi–
drug resistant parasites,[9] coupled to a widespread adoption of
ACT as a first line treatment in most malaria endemic countries,[10]
has contributed to the significant progress made in reducing rates
of malarial transmission.[11] However, reports of delayed parasite
clearance after standard ACT dosing regimens, indicate that the
current trends in reducing malaria burden may be under threat,[12–
14] and highlights the urgent need to discover chemotypes which
disrupt vital malarial biochemical processes, either via new
targets, or overcoming resistance pathways in current validated
targets. The post-genomic era of drug discovery, has providing
stunning insight into biological pathways, as well as facilitating
target based drug discovery against human derived diseases.[15]
However, targeted screening approaches have proven more
challenging against infectious disease.[16] Conversely, phenotypic
screening takes advantage of the greater chemical space
sampled in the complex cellular environment, thereby increasing
the probability of unearthing new classes of biological targets, or
the discovery of active compounds which act against multiple
targets.[17–20]
Abstract: Despite major strides in reducing Plasmodium falciparum
infections, this parasite still accounts for roughly half a million annual
deaths. This problem is compounded by reduced efficacy of
artemisinin combination therapies. Therefore, the development
and optimisation of novel antimalarial chemotypes is critical. In
this study, we describe our strategic approach to optimise a class
of previously reported antimalarials, resulting in the discovery of
1-(5-Chloro-1H-indol-3-yl)-2-[(4-cyanophenyl)thio]ethanone (13)
and
1-(5-Chloro-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(14) whose activity was equipotent to chloroquine against the P.
falciparum 3D7 strain. Furthermore, these compounds were non-
toxic to HeLa cells as well as being non-haemolytic to uninfected
red blood cells. Intriguingly, several of our most promising compounds
were less active against the isogenic NF54 strain, highlighting
possible issues with long-term dependability of malarial strains.
Finally compound 14 displayed similar activity against both the NF54
and K1 strains suggesting that it inhibits a pathway, which is
uncompromised by K1 resistance.
Introduction
Plasmodium falciparum induced malaria, remains one of the most
prevalent parasitic disease worldwide.[1] The WHO reported 212
million new cases of malaria, and 429 000 deaths in 2015, with
children under five years of age and severely disadvantaged
populations being particularly vulnerable.[2–4] Chemotherapeutics
have formed a corner stone of efforts to reduce the burden of
malaria, but have been hampered by the continuous emergence
Recent studies conducted by our group, focusing on indole based
antimalarials,[21,22] identified a new class of compounds, two
members of which (1 and 2, Figure 1) possessed good
1
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antimalarial activity in an in vitro phenotypic screening campaign
against the chloroquine sensitive 3D7 strain, with no observable
mammalian cytotoxicity against a control HeLa cell line.
to their contribution to hydrophobic interactions, as well as
halogen bond formation.[23] Moreover, their varying atomic radii
allow for subtle interrogation of substituent size limitations on
biological activity. We therefore expanded our ‘halogenated set’
through the incorporation of 5-fluoro indole analogues. The
introduction of supposedly chemically inert methyl moieties, does
on occasion reduce the free energy required to desolvate the
ligand in question, thereby energetically favouring binding in a
hydrophobic pocket, in what is referred to as the ‘magic methyl
effect’.[24] Furthermore, melatonin features a C-5 methoxylated
indole, which has been implicated as a critical factor in malaria
biology, and exploited as an endogenous ligand for antimalarial
drug design.[25–27] Consequently, we opted to synthesise
analogues featuring a C-5 methyl or methoxy indole. In addition
to this strategic structural modification, a C-2 methyl indole was
incorporated to explore potential effects of this substitution.
Figure 1. Promising early antimalarial compounds identified through phenotypic
screening against the 3D7 P. falciparum strain
Accordingly, we rationalised a cohort of analogues (3 – 22) of
compounds 1 and 2, which were designed to answer specific
questions pertaining to our structure activity relationship (SAR).
This study provided new insight into the SAR and pharmacophore
of this class of compounds, which will be discussed in detail below,
and led to the discovery of two new compounds (13 and 14),
which displayed potent activity against the 3D7 strain,
comparable to that of chloroquine, in the absence of notable HeLa
cytotoxicity or haemolysis. Interestingly, biological assessment of
our most active compounds, against chloroquine sensitive NF54
resulted in a reduction in biological activity into the mid-nanomolar
range, which was maintained against the multidrug resistant K1
strain.
The electronegativity of halogens renders them net electron-
withdrawing groups, which can potentially alter the electronic
environment of the indole ring. We therefore sought to incorporate
a bioisosteric nitrile moiety,[28,29] which has also demonstrated the
capacity to displace bound waters for a net entropic gain.[30]
Region C analogues had focussed mainly on thiophenols, which
had indicated firstly, that para-substituted analogues were
preferred for activity, with the strongly electron withdrawing nitro
group providing the greatest antimalarial activity. Electron
donating groups, particularly amines negatively influenced activity,
and introduced significant mammalian cytotoxicity.
Results and Discussion
SAR strategy
SAR analysis from our initial study had provided suitable insight
into positional substituents required for optimal activity (Figure 2).
In this study, we sought to resolve greater detail of specific
requirements at each position through strategic bioisosteric
replacement of specific moieties, as well as combinations of
promising substituents in order to enhance antimalarial potency
without compromising mammalian toxicity.
Region A analogues had indicated a preference for C-5
substitution, however, our C-5 substituted analogues were limited
to a chlorine or bromine derivative, providing limited information
pertaining to what properties of the substituents was responsible
for enhanced activity. We were particularly interested in whether
the activity was possibly due to substituent size, enhanced
lipophilicity, or the electron withdrawing nature of the substituent.
Halogens are common substituents in medicinal chemistry, due
Figure 2. SAR analysis from our previous study identified three regions of
interest for further investigation. Region A: C-5 substituent at R1 increased
antimalarial activity, while C-6, and C-7 reduces activity. The physicochemical
properties of the R1 substituent which contribute toward activity is unresolved.
Region B: Influence of R2 substitution on activity unexplored. Region C: X =
NH increases toxicity. Unclear whether an O or S in this region is superior for
activity. Extension of chain between X and the phenyl ring not tolerated. Para-
substitution on ring essential. Electron donating groups at R3 decrease activity
and introduce toxicity. Strong electron withdrawing groups increase activity.
2
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In addition, we had determined that extension of the chain
between the sulfur and the aromatic ring was not well tolerated
from an antimalarial and cytotoxicity perspective as well as ruling
out amides and α-amino carbonyls as viable linker moieties
between the indole and phenyl rings. However, it remained
unresolved whether an α-oxo or thio carbonyl, was a preferable
linker group. Accordingly, we generated a small cohort of oxygen
and sulfur containing compounds featuring an electron
withdrawing nitro group or related bioisosteres.
(Scheme 5). The remaining indoles, phenols and thiophenols
were all commercially available. Finally, we applied an m-CPBA
oxidation method[34] to derive a sulfone analogue (22) of an
antimalarial analogue (29), previously reported by us[21] (Scheme
6).
