University of Dundee
Anti-trypanosomatid drug discovery
Field, Mark C.; Horn, David; Fairlamb, Alan H.; Ferguson, Michael A. J.; Gray, David W.;
Read, Kevin D.
Published in:
Nature Reviews Microbiology
DOI:
10.1038/nrmicro.2016.193
Publication date:
2017
Document Version
Peer reviewed version
Link to publication in Discovery Research Portal
Citation for published version (APA):
Field, M. C., Horn, D., Fairlamb, A. H., Ferguson, M. A. J., Gray, D. W., Read, K. D., De Rycker, M., Torrie, L. S.,
Wyatt, P. G., Wyllie, S., & Gilbert, I. H. (2017). Anti-trypanosomatid drug discovery: an ongoing challenge and a
continuing need. Nature Reviews Microbiology, 15(4), 217-231. https://doi.org/10.1038/nrmicro.2016.193
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Download date: 26. Aug. 2022
1
Vector-borne diseases series
Antitrypanosomatid drug discovery: an ongoing challenge and a continuing need
Mark C. Field, David Horn, Alan H. Fairlamb, Michael A. J. Ferguson, David W. Gray, Kevin D. Read,
Manu De Rycker, Leah S. Torrie, Paul G. Wyatt, Susan Wyllie and Ian H. Gilbert
Division of Biological Chemistry and Drug Discovery, University of Dundee, Dundee, DD1 5EH, UK
Correspondence to I.H.G. i.h.gilbert@dundee.ac.uk
Abstract | The World Health Organization recognizes human African trypanosomiasis, Chagas’
disease and the leishmaniases as neglected tropical diseases. These diseases are caused by parasitic
trypanosomatids and range in severity from mild and self-curing to near invariably fatal. Public
health advances have substantially decreased the impact of these diseases in recent decades, but
alone will not eliminate these diseases. Here we discuss why new drugs against trypanosomatids
are needed, approaches that are under investigation to develop new drugs and why the drug
discovery pipeline remains essentially unfilled. Additionally, we consider the important challenges
to drug discovery strategies and the new technologies that can address them. The combination of
new drugs, new technologies and public health initiatives are essential for the management and
hopefully eventual elimination of trypanosomatid diseases from the human population.
Trypanosomatid parasites cause several neglected diseases of humans and animals,
which range in severity from comparatively mild to near invariably fatal
1,2
. The organisms
responsible for human diseases are: Trypanosoma brucei spp., which cause human African
trypanosomiasis (HAT); T. cruzi, which causes Chagas’ disease; and Leishmania spp., which cause
the leishmaniases. Together, these insect-transmitted parasites threaten millions of people. All of
these organisms have complex life-cycles, with substantial differences in morphology, cell biology
and biochemistry between lifecycle stages and in some cases between species (Box 1).
Control of trypanosomatid diseases has had a mixed history, although public health campaigns
are showing success in many instances. For example, the Southern Cone and Andean initiatives are
tackling Chagas’ disease with a combination of insecticide spraying of dwellings, improved housing,
screening of people in endemic zones and blood bank monitoring
3
. However, South America has
considerable numbers of T. cruzi-infected individuals and many infected individuals have migrated
to North America and Europe, where the disease is non-endemic. In the case of leishmaniasis, co-
infection with Leishmania spp. and HIV can increase the disease burden and severity, and recent
refugee movements from the Middle East into Europe are likely to increase the prevalence of
leishmaniasis in Europe. In the immediate post-colonial period, HAT resurged, but vector control,
active case-finding and treatment have all helped control the disease
4
. However, many
2
trypanosomatid diseases are zoonotic, which will make eradication extremely unlikely. The current
target is elimination, which is still an ambitious goal. Despite progress, trypanosomatid diseases
remain a substantial public health problem and there is an urgent need for new drugs to tackle
them.
None of the available drugs for treatment of trypanosomatid disease (Table 1) are satisfactory
and new drugs are needed, especially those suitable for rural health systems with limited resources.
The current standard of care is monotherapy, with the exception of nifurtimox-eflornithine
combination therapy (NECT) for HAT, although various drug combinations are in clinical trials.
