Discovery of novel and potent benzhydryl-tropane trypanocides highly selective for Trypanosoma cruzi

Article abstract of DOI:10.1016/j.bmcl.2010.02.022

A benzhydryl tropinone oxime that is potently toxic to Trypanosoma cruzi has been previously identified. An SAR investigation determined that no part of the original compound was superfluous and all early SAR probes led to significant drops in activity. The only alteration that could be achieved without loss of activity was replacement of the aryl chloride substituent with chloro homologues. This led to the discovery of a trifluoromethyl-containing analogue with an EC₅₀ against T. cruzi of 30 nM and a cytotoxicity selectivity index of over 1000 relative to rat skeletal myoblast L-6 cells.

Full text of DOI:10.1016/j.bmcl.2010.02.022

Bioorganic & Medicinal Chemistry Letters 20 (2010) 1816–1818
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of novel andpotent benzhydryl-tropane trypanocides highly selective
for Trypanosoma cruzi
G. A. Holloway a, J. P. Parisot a, P. M. Novello a, K. G. Watson a, T. Armstrong b, R. C. A. Thompson b, I. P. Street a,
J. B. Baell a,
*
a Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
b Murdoch University, Murdoch, Western Australia, Australia
a r t i c l e i n f o
a b s t r a c t
Article history:
A benzhydryl tropinone oxime that is potently toxic to Trypanosoma cruzi has been previously identified.
An SAR investigation determined that no part of the original compound was superfluous and all early SAR
probes led to significant drops in activity. The only alteration that could be achieved without loss of activ-
ity was replacement of the aryl chloride substituent with chloro homologues. This led to the discovery of
a trifluoromethyl-containing analogue with an EC50 against T. cruzi of 30 nM and a cytotoxicity selectivity
index of over 1000 relative to rat skeletal myoblast L-6 cells.
Received 16 December 2009
Revised 3 February 2010
Accepted 3 February 2010
Available online 8 February 2010
Keywords:
Trypanosoma
Chagas disease
Ó 2010 Elsevier Ltd. All rights reserved.
The protozoan parasite Trypanosoma cruzi is the causative agent
of Chagas disease which is found in 18 countries in Latin America.
In the 2004 World Health Report1 Chagas disease was estimated to
cause 14,000 deaths and a disease burden of 0.7 million DALYs
annually. There are currently only two significant therapies for
Chagas disease, nifurtimox and benznidazole (Fig. 1).2 These drug
treatments are ineffective at preventing the development and
treatment of chronic Chagas disease3 and induce a number of ad-
verse effects. Hence, there is an urgent need for the development
of new cost effective therapy against T. cruzi with minimal side
effects.
was found to be potently toxic towards T. cruzi with an EC50 of
110 nM and highly selective for this organism relative to other try-
panosomatids such as Leishmania donovani and Trypanosoma bru-
cei.5 These data are summarised in Figure 2.
For several reasons, significantly amongst these being the very
weak trypanothione reductase inhibitory activity of compound 1
(EC50 35 lM, Fig. 2), we have previously argued that the observed
potent T. cruzi-selective trypanocidal activity could likely be due to
We have previously reported on a high-throughput screening
campaign conducted to identify inhibitors of trypanothione reduc-
tase (TR) that might consequently be toxic to trypanosomatids.4,5
Amongst the identified hits was a benzhydryl tropinone oxime that
Figure 1. Current T. cruzi therapies.
* Corresponding author.
E-mail address: jbaell@wehi.edu.au (J.B. Baell).
Figure 2. Structure and antiparasitic activity of benzhydryl tropinone oxime 1.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.02.022