Synthesis and Biological Evaluation
Our general synthetic pathway closely followed that of our original
study,[21,22] whereby appropriately substituted indole underwent a
Friedel-Crafts acetylation, followed by selective bromination to
yield the requisite α-bromo ketone intermediates. These were
then subjected to nucleophilic displacement with a suitable phenol
or thiophenol in the presence of potassium carbonate, to generate
our final compounds at yields varying from poor to
excellent(Scheme 1). Thiophenols featuring a strongly electron
withdrawing substituent, have demonstrated the capacity to
induce reductive dehalogenation of α-halogenated ketones.[31,32]
However, this phenomenon was not observed to any significant
extent in this study.
Compounds
3 – 22 were initially subjected to a single
5-Cyano indole (23) was prepared in good yield from 5-iodo indole
utilising a CuI/PPh3 catalytic system with CuCN as a nitrile source
(Scheme 2), while 3-acetyl-5-methoxyindole (24) was
synthesised from 5-bromo-3-acetyl indole via an Ullman type
nucleophilic displacement (Scheme 3).
concentration screen against both the 3D7 and HeLa lines at 20
µM (Figure 3). This screen identified several compounds which
inhibited P. falciparum cell viability to below 15%. Compounds 6,
11, 17 and 22 were unable to reduce 3D7 cell viability below 50%
and were not subjected to further assessment. Compound 17
seemingly enhanced malarial proliferation, indicating possible
assay interference. Importantly, none of the compounds inhibited
HeLa cell viability to below 77% indicating that these compounds
display a high degree of selectivity. In order to refine our SAR
understanding, compounds which inhibited 3D7 viability to below
50% were submitted for IC50 determination (Table 1).
Tetrazole 25 was generated via a 1,3 dipolar cycloaddition of 4-
cyanophenol as per the method of Koyama et al.[33] (Scheme 4).
4-Mercaptomethylbenzoate (26) was synthesised from 4-
mercaptobenzoic acid via the acid chloride, while a similar method
was used to generate the corresponding primary amide (27),
which was dehydrated into the desired 4-cyanothiophenol
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Figure 3. Cell viability (%) of 3D7 and HeLa cell lines at 20 µM. Vales are the mean ± SD of experiments performed in duplicate. Compounds
which reduced cell viability below the 50% threshold were submitted for IC50 determination. In general, low mammalian cytotoxicity was observed.
Table 1. IC50 values (Pf 3D7) of compounds 1 – 22
Compounds 3 – 10 were designed as oxygen containing
IC50
IC50
(µM)
No.
Structure
No.
Structure
(µM)[a]
analogues of compound 1 and 2. The negligible loss of biological
activity observed through compounds 3 and 4 suggested that both
an oxygen and nitrile moiety were relatively well tolerated.
However, activity was not maintained with an electron
withdrawing tetrazole (5). Furthermore, the incorporation of a
pyridine (6) resulted in a significant reduction in activity. Activity
was somewhat restored, upon incorporation of a hydrophobic 5-
methyl moiety (7), and to a lesser extent with the 2-methyl (8),
while compound 9, which featured a strongly electron withdrawing
nitrile moiety at the indole C-5 position, possessed no observable
activity. Finally, biological activity was significantly improved
through the combination of a para cyanophenol and a 5-
chloroindole (10). We proceeded to prepare a series of sulfur
containing analogues (11 – 22). As a direct comparison of 1 and
3.5 ±
0.7
1
0.20[a]
12
0.027 ±
0.0
2
0.090[b]
13
14
15
16
17
18
19
20
21
22
0.33 ±
0.1
0.044 ±
0.0
3
0.41 ±
0.0
1.1 ±
0.2
4
8.3 ±
0.3
0.098 ±
0.0
5
6
ND[c]
ND
4, compound 11 resulted in
a moderate yet noticeable
improvement in biological activity, while replacement of the nitrile
with a methyl ester (12) dramatically reduced activity. Compounds
13 and 14, which combined design elements of compounds 1, 2
and 11 increased biological activity into the low nanomolar range,
with activity comparable to that of chloroquine.
0.87 ±
0.1
0.25 ±
0.0
7
2.2 ±
0.2
1.4 ±
0.3
8
Activity was reduced upon replacement of the C-5 chlorine with
fluorine (15 and 16), while the nitrile analogue of compound 2 and
15 (17) was inactive. Compounds 18 – 20 were designed as sulfur
analogues of 7, 8 and 9 respectively. Compounds 18 and 20 in
particular, showed a vast improving in biological activity over their
corresponding oxygen analogues. Interestingly, incorporation of a
C-5 methoxy substituent (21) showed a moderate but discernible
improvement in biological activity in comparison to 18.
0.54 ±
0.0
9
ND
0.1 ±
0.0
0.13 ±
0.0
10
11
0.13 ±
0.0
ND
0.03 ±
0.0
Chloroquine
[a] Values are the mean ± SD of experiments biological repeats performed in
technical duplicate. [b] Values determined in previous study. [c] ND = Not
determined
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The final compound tested in this series was a sulfone derivative
(22) of a low micromolar inhibitor from our original study, was
found to be inactive. With respect to our study questions, these
data provided several new elements to our SAR hypothesis. The
differences in activity between compounds 7 – 10 and 13, 18 – 20
indicates that the sulfur moiety is seemingly superior to an oxygen
in terms of 3D7 inhibition, however, oxidation of the sulfur, which
is a possible metabolic pathway of this series reduces activity,
with a negligible effect on cytotoxicity. Strongly electron-
withdrawing groups are beneficial for biological activity, but the
relative loss of activity observed with the tetrazole and ester
moieties suggests that this effect is not ubiquitous to all electron-
withdrawing groups, and a size limitation may exist at the putative
binding site. C-2 methylation was seemingly detrimental to activity
overall, while hydrophobicity is seemingly more important than
election withdrawing capacity with respect to C-5 substitution.
Furthermore, the moderate reduction in activity observed
between compounds 14 and 16 in addition to the moderate gain
observed between 18 and 21, point toward an optimal substituent
size in this region, which is currently best exploited with a chlorine
analogue. However, this may still be as a result of a halogen
bonding effect. Due to their encouraging activity against 3D7,
compounds 13 and 14, were investigated for possible haemolysis,
where they were found to be non-lytic, at concentrations far
exceeding the HC50 of haemolytic peptide gramicidin D (Figure
4).
compounds 11, 14 and 21 all retained reduced albeit encouraging
mid nanomolar activity. Furthermore, this strain was seemingly
more susceptible to chloroquine than 3D7. Compound 14 was the
most active of our compounds against the K1 stain, in the mid
nanomolar range, while notable improvements in susceptibility
were seen with compounds 13 and 20. The general consistency
and /or improvement in compound activity between the NF54 and
K1 suggests that these compounds do not inhibit a pathway which
is compromised by K1 resistance, as shown by a >2 fold
difference between the isolates. However, the question as to the
major changes in biological activity between the isogenic
chloroquine sensitive strains is more perplexing.
Table 2. IC50 (µM) of compounds 10, 11, 13,
14,18 and 21 against Pf NF54 and K1.[a]
no.
NF54
K1
10
0.77 ± 0.0
0.5 ± 0.0
>1
0.9 ± 0.1
0.45 ± 0.1
0.57 ± 0.1
0.36 ± 0.0
0.66 ± 0.0
0.48 ± 0.1
11
13
14
0.28 ± 0.0
0.9 ± 0.0
0.65 ± 0.0
0.0034 ± 0.0
18
21
Chloroquine
Artensunate
0.0027 ± 0.0
[a] Values are the mean ± SEM of experiments
performed in triplicate
It must be noted that these assays were conducted independently
of one another in separate laboratories, where subtle inter-lab
variability may account for variations in the biological data.