Importantly, many of the current treatments require parenteral administration
5
, and also suffer
from poor efficacy, major side effects and increasing levels of resistance
6-8
. Most of the drugs in use
probably have multiple modes of action, due to acting on multiple parasite targets
9
. Goals for drug
discovery include the development of completely new classes of therapeutics, reduced host toxicity,
improved administration regimens and the development of combination therapies.
Vaccine development is a powerful approach to disease management but remains challenging
in the trypanosomatid diseases due to efficient immune evasion mechanisms, such as antigenic
variation in the African trypanosomes, and the intracellular locations of T. cruzi and Leishmania spp.
in the human host. Progress towards human
10
and canine
11
leishmania vaccines and the challenges
in developing vaccines for HAT
12
and Chagas’ disease
13
have been reviewed recently and will not be
discussed further here.
In this Review, we discuss the potential for the development of new drug therapies against
trypanosomatids. We highlight unique biological features of these parasites that suggest potential
targets, methods used to identify bioactive compounds and consider some of the outcomes of
recent campaigns. We encourage the reader to consider excellent reviews of life cycles, genomes,
pathogenesis and more general aspects of the biology of trypanosomatids published elsewhere
14-
19
.
[H1] Drug discovery
A successful drug discovery campaign typically takes 10-15 years (Fig. 1). High attrition rates,
together with relatively few organizations working on drug discovery for trypanosomatid parasites
mean that the number of new compounds in clinical development is very low (Fig. 2) and unlikely
to meet the clinical need. Ideally, the pipeline would contain multiple new agents that are suitable
for combination therapy. The advantages of combination therapies are manifold: they can increase
the clinical efficacy of treatments; they can reduce side effects by allowing lower dosing of individual
agents; and they can reduce the risk of resistance development. Reducing resistance is critical for
safeguarding whatever new medicines do emerge from the drug discovery pipeline.
Three broad approaches are used for drug discovery against trypanosomatids: (1) target-
based approaches involve screening for inhibitors against a purified protein, for example an enzyme.
Compounds identified through the screening (or structure-based) process are subsequently
optimized to show efficacy in a cellular model; (2) phenotypic approaches involve screening for
growth-inhibitors directly against an intact parasite, usually in an in vitro culture; (3) compound re-
3
positioning is re-deployment of compounds previously developed for an alternative indication as
anti-trypanosomatid therapies.
The drug discovery process is ideally driven by target product profiles (TPPs), which define the
properties required of a drug for clinical application
20-22
. Such factors include: route of
administration (oral, inhaled, intravenous, etc.), acceptable dosing regimen and course of
treatment, acceptable safety and tolerability levels, cost and shelf-life. TPPs allow for the
development of compound progression criteria, which define parameters for compounds at each
stage in the drug discovery process (hit, validated hit, lead, preclinical candidate, etc. –see Fig. 1).
Progression criteria include assessment of the physicochemical properties (such as solubility in
physiological media, lipophilicity, molecular weight, hydrogen bond donors and acceptors), potency
(against the molecular target and intact organism), selectivity, chemical and metabolic stability,
pharmacokinetics, efficacy and safety. Additional criteria for parasitic infections can include factors
such as cytocidal activity and the rate of parasite killing. The Drugs for Neglected Diseases Initiative
(DNDi) is a public private partnership that focuses on drug discovery and clinical development for
these organisms. It has developed TPPs and compound progression criteria for trypanosomatid
diseases (www.dndi.org)
21
.
[H1] Target-based approaches
For target-based approaches, the key is careful selection of the most promising molecular
targets. A recent review highlights some examples of target-based drug discovery against
trypanosomatids
23
. For neglected diseases in general, including the trypanosomatid diseases, there
has been very limited success from target-based approaches. This is often due to lack of translation
from inhibition of the target (enzyme) in a purified cell-free context to inhibition of proliferation of
the parasite and/or subsequent activity in an animal disease model. In part, this reflects the absence
of robustly validated targets (for example, enzymes whose activity is essential to the parasite) and
highlights the need for fundamental research into trypanosomatid biology and for thorough genetic
and chemical validation of potential targets
24
. However, this is only part of the problem. As will be
discussed below, an improved understanding of how to translate compounds that are active in vitro
into therapeutics is required, which includes better defining the cellular and animal models (Box 2)
that predict clinical efficacy in humans.