G. A. Holloway et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1816–1818
1817
Figure 3. Benzhydryl tropane oxime synthesis. See Supplementary data.
off-target activity.5 Subsequently, we gained access to a higher
throughput parasite assay and it was principally for this reason
that we initiated a hit optimisation campaign around compound
1, using a whole parasite primary assay. This communication will
disclose the results of our SAR investigation into this compound.
Two main synthetic methods were utilised (Fig. 3). One method
enabled rapid analogue production of the tropane portion of 1 by
reaction of a biaryl oxylamine with an appropriate ketone. The
biaryl oxylamine was synthesised via deprotection of the corre-
sponding benzhydryloxyphthalimide6 which in turn was synthes-
ised from the biaryl alcohol, either directly through a Mitsunobu
reaction6 or via its corresponding chloride.7,8 The second method
enabled rapid analogue production of the benzhydryl portion,
through reaction of the tropinone oxime with an appropriate aryl-
methyl chloride. Results from the whole cell T. cruzi assay of the
first set of compounds (Fig. 4) confirmed that two aryl groups were
essential for activity and removal of one of the phenyl rings (2 and
3) resulted in loss of activity. It was also determined that the basic
nitrogen atom in the tropane ring was important, as replacement
with cycloalkyl (4) or alkyl (5) resulted in complete loss of activity.
Even removal of the N-methyl group (6) or the tropane bridge (7)
led to significant loss of activity.
Figure 5. SAR probing of the benzhydryl group.
Having established that the tropane ring and two aryl groups
are important, a second set of compounds was assembled to inves-
tigate changes in aryl ring connectivity. Replacing the unsubstitut-
Figure 6. Benzhydryl and tropane substituent SAR.
ed phenyl ring with a pyridine or thiazole led to a significant loss in
activity (8–13, Fig. 5), though to greatly varying extents, as the 4-
pyridyl analogue 10 was 12 times weaker again than the 3-pyridyl
analogue 9. Similarly, introducing a carbon chain between the two
phenyl rings on either side led to variable losses in activity (14–
18), and constraining the rings (19) led to complete loss of activity.
This tight SAR suggested that the benzhydryl group should be re-
tained for optimum activity. Therefore, by way of finer SAR probing,
a series of compounds with electron withdrawing and donating
groups in different positions on the benzhydryl phenyl rings were
synthesised (20–28, Fig. 6). These compounds all lost significant
Figure 4. Benzhydryl tropane oxime SAR: deletion analogues.

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G. A. Holloway et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1816–1818
In summary, we have described a series of potent trypanocides,
benzhydryl tropinone oximes, which do not tolerate variation to
the structure apart from substitution of the chloro-aryl substituent
by other hydrophobic electronegative substituents. Compound 34,
where the chlorine atom is replaced by a trifluoromethyl group,
has an EC50 against T. cruzi of 30 nM with cytotoxicity to mamma-
lian cells more than 1000-fold weaker.
Acknowledgments
Financial support was received from the drugs for neglected
diseases initiative (DNDi) and the Victorian State Government OISS
Grant and NHRMC IRIISS Grant #361646.
Figure 7. Chloro homologue SAR.
Supplementary data
activity, except the p-methoxy compound (24) which retained rea-
sonable activity with an EC50 of 300 nM. We similarly investigated
fine alterations of the tropane ring, namely by extension of the N-al-
kyl substituent. As shown for compounds 27 and 28, significant loss
of activity was observed for the tropane N-hydroxyethyl and N-
(dimethylamino)ethyl substituents, respectively.
Thus far, no improvement in the activity of the initial hit 1 had
been obtained, suggesting that all structural aspects of this com-
pound were important for biological activity. With this informa-
tion in hand, a final set of very fine SAR probes was assembled,
where the chlorine atom was replaced with homologues. These re-
sults are shown in Figure 7. Gratifyingly, good activity was consis-
tently observed and in three cases was better than the original hit.
Thus the bromo (30), iodo (31) and trifluoromethyl compounds
(34) displayed outstanding activity with whole organism EC50 val-
ues of 40, 40 and 30 nM, respectively. The weaker activity for the
fluoro, methyl and cyano-containing analogues suggests a combi-
nation of electronegativity but principally hydrophobicity is
important for most potent activity.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.02.022.
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