Furthermore, recent studies have demonstrated that phenotypic
and genotypic characteristics of P. falciparum isolates are
unstable in long term culture, which alter the susceptibility of
different P falciparum strains to different drugs.[35,36] To illustrate,
cultures of the 3D7 strain of P. falciparum have been found to
express significantly higher levels of the adhesion protein
encoding var genes than NF54[37] which is particularly active in
the ring stage.[38] While this specific alteration may not adequately
answer our specific alterations in biological activity, it does
highlight the challenges in antimalarial drug discovery, and the
need for robust multifaceted datasets to adequately assess
promising compounds.
Figure 4. Haemolysis assay of compounds 13 and 14 vs gramicidin D, which
indicates low levels of haemolysis at concentrations in excess of the gramicidin
D HC50. Values are the mean ± SD of experiments biological repeats performed
in technical duplicate
Compounds 10, 11, 13, 14, 18 and 21 were selected for biological
evaluation against the isogenic NF54 strain as well as the
multidrug resistant K1 strain (Table 2). In this screen, all
compounds, suffered from a dramatic reduction in biological
activity against NF54, particularly 10 and 13, 18. The remaining
Conclusions
In conclusion, this study sought to expand the SAR of a promising
class of non-toxic antimalarial compounds, through specific
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under reflux, following which the reaction mixture was allowed to cool
before filtering. The crude organics were re dissolved in EtOAc and
washed with 10% NaOH (30 ml). The mixture was dried over MgSO4,
concentrated in vacuo and purified by flash column chromatography
(DCM: EtOAc, 4:1) to yield a brown solid (20%): IR (cm-1) 3150, 2934,
1613, 1518, 1427, 804, 652, 587; 1H NMR (CD3OD, 300 MHz): δH 8.08
(1H, s, H-2), 7.77 (1H, d, J = 2.4 Hz, H-4), 7.32 (1H, d, J = 8.9 Hz, H-7),
6.86 (1H, dd, J = 8.9, 2.4 Hz, H-6), 3.83 (3H, s, H-9), 2.50 (3H, s, H-2’);
13C NMR (CD3OD, 75 MHz): δC 195.0, 156.2, 134.3, 131.9, 126.2, 116.9,
112.9, 112.1, 103.1, 54.6, 25.6 ppm.
compound modifications. Through this strategy, we have refined
our SAR hypothesis against
a currently unknown target.
Furthermore, compounds 13 and 14 were identified as potent
non-haemolytic inhibitors of P. falciparum 3D7 with negligible
cytotoxicity towards a HeLa cell line. However, this activity
reduced significantly when scrutinised against two further P.
falciparum strains. Whilst the reasons for this remain unclear at
this stage, compound 14 maintained encouraging activity against
NF54 as well as K1, indicating that it inhibits a biological pathway
independent of those compromised by resistance in K1, and it
remains a promising compound in malaria drug discovery. Further
studies are planned to elucidate the mechanism of action of this
class as well as to fully elucidate the pharmacophore.
The synthesis and characterisation of several 3-acetylindoles and
corresponding α-bromoketones has previously been reported by us.[41,42]
Below are compounds used in this study, which are yet to be reported by
us.
1-(5-Methyl-1H-indol-3-yl)ethanone: (19%), brown solid; IR (film) νmax
cm-1 3132, 2918, 1621, 1431, 1374, 943, 798, 757; 1H NMR (DMSO-d6,
300 MHz): δH 11.77 (1H, brs, NH-1), 8.22 (1H, s, H-2), 7.98 – 7.96 (1H, m,
H-4), 7.34 (1H, d, J = 8.3 Hz, H-7), 7.02 (1H, dd, J = 8.3, 1.6 Hz, H-6), 2.42
(3H, s, H-2’), 2.39 (3H, s, H-8); 13C NMR (DMSO-d6, 75 MHz): δC 193.0,
135.3, 134.6, 130.7, 126.0, 124.5, 121.4, 117.0, 112.1, 27.6, 21.7 ppm.
Experimental Section
Chemistry
General methods: NMR spectra were obtained using either a Bruker
Fourier 300 MHz, 400 MHz or 600 MHz Avance II spectrometer. Chemical
shifts are reported in ppm with residual reference solvents resonances as
follows: Acetone-d6 δH 2.05, δC 29.8, 206.2; CD3OD δH 3.31, δC 49.0;
DMSO-d6 δH 2.50, δC 39.5 ppm.[39] High resolution mass spectrometry was
performed on a Waters Synapt G2 TOF instrument with an ESI source. IR
spectra were obtained using a Perkin Elmer Spectrum 2000 FT-IR. Melting
points were determined using a SMP30 Advanced Digital Melting Point
apparatus. Flash column chromatography was performed using Kieselgel
60 (230–400 mesh) silica gel. All bulk solvents were distilled prior to use,
while anhydrous solvents were prepared by standard procedures outlined
by Perrin and Armarego[40]. Compounds 13 and 14 were determined to
have a measured purity ≥ 95% by LC-UV/vis-MS.
1-(2-Methyl-1H-indol-3-yl)ethanone: (43%), brown solid; IR (film) νmax
1
cm-1 3162, 3050, 2970, 1609, 1528, 1444, 1413, 1390, 969, 905, 739; H
NMR (DMSO-d6, 400 MHz): δH 11.81 (1H, brs, NH-1), 8.02 – 8.00 (1H, m,
H-4), 7.37 – 7.35 (1H, m, H-7), 7.14 – 7.11 (2H, m, H-5, H-6), 2.67 (3H, s,
H-8), 2.51 (3H, s, H-2’); 13C NMR (DMSO-d6, 100 MHz): δC 193.5, 144.6,
135.1, 127.3, 122.2, 121.7, 121.0, 113.9, 111.6, 31.3, 15.4 ppm.
1-(5-Cyano-1H-indol-3-yl)ethanone: (46%), cream solid; IR (film) νmax
cm-1 3109, 2220, 1739, 1628, 1615, 1444, 1376, 805, 655; 1H NMR
(DMSO-d6, 300 MHz): δH 12.40 (1H, brs, NH-1), 8.52 (1H, dd, J = 1.6, 0.8
Hz, H-4), 8.51 (1H, s, H-2), 7.65 (1H, dd, J = 8.5, 0.8 Hz, H-7), 7.58 (1H,
dd, J = 8.5, 1.6 Hz, H-6), 2.48 (3H, s, H-2’); 13C NMR (DMSO-d6, 75 MHz):
δC 192.9, 138.5, 136.6, 126.3, 125.7, 125.0, 120.2, 116.9, 113.7, 103.9,
27.3 ppm.