We have published some criteria to help in selection of molecular targets (Box 3)
9,20,24
. Many
target-based drug discovery programmes can be initially viewed as target-validation
25
. It is therefore
vital to obtain proof-of-concept (POC) of anti-parasitic activity for new target-derived chemical
series at the earliest possible stage, ideally both in cellular and animal models, to minimize waste of
resources, should the target fail to progress.
[H3] Drug targets with the highest degree of validation
The best validated drug-target in T. brucei is ornithine decarboxylase (ODC), which is the target
of eflornithine, a drug used clinically for treatment of HAT. Eflornithine is a suicide inhibitor that was
initially developed for the treatment of cancer, but subsequently re-purposed for HAT
26
. Selectivity
is thought to arise from the more rapid turnover of human ODC compared to the trypanosome
4
enzyme
27
, or due to inhibition of trypanothione biosynthesis
28
, which is a metabolite unique to
trypanosomatids.
The enzyme N-myristoyltransferase (NMT) has also been well validated as a molecular target
for HAT
29-31
. In a programme initiated with a high-throughput screen against NMT, a compound
series was identified and subsequently optimized (typified by DDD85646, Fig. 3b) to be active in a
mouse model of the first stage of HAT, which does not involve the central nervous system. There
was strong evidence that the compounds inhibit NMT in cells and that this inhibition kills parasites,
which validates both the target and the mode of action. NMT is also present in humans, but T. brucei
is acutely sensitive to NMT inhibition, probably because endocytosis, which occurs at a very high
rate in T. brucei, is affected. NMT has also been validated as a target in a second stage mouse model
of HAT (K.D.R., personal communication). The challenge with second stage disease is that
compounds need to penetrate the blood brain barrier and achieve therapeutic concentrations in
the central nervous system without causing host toxicity
Very recently the proteasome has been shown to have great potential as a target in all three
types of trypansomatids
32
. This study used a phenotypic approach to develop a parasite-specific,
selective inhibitor (GNF6702) that does not inhibit the human proteasome This is an excellent
example of taking a phenotypic hit and subsequently deconvoluting the target. The initial
experiments to determine the mode of action involved generating compound-resistant T. cruzi
mutants, followed by whole genome sequencing, which revealed mutations in the β4 subunit of the
proteasome. Various additional biochemical experiments demonstrated that GNF6703 specifically
inhibits the chymotrypsin-like activity of the parasite proteasome.
[H3] Biological features of trypanosomatids that might be targeted
Trypanosomatids are one of the most evolutionary divergent eukaryotic lineages from
mammals, a feature that is reflected in their distinct biology (Fig. 3a). Conversely, there are many
similarities between T. brucei, T. cruzi and Leishmania spp. and many molecular mechanisms are
conserved between all three lineages. Trypanosomatid-specific metabolic and cellular pathways
(discussed below) should represent excellent drug targets as specificity should be an easier criterion
to control, but no candidate drugs have been developed that inhibit such targets. In fact, most
potential trypanosome-specific targets remain unexplored for drug discovery and/or are of
unknown druggability. Ironically, the best validated targets in trypanosomatids are one repurposed
from oncology (ODC) and two pan-eukaryotic essential targets (NMT and the proteasome),
discussed above.
Uniquely, trypanosomatids package the first six or seven enzymes of glycolysis into the
glycosome, a specialized form of the peroxisome. Glycolysis is especially important for the
bloodstream forms of African trypanosomes, which rely exclusively on this pathway for ATP
production. The compartmentalization of glycolysis in trypanosomatids is accompanied by
fundamental differences in allosteric regulation of the pathway compared to most other
eukaryotes. Because of this, phosphofructokinase, for example, is being pursued as a target
33
.
However, computational modelling of glycolysis suggests that there is little prospect of killing
trypanosomes by suppressing glycolysis unless inhibition is irreversible or uncompetitive, due to the