5-Cyanoindole (23): CuI (80 mg, 0.41 mmol) was suspended in
anhydrous N,N-DMF (20 ml) in a 2-neck round bottom flask under a
nitrogenous atmosphere. To this, 5-iodoindole (500 mg, 2.05 mmol), PPh3
(107 mg, 0.41 mmol) and CuCN (552 mg, 6.17 mmol) were added, and
allowed to stir for 24 h at 140 °C, after which time the reaction mixture was
allowed to cool. Organic material was dissolved in EtOAc, and washed
with saturated brine solution (3 x 10 ml). The extract was dried over
anhydrous MgSO4 before concentrating in vacuo. Compound 23 was
purified via flash column chromatography (Hexane: EtOAc, 7:3) to yield a
brown amorphous solid (77%): IR (film) νmax cm-1 3400, 2224, 1610, 1414,
1-(5-Fluoro-1H-indol-3-yl)ethanone: (50%), yellow solid; 1H NMR
(DMSO-d6, 300 MHz): δH 11.96 (1H, s, NH-1); 8.36 (1H, s, H-2); 7.86 
7.80 (1H, dd, J = 10.0, 2.6 Hz, H-4); 7.50  7.45 (1H, m, H-7); 7.10  7.02
(1H, m, H-6); 2.44 (s, 3H, H-2’) ppm; 13C NMR (DMSO-d6, 75 MHz): δC
192.7,160.6 (d, JF.C = 236 Hz), 135.9, 133.2, 125.8 (d, JF.C = 11.2 Hz),
116.8 (d, JF.C = 4.4 Hz); 113.4 (d, JF.C = 10.5 Hz), 110.9 (d, JF.C = 25.6 Hz),
106.0 (JF.C = 22.6 Hz); 27.1 ppm.
1
769, 732; H NMR (DMSO-d6, 300 MHz): δH 11.65 (1H, brs, NH-1), 8.08
2-Bromo-1-(5-methyl-1H-indol-3-yl)ethanone: (19%), yellow solid; IR
(film) νmax cm-1 3185, 2919, 1643, 1514, 1430, 1365, 801, 761, 672; 1H
NMR (CH3OD, 300 MHz): δH 8.21 (1H, s, H-2), 8.03 (1H, s, H-4), 7.34 (1H,
d, J = 8.3 Hz, H-7), 7.09 (1H, d, J = 8.3 Hz, H-6), 4.47 (2H, s, H-2’), 2.45
(3H, s, H-8); 13C NMR (CH3OD, 75 MHz): δC 188.0, 135.4, 134.6, 131.9,
126.0, 124.8, 121.2, 113.6, 111.3, 31.1, 20.4 ppm.
(1H, dd, J = 1.6, 0.7 Hz, H-4), 7.57 – 7.54 (2H, m, H-2, H-7), 7.41 (1H, dd,
J = 8.4, 1.6 Hz, H-6), 6.58 (1H, dd, J = 3.2, 0.7 Hz, H-3); 13C NMR (DMSO-
d6, 75 MHz): δC 137.6, 128.1, 127.4, 125.7, 123.7, 120.8, 112.7, 102.1,
ppm.
1-(5-Methoxy-1H-indol-3-yl)ethanone (24): Sodium methoxide (sodium
362 mg, methanol, 10 ml) was added to a stirring mixture of CuI (484 mg,
2.5 mmol), 1-(5-bromo-1H-indol-3-yl)ethanone (250 mg, 1.1 mmol) in
anhydrous N,N-DMF (15 ml). The reaction was allowed to proceed for 6 h
2-Bromo-1-(2-methyl-1H-indol-3-yl)ethanone (44%), brown solid; IR
(film) νmax cm-1 3302, 3114, 2946, 1715, 1624, 1454, 1411, 980, 890, 757,
589; 1H NMR (CD3OD, 300 MHz): δH 7.97 – 7.94 (1H, m, H-4), 7.38 – 7.34
6
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ChemMedChem
FULL PAPER
(1H, m, H-7), 7.21 – 7.17 (2H, m, H-5, H-6), 4.51 (2H, s, H-2’), 2.74 (3H, s,
H-8); 13C NMR (CD3OD, 75 MHz): δC 188.1, 146.6, 135.3, 126.6, 122.2,
121.8, 120.5, 110.9, 34.4, 14.0 ppm.
4-Mercaptobenzonitrile (28): To
a
stirred solution of 4-
mercaptobenzamide (±3.55 mmol, 1 eq.) in DMF was added POCl3 (8.88
mmol, 2.5 eq.) at 0 ᵒC. The mixture was allowed to react for 2 h, following
which the reaction mixture was dissolved in EtOAc (20 ml) and washed
with sat. brine (3 x 10 ml). In vacuo solvent removal afforded a yellow waxy
solid which required no further purification (93%0 1H NMR (DMSO-d6,
3-(2-Bromoacetyl)-1H-indole-5-carbonitrile: Inseparable mixture with 1-
(5-Cyano-1H-indol-3-yl)ethanone and was taken forward into the next step
without further purification
300MHz): δH 7.88  7.84 (2H, m, H-3); 7.73  7.69 (2H, m, H-2) ppm; 13
NMR (DMSO-d6, 75 MHz): δC 141.5, 133.3, 126.6, 118.4, 109.9 ppm.
C
2-Bromo-1-(5-methoxy-1H-indol-3-yl)ethanone (37%), cream solid; IR
(film) νmax cm-1 3186, 2928, 1639, 1514, 1426, 1029, 951, 920, 886, 743,
670; 1H NMR (Acetone-d6, 300 MHz): δH 11.07 (1H, brs, NH-1), 8.34 (1H,
d, J = 3.3 Hz, H-2), 7.82 (1H, d, J = 2.6 Hz, H-4), 7.43 (1H, d, J = 8.9 Hz,
H-7), 6.89 (1H, dd, J = 8.9, 2.6 Hz, H-6), 4.51 (2H, s, H-2’), 3.84 (3H, s, H-
9); 13C NMR (Acetone-d6, 75 MHz): δC 186.1, 156.4, 134.0, 131.8, 126.9,
114.1, 113.5, 112.8, 103.2, 54.9, 32.2 ppm.
General synthetic procedure for compounds (3 – 22): A relevant
phenol or thiophenol (2 eq.) was added to a stirring suspension of a
corresponding 2- bromo-1-(1H-indol-3-yl)ethanone ( 1 eq.) and K2CO3 (2
eq.) in acetone. The reaction mixture was heated to reflux for 5 h, following
which time the reaction mixture was cooled and dissolved in EtOAc. The
organic phase was washed with water (3 x 15 ml) and sat. brine (10 ml).
Solvent was removed in vacuo and the resultant crude product was
purified by normal phase flash chromatography (Hexane: EtOAc 4:1) to
yield a crystalline solid. The solid was re-dissolved in acetone, which was
allowed to crystallise through slow evaporation. These resultant crystals
were washed with cold chloroform to yield the desired product.
2-Bromo-1-(5-fluoro-1H-indol-3-yl)ethanone (34%), cream solid; 1H
NMR (DMSO-d6, 300 MHz): δH 12.27 (1H, s, NH-1); 8.54 (1H, d, J = 3.31
Hz, H-2); 7.81 (1H, dd, J = 9.9, 2.5 Hz H-4); 7.54  7.49 (1H, m, H-7); 7.14
 7.07 (1H, m, H-6); 4.65 (2H, s, H-2”); 13C NMR (DMSO-d6, 75- MHz) : δC
186.9, 157.4 (d, JF.C = 243 Hz), 136.8, 133.3, 126.2 (d, JF.C = 11.8 Hz),
115.5, 113.6 (d, JF.C = 10.5 Hz); 111.5 (d, JF.C = 25.7 Hz) 106. (d, JF.C
=
26.9 Hz) 33.5 ppm
1-(1H-indol-3-yl)-2-(4-nitrophenoxy)ethanone (3): White crystalline
solid, m.p: 220 – 222 C, (66 %); IR (film) νmax cm-1 3172, 2922, 1635, 1581,
1505, 1428, 1340, 1235, 1111; 1H NMR (DMSO-d6, 600 MHz): δH 8.52
(1H, s, H-2 ); 8.24  8.18 (2H, m, H-6’); 8.14  8.11 (1H, m, H-4); 7.53 
7.50 (1H, m, H-7); 7.27  7.14 (2H, m, H-5, H-6, H-5’); 5.55 (2H, s, H-2’).
13C NMR (DMSO-d6, 150 MHz) ): δC 188.3, 163.8, 141.0, 136.5, 134.3,
125.9, 125.4, 123.3, 122.2, 121.2, 115.3, 113.1, 112.4, 70.3 ppm;
HRESMS m/z 297.0871 (calcd for C16H13N2O4 [M+H]+ 297.0875).
4-(1H-tetrazol-5-yl)phenol (25): NaN3 (272 mg, 4.2 mmol) and
ammonium chloride (224 mg, 4.2 mmol) were added to a stirring mixture
of 4-cyanophenol (500 mg, 4.2 mmol) in anhydrous N,N-DMF (2 ml) under
a nitrogenous atmosphere. The reaction proceeded for 7 h at 125 °C,
following which time it was allowed to cool. 1M NaOH (10 ml) was added
to the mixture and stirred for a further 30 minutes, followed by adjustment
of the pH to 1 through the addition of small portions of 32 % HCl solution.
The organic material was extracted with EtOAc (3 x 15 ml) prior to drying
over MgSO4. Solvent was removed in vacuo, and the crude organic sold
was further dried at 80°C for 24 h at low pressure. Compound 25 was
purified using flash column chromatography (Hexane: EtOAc, 1:1)
followed by crystallisation from EtOAc and washing with CHCl3 to yield a
white crystalline solid (31%): IR (film) νmax cm-1 3414, 2843, 2620, 1614,
1599, 1415, 1279, 1249, 1079, 830, 751; 1H NMR (CD3OD, 400 MHz): δH
1-(1H-indol-3-yl)-2-(4-cyanophenoxy)ethanone (4): White crystalline
solid, m.p: 237 – 239 C, (76 %); νmax cm-1 3171, 2221, 1643, 1605, 1506,
1429, 1300, 1156, 938. 790; 1H NMR (DMSO-d6, 400 MHz): δH 12.12 (1H,
s, NH-1); 8.51 (1H, s, H-2); 8.14  8.12 (1H, m, H-4); 7.79 – 7.75 (2H, m,
H-6’); 7.52 – 7.49 (1H, m, H-7); 7.26 – 7.18 (2H, m, H-5, H-6); 7.15 – 7.11
(2H, m, H-5’); 5.47 (s, 2H, H-2’);13C NMR (DMSO-d6, 100 MHz) ): δC 188.5,
161.9, 136.4, 134.2 (2 x CH), 125.3, 123.2, 122.1, 121.2, 119.2, 115.7,
113.2, 112.3, 102.9, 69.9 ppm; HRESMS m/z 277.0968 (calcd for
C17H13N2O2 [M+H]+ 277.0977).
7.84 (2H, d, J = 8.7 Hz, H-2’, H-6’), 6.95 (2H, d, J = 8.7 Hz, H-3’, H-5’); 13
NMR (CD3OD, 100 MHz): δC 160.6, 155.4, 128.6, 115.8, 114.4 ppm.
C
Methyl 4-mercaptobenzoate (26): 4-Mercaptobenzoic acid (500 mg, 3.24
mmol) was dissolved in thionyl chloride (3ml) under a nitrogenous
atmosphere and heated to 70 ºC for 2 h. Following cooling, sodium
methoxide solution was added dropwise, allowing the fumes to purge. The
reaction mixture was dissolved in EtOAc (45 ml) and washed with sat.
brine (3 x 10 ml). Solvent was removed in vacuo affording a beige waxy
solid, which required no further purification (43%): 1H NMR (DMSO-d6,
300MHz): δH 7.96  7.93 (2H, m, H-3); 7.67  7.64 (2H, m, H-2); 3.82 (3H,
s, H-7) ppm; 13C NMR (DMSO-d6, 75 MHz): δC 165.7, 141.3, 130.3, 128.5
126.5, 52.4 ppm.
1-(1H-indol-3-yl)-2-(4-[1H-tetrazol-5-yl]phenoxy)ethanone (5): White
needles, m.p: 225 – 227 °C (20%); IR (film) νmax cm-1 3473, 3307, 2957,
2737, 1640, 1617, 1595, 1523, 1442, 1379, 1244, 1141, 836, 759; 1H NMR
(Acetone-d6, 600 MHz): δH 8.57 (1H, s, H-2), 8.26 – 8.24 (1H, m, H-4), 8.02
– 7.99 (2H, m, H-6’), 7.59 – 7.57 (1H, m, H-7), 7.31 – 7.23 (2H, m, H-5, H-
6), 7.01 – 6.99 (2H, m, H-5’), 6.27 (2H, s, H-2’); 13C NMR (Acetone-d6, 100
MHz): δC 185.3, 165.8, 160.2, 137.8, 134.7, 129.0, 126.6, 124.5, 123.3,
122.5, 120.1 , 116.7, 115.0, 113.0, 58.9, ppm; HRESMS m/z 320.1142
(calcd for C17H14N5O2 [M+H]+ 320.1147).
1-(1H-indol-3-yl)-2-(5-nitropyridin-2-yloxy)ethanone
(6):
White
The same method was used to generate 4-mercaptobenzamide 27, which
crystalline solid, m.p: 258 – 260 C, (13%); IR (film) νmax cm-1 3403, 3126,
1602, 1458, 1418, 1280, 1167, 1108.6, 1027, 837, 745; 1H NMR (DMSO-
was taken forward without full characterisation.
7
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10.1002/cmdc.201800235
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FULL PAPER
d6, 300 MHz): δH 12.19 (s, 1H, NH-1); 9.22 (1H, d, J = 3.1 Hz, H-6’); 8.61
(1H, s, H-2); 8.22 (1H, dd, J = 10.0, 3.1 Hz, H-8’); 8.11 – 8.08 (1H, m, H-
4); 7.54 – 7.51 (1H, m, H-7); 7.28 – 7.19 (2H, m, H-5, H-6); 6.55 (1H, d, J
= 10.0 Hz, H-9’); 5.52 (2H, s, H-2’); 13C NMR (DMSO-d6, 75 MHz) ): δC
186.2, 160.9, 143.1, 136.6, 134.5, 133.9, 129.8, 125.2, 123.2, 122.2, 121.0,
118.1, 113.5, 112.5, 55.2 ppm; HRESMS m/z 298.0824 (calcd for
C15H12N3O4 [M+H]+ 298.0828).
(Acetone-d6, 150 MHz) ): δC 187.8, 144.8, 136.8, 133.7, 132.0, 126.8,
125.8, 123.2, 122.0, 121.7, 118.4, 115.4, 111.8, 107.8, 38.9 ppm;
HRESMS m/z 293.0739 (calcd for C17H13N2OS [M+H]+ 293.0749).
1-(1H-indol-3-yl)-2-[(4-methylbenzoate)thio]ethanone (12): White
crystalline solid, m.p: 99 – 101 C, (26%); IR (film) νmax cm-1 3261, 2920,
2850, 1713, 1628, 1518, 1432, 1274, 1274, 1109; 1H NMR (DMSO-d6,
600 MHz): δH 12.11 (1H, s, NH-1); 8.53 (1H, d, J = 3.2 Hz, H-2); 8.13 (1H,
d, J = 8.1 Hz, H-4); 7.85  7.82 (2H,m, Hz, H-6’); 7.50  7.46 (3H, m, H-5’,
H-7); 7.24  7.18 (2H, m, H-5, H-6); 4.61 (2H, s, H-2’); 3.81 (3H, s, H-10’)
ppm;13C NMR (DMSO-d6, 150 MHz): δC 188.4, 165.9, 144.0, 136.6, 135.1,
129.5, 126.3, 125.9, 125.4, 123.1, 122.1, 121.2, 114.9, 112.3, 52.0, 39.2
ppm; HRESMS m/z 326.0842 (calcd for C18H16NO3S [M+H]+ 326.0851).
1-(5-Methyl-1H-indol-3-yl)-2-(4-nitrophenoxy)ethanone (7): Orange
crystalline solid, m.p: 235 – 237 °C, (82%); IR (film) νmax cm-1 3253, 2920,
2854, 1709, 1650, 1591, 1507, 1422, 1339, 1251, 1234, 1110, 820, 750,
689; 1H NMR (DMSO-d6, 400 MHz): δH 12.02 (1H, brs, NH-1), 8.44 (1H, s,
H-2), 8.21 – 8.19 (2H, m, H-6’), 7.94 (1H, s, H-4), 7.39 (1H, d, J = 8.3 Hz,
H-7), 7.18 – 7.15 (2H, m, H-5’), 7.07 (1H, d, J = 8.3 Hz, H-6), 5.52 (2H, s,
H-2’), 2.39 (3H, s, H-8); 13C NMR (DMSO-d6, 100 MHz): δC 188.6, 164.2,
141.4, 135.2, 134.5, 131.4, 126.2, 126.1, 125.1, 121.3, 115.7, 113.2, 112.4,
70.7, 21.7 ppm; HRESMS m/z 311.1025 (calcd for C17H15N2O4 [M+H]+
311.1032).
1-(5-Chloro-1H-indol-3-yl)-2-[(4-cyanophenyl)thio]ethanone
(13):
White crystalline solid, m.p: 249 – 251 C, (36 %); IR (film) νmax cm-1 3203,
2907, 2219, 1634, 1592, 1434, 1229, 1142, 1088, 952, 894; 1H NMR
(DMSO-d6, 300 MHz): δH 12.33 (1H, s, NH-1); 8.63 (1H, d, J = 2.7 Hz, H-
2); 8.09 (1H, d, J = 2.1 Hz, H-4); 7.74 – 7.71 (2H, m, H-6’); 7.55 – 7.49 (3H,
1-(2-Methyl-1H-indol-3-yl)-2-(4-nitrophenoxy)ethanone (8): Yellow
crystalline solid, m.p: 217 – 219 °C, (98%); IR (film) νmax cm-1 3274, 2923,
1637, 1590, 1508, 1427, 1333, 1250, 1171, 974, 838, 740; 1H NMR
(Acetone-d6, 600 MHz): δH 11.08 (1H, brs, NH-1), 8.22 – 8.20 (2H, m, H-
6’), 8.06 – 8.04 (1H, m, H-4), 7.45 – 7.43 (1H, m, H-7), 7.21 – 7.19 (2H, m,
H-5, H-6), 7.19 – 7.17 (2H, m, H-5’, H-9’), 5.56 (2H, s, H-2’), 2.81 (3H, s,
H-8); 13C NMR (Acetone-d6, 150 MHz): δC 188.3, 164.2, 145.0, 141.4,
135.3, 126.7, 125.5, 122.3, 121.9, 120.9, 115.1, 111.4, 111.3, 72.3, 14.6
ppm; HRESMS m/z 311.1026 (calcd for C17H15N2O4 [M+H]+ 311.1032).
m, H-5’, H-7); 7.26 (1H, dd, J = 8.6, 2.1 Hz, H-6); 4.66 (2H, s, H-2’); 13
C
NMR (DMSO-d6, 75 MHz) ): δC 188.5, 144.5, 136.5, 135.2, 132.4, 126.9,
126.7, 126.6, 123.3, 120.4, 119.0, 114.5, 114.1, 107.1, 39.8 ppm;
HRESMS m/z 327.0353 (calcd for C17H1235ClN2OS [M+H]+ 327.0359).
1-(5-Chloro-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(14):
Yellow crystalline solid, m.p: 249 – 251 °C (46%); IR (film) νmax cm-1 3170,
3121, 2922, 1713, 1637, 1577, 1507, 1424, 1329, 1087, 843, 739, 680; 1H
NMR (Acetone-d6, 600 MHz): δH 11.37 (1H, brs, NH-1), 8.59 (1H, d, J =
2.4 Hz, H-2), 8.28 (1H, d, J = 2.1 Hz, H-4), 8.16 – 8.13 (2H, m, H-5’), 7.64
– 7.61 (2H, m, H-6’), 7.57 (1H, d, J = 8.7 Hz, H-7), 7.27 (1H, dd, J = 8.7,
2.1 Hz, H-6), 4.69 (2H, s, H-2’); 13C NMR (Acetone-d6, 150 MHz): δC 188.7,
148.3, 145.9, 136.0, 135.8, 128.5, 127.9, 127.3, 124.5, 124.3, 121.9, 115.9,
114.5, 39.9 ppm; HRESMS m/z 347.0255 (calcd for C16H1235ClN2O3S
[M+H]+ 347.0257).
1-(5-Cyano-1H-indol-3-yl)-2-(4-nitrophenoxy)ethanone (9): Cream
crystalline solid, m.p: 254 – 256 °C, (17% over two steps); IR (film) νmax
cm-1 3180, 3116, 2924, 2216, 1729, 1657, 1505, 1430, 1344, 1261, 1239,
945, 779; 1H NMR (DMSO-d6, 400 MHz): δH 8.63 (1H, s, H-2), 8.46 (1H, d,
J = 1.4 Hz, H-4), 8.20 – 8.16 (2H, m, H-6’), 7.70 (1H, d, J = 8.5 Hz, H-7),
7.61 (1H, dd, J = 8.5, 1.4 Hz, H-6), 7.18 – 7.14 (2H, m, H-5’), 5.54 (2H, s,
H-2’); 13C NMR (DMSO-d6, 100 MHz): δC 189.1, 164.0, 141.5, 138.7, 136.9,
126.7, 126.6, 126.3, 125.5, 120.5, 115.7, 114.4, 113.7, 104.8, 70.8 ppm;
HRESMS m/z 320.0676 (calcd for C17H10N3O4 [M-H]- 320.0671).
1-(5-Fluoro-1H-indol-3-yl)-2-[(4-bromophenyl)thio]ethanone
(15):
White crystalline solid, m.p: 177 – 179 C, (63%); IR (film) νmax cm-1 3197,
2921, 1714, 1612, 1467, 1430, 1168, 810, 706; 1H NMR (DMSO-d6-, 600
MHz): δH 12.19 (1H, s, NH-1); 8.56 (1H, s, H-2); 7.79 (1H, dd, J= 9.8, 2.7
Hz, H-4); 7.51  7.46 (3H, m, H-7, H-5’); 7.34  7.32 (2H, m, H-6’); 7.10 
7.07 (1H, m, H-6); 4.48 (2H, s, C-2’) ppm; 13C NMR (DMSO-d6, 150 MHz):
δC 188.8; 158.7 (d, JF.C = 235 Hz); 136.4, 136.0, 133.2, 131.6, 129.8, 126.0
(d, JF.C = 11.1 Hz), 118.5, 114.9 (d, JF.C = 4.3 Hz), 113.5 (d, JF.C = 9.7 Hz),
111.2 (d, JF.C = 25.8 Hz), 106.0 (d, JF.C = 24.3 Hz), 39.8 (CH2, C-2’) ppm;
HRESMS m/z 363.9803 (calcd for C16H1279BrFNOS [M+H]+ 363.9807).
1-(5-Chloro-1H-indol-3-yl)-2-(4-cyanophenoxy)ethanone (10): White
crystalline solid; m.p: 229 – 231 C, (36 %); IR (film) νmax cm-1 3177, 2917,
1
2224, 1659, 1509, 1250, 1153, 946, 807; H NMR (DMSO-d6, 400 MHz):
δH 12.32 (1H, s, NH-1); 8.57 (1H, s, H-2); 8.10 (1H, d, J = 2.0 Hz, H-4);
7.78 – 7.75 (2H, m, H-6’); 7.54 (1H, d, J = 8.7 Hz, H-7); 7.26 (1H, dd, J =
8.7, 2.0 Hz, H-6); 7.15 – 7.12 (2H, m, H-5’); 5.47 (2H, s, H-2’); 13C NMR
(DMSO-d6, 100 MHz) ): δC 188.7, 161.8, 135.5, 134 .9, 134.2, 126.9, 126.5,
123.3, 120.3, 119.2, 115.8, 114.1, 112.8, 103.1, 69.9 (CH2, C-2’) ppm;
HRESMS m/z 311.0591 (calcd for C17H1235ClN2O2 [M+H]+ 311.0587).
1-(5-Fluoro-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone (16): White
crystalline solid, m.p: 210 – 212 C, (55%); IR (film) νmax cm-1 3171, 2917,
2850, 1727, 1613, 1505, 1174, 840, 737; 1H NMR (DMSO-d6, 600MHz):
δH 12.25 (1H, s, NH-1); 8.65 (1H, d, J = 3.3 Hz, H-2); 8.14  8.12 (2H, m,
H-6’); 7.79 (1H, dd, J = 9.8, 2.5 Hz, H-4); 7.59  7.57 (2H, m, H-5’); 7.53 
1-(1H-indol-3-yl)-2-[(4-cyanophenyl)thio]ethanone
(11):
Brown
crystalline solid, m.p: 204 – 206 C, (22%); IR (film) νmax cm-1 3211, 2220,
1627, 1402, 1239, 1137, 1088, 947, 813; 1H NMR (Acetone-d6, 600 MHz):
δH 11.25 (1H, s, NH-1); 8.49 (1H, d, J = 3.1 Hz, H-2); 8.29 – 8.28 (1H, m,
H-4); 7.67  7.65 (2H, m, H-5’); 7.58 – 7.56 (2H, m, H-6’); 7.54 – 7.53 (1H,
m, H-7); 7.27 – 7.22 (2H, m, H-5, H-6); 4.61 (2H, s, H-2’); 13C NMR
7.51 (1H, m, H-7); 7.12  7.08 (1H, m, H-6); 4.73 (2H, s, H- 2’) ppm; 13
NMR (DMSO-d6, 150 MHz): δC 187.8, 158.7 (d, JF.C = 236 Hz), 147.4, 144.4,
C
8
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10.1002/cmdc.201800235
ChemMedChem
FULL PAPER
136.6, 133.2, 126.4, 125.9 (d, JF.C = 11.4 Hz), 125.9, 123.8, 114.8 (d, JF.C
= 4.4 Hz), 113.6 (d, JFC = 9.9 Hz), 111.3 (d, JF.C = 26.2 Hz), 106.0 (d, JF.C
= 24.1 Hz), 38.7 ppm; HRESMS m/z 331.0545 (calcd for C16H12FN2O3S
[M+H]+ 331.0553).
113.5, 113.4, 103.3, 55.7, 39.1 (CH2, C-2’) ppm; HRESMS m/z 343.0744
(calcd for C17H15N2O4S [M+H]+ 343.0753).
1-(1H-indol-3-yl)-2-[(4-bromophenyl)sulfonyl]ethanone
(22):
A
solution of m-CPBA (2.5 eq.) in DCM (3 ml) was added dropwise to a
solution of 1-(1H-indol-3-yl)-2-(4-bromophenyl)ethanone (100 mg, mmol)
in THF (3 ml). The reaction mixture was allowed to stir for 3 hours at room
temperature, followed by the addition of a saturated sodium bicarbonate
solution, and extracted with EtOAc (3 x 15 ml). Solvent was removed in
vacuo and crude organics were purified by normal phase flash
chromatography (Hexane: EtOAc 1:1) to yield a white solid (75%), 1H NMR
(Acetone-d6, 600 MHz): δH 11.29 (1H, s, NH-1); 8.42 (1H, d, J = 3.1 Hz, H-
2); 8.21 (1H, d, J = 7.7 Hz, H-4); 7.87  7.82 (4H, m, H-5’, H-6’); 7.53 (1H,
1-(5-Cyano-1H-indol-3-yl)-2-[(4-bromophenyl)thio]ethanone
(17):
Cream crystalline solid, m.p: 242 – 244 °C, (6% over two steps); IR (film)
νmax cm-1 3112, 2930, 2220, 1784, 1614, 1523, 1440, 1374, 1085, 805, 784;
1H NMR (DMSO-d6, 400 MHz): δH 12.56 (1H, brs, NH-1), 8.67 (1H, s, H-
2), 8.48 (1H, d, J = 1.5 Hz, H-4), 7.68 (1H, d, J = 8.4 Hz, H-7), 7.61 (1H,
dd, J = 8.4, 1.5 Hz, H-6), 7.49 – 7.46 (2H, m, H-6’), 7.35 – 7.31 (2H, m, H-
5’), 4.52 (2H, s, H-2’); 13C NMR (DMSO-d6, 100 MHz): δC 189.2, 138.5,
137.2, 135.8, 131.7, 129.9, 126.2, 126.0, 125.2, 120.0, 118.7, 115.1, 113.9,
104.3, 39.6* ppm, HRESMS m/z 370.9849 (calcd for C17H1279BrN2OS
[M+H]- 370.9854. *resolved by HSQC.
d, J = 7.8 Hz H-7); 7.29 – 7.22 (2H, m, H-5, H-6); 4.87 (2H, s, H-2’); 13
C
NMR (Acetone-d6, 150 MHz) ): δC 181.5, 139.5, 137.1, 136.1, 132.3, 130.5,
128.3, 125.9, 123.7, 122.5, 121.8, 117.4, 112.2, 64.2 ppm; HRESMS m/z
377.9790 (calcd for C16H1379BrNO3S [M+H]+ 377.9800).
1-(5-Methyl-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(18):
Yellow crystalline solid, m.p: 218 – 221 °C (62%); IR (film) νmax cm-1 3159,
3112, 2914, 1703, 1611, 1502, 1431, 1338, 1088, 960, 800, 739; 1H NMR
(DMSO-d6, 400 MHz): δH 12.02 (1H, brs, NH-1), 8.52 (1H, m, H-2), 8.14 –
8.11 (2H, m, H-6’), 7.95 (1H, s, H-4), 7.60 – 7.56 (2H, m, H-5’), 7.37 (1H,
m, H-7), 7.05 (1H, m, H-6), 4.70 (2H, s, H-2’), 2.39 (3H, s, H-8); 13C NMR
(DMSO-d6, 100 MHz): δC 188.3, 148.1, 144.9, 135.6, 135.4, 131.4, 126.8,
126.1, 125.0, 124.2, 121.4, 114.9, 112.4, 39.2, 21.8 ppm; HRESMS m/z
327.0798 (calcd for C17H15N2O3S [M+H]+ 327.0803).
Biological assessment
Antiplasmodial assay (3D7): P. falciparum (3D7 strain) parasites were
maintained at 37oC under an atmosphere of 5% CO2, 5% O2, 90% N2 in
RPMI 1640 medium containing 25 mM HEPES, 22 mM glucose, 0.65 mM
hypoxanthine, 0.5% (w/v) Albumax II, 0.05 mg/ml gentamicin and 2-4%
haematocrit human erythrocytes. For compound assessment, parasite
cultures were adjusted to 2% parasitaemia and 1% haematocrit,
distributed in 96 well plates and compound stocks of 20 mM in dimethyl
sulfoxide (DMSO) added to a final concentration of 20 µM. Incubation was
continued for 48 hours, after which parasite levels in the culture wells were
determined using a parasite lactate dehydrogenase (pLDH) assay.[43]
Briefly, 20 µl of culture was removed from each well and transferred to a
separate plate containing 125 µl/well pLDH reagent (44 mM Tris, pH 9,
1-(2-Methyl-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(19):
Orange crystalline solid, m.p: 215 – 217 °C (69%); IR (film) νmax cm-1 3288,
1
2906, 1732, 1610, 1500, 1455, 1416, 1332, 980, 832, 738, 715; H NMR
(DMSO-d6, 400 MHz): δH 12.05 (1H, brs, NH-1), 8.15 – 8.12 (2H, m, H-6’),
8.06 – 8.02 (1H, m, H-4), 7.55 – 7.52 (2H, m, H-5’), 7.41 – 7.39 (1H, m, H-
7), 7.19 – 7.16 (2H, m, H-5, H-6), 4.80 (2H, s, H-2’), 2.76 (3H, s, H-8); 13
C
NMR (DMSO-d6, 100 MHz): δC 187.7, 147.8, 145.6, 144.3, 134.8, 126.7,
126.5, 123.8, 122.2, 121.8, 120.8, 111.9, 111.4, 42.4, 15.3 ppm; HRESMS
m/z 327.0801 (calcd for C17H15N2O3S [M+H]+ 327.0803).
containing 0.18
M L-lactic acid, 0.13 mM acetylpyridine adenine
dinucleotide, 0.39 mM nitrotetrazolium blue chloride, 0.048 mM
phenazineethosulphate and 0.16% (v/v) Triton X-100). Colour
development was monitored as absorbance at 620 nm and converted to %
parasite viability relative to untreated control wells. Dose-response assays
were conducted as described above, except that parasite cultures were
incubated in 96 well plates with 3-fold serial dilutions of test compounds.
Plots of % parasite viability vs. log[compound] were used to derive IC50
values by non-linear regression analysis using GraphPad Prism.
1-(5-Cyano-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(20):
Yellow crystalline solid, m.p: 226 – 230 °C, (10% over two steps); IR (film)
νmax cm-1 3182, 2920, 2219, 1637, 1580, 1513, 1434, 1335, 1080, 843, 742;
1H NMR (Acetone-d6, 300 MHz): δH 11.65 (1H, brs, NH-1), 8.71 (1H, s, H-
2), 8.64 (1H, d, J = 1.4 Hz, H-4), 8.17 – 8.13 (2H, m, H-6’), 7.76 (1H, d, J
= 8.6 Hz, H-7), 7.65 – 7.61 (2H, m, H-5’), 7.59 (1H, dd, J = 8.6, 1.4 Hz, H-
6), 4.72 (2H, s, H-2’); 13C NMR (DMSO-d6, 100 MHz): δC 188.8, 147.7,
145.0, 138.9, 137.9, 126.9, 126.6, 126.5, 125.0, 124.3, 120.4, 115.5, 114.4,
104.8, 39.3* ppm. HRESMS m/z 336.0454 (calcd for C17H10N3O3S [M-H]-
336.0443) *resolved by HSQC.
Antiplasmodial assay (NF54 and K1): Both parasite strains were
cultured continuously in normal type
A human erythrocytes (2%
haematocrit) using a method modified from Trager and Jensen.[44] Parasite
cultures were maintained in complete tissue culture medium of RPMI 1640
supplemented with 25 mM HEPES buffer, 20 μg/ml gentamicin, 27 mM
sodium hydrogen carbonate and 0.5% Albumax II serum supplement,
1-(5-Methoxy-1H-indol-3-yl)-2-[(4-nitrophenyl)thio]ethanone
(21):
Yellow crystalline solid, m.p: 218 – 223 °C (87%); IR (film) νmax cm-1 3165,
3122, 2928, 1735, 1610, 1501, 1427, 1337, 1275, 842, 798, 739; 1H NMR
(DMSO-d6, 400 MHz): δH 12.05 (1H, brs, NH-1), 8.53 (1H, d, J = 3.3 Hz,
H-2), 8.14 – 8.12 (2H, m, H-6’), 7.63 (1H, d, J = 2.4 Hz, H-4), 7.59 – 7.57
(2H, m, H-5’), 7.39 (1H, d, J = 8.8 Hz, H-7), 6.86 (1H, dd, J = 8.8, 2.4 Hz,
H-6), 4.70 (2H, s, H-2’), 3.76 (3H, s, H-9); 13C NMR (DMSO-d6, 100 MHz):
δC 188.3, 156.2, 148.1, 144.9, 135.7, 131.8, 126.8, 126.7, 124.2, 115.1,
which was incubated at 37 ̊C in an atmosphere of 3% O2, 4% CO2 and
91% N2. In vitro antiplasmodial activity was again determined using a
pLDH assay.[43] Starting from a concentration of 1000 ng/ml, test
compounds were serially diluted 2-fold to 2 ng/ml in complete medium.
Each dilution was distributed in a 96-well microtiter plate with parasite
suspension in triplicate and incubated at 37 °C for 72 hours. Cultures in
each well were re-suspended carefully and aliquots were removed and
9
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10.1002/cmdc.201800235
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FULL PAPER
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values were obtained by the non-linear dose-response curve fitting
analysis using GraphPad Prism v.4.0 for windows.
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Entry for the Table of Contents
Strategic fine tuning: We report the optimisation of a novel antimalarial chemotype, resulting in two non-haemolytic compounds
which are equipotent to chloroquine against the 3D7 strain with negligible cytotoxicity. Further studies against the isogenic NF54
strain, resulted in a reduction in activity into the mid nanomolar range. However, similar activity against both the NF54 and K1 strains
suggest that these compounds inhibits a pathway which is uncompromised by K1 resistance.
12
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