The search for novel antimalarial compounds has shifted in recent years from target- 878
based approaches towards whole cell ‘phenotypic’ screening. Whole cell screening 879
involves assessing diverse compound libraries for the ability of compounds to inhibit 880
parasite growth (Guiguemde et al., 2012). An advantage of this approach over target- 881
based approaches is that the discovery of antimalarial compounds does not require 882
prior knowledge of the biological target. This strategy has led to the identification of 883
numerous antimalarial compounds with diverse chemical structures (Plouffe et al., 884
2008, Gamo et al., 2010, Guiguemde et al., 2010, Duffy and Avery, 2013, Avery et al., 885
2014, Plouffe et al., 2016). 886
887
Identifying the molecular target(s) of antimalarial compounds identified in phenotypic 888
screens is not necessarily straightforward. While it is not necessary to have knowledge 889
of a compound’s chemical target prior to that compound entering the clinical pipeline, 890
knowledge of the compound target enables the development of specific biochemical 891
assays for compound/target interaction that aid in medicinal chemistry efforts to 892
improve the pharmacodynamic profile of a compound (Flannery et al., 2013). For a 893
number of compounds identified in whole cell screens as having activity against the 894
asexual stage parasite, the exposure of malaria parasites to either static or increasing 895
concentrations of the compound, typically over a period of weeks to months, has given 896
rise to resistant parasites (Rottmann et al., 2010, Jimenez-Diaz et al., 2014, Lehane et 897
al., 2014, Vaidya et al., 2014, Flannery et al., 2015, Hameed et al., 2015, Golldack et al., 898
2017, Hapuarachchi et al., 2017). Genomic techniques have been used to identify 899
mutations in the resistant parasites, thereby providing insight into mechanisms of 900
resistance and, potentially, the identity of the target. 901
902
Confirming the target of a particular antimalarial compound entails a demonstration of 903
inhibition of the activity of a particular pathway and/or protein, using one or more 904
appropriate biochemical assays. One of the challenges with regard to the screening of 905
the large number of antimalarial compounds that have emerged from whole cell 906
phenotypic screens is the adaptation of biochemical assays to a high-throughput 907
format (Flannery et al., 2013). It is with this in mind that 400 structurally diverse 908
antimalarial compounds, representing some of the most promising compounds 909
identified in phenotypic screens, were curated by the Medicine’s for Malaria Venture 910
into a set of compounds called the ‘Malaria Box’ (Spangenberg et al., 2013). The 911
Malaria Box was distributed free of charge and this has enabled researchers across the 912
world to discover the targets for some of these compounds (Lehane et al., 2014, 913
Ahyong et al., 2016, Dickerman et al., 2016, Van Voorhis et al., 2016, Golldack et al., 914
2017, Hapuarachchi et al., 2017). 915
916
The high-throughput phenotypic screening of chemical libraries against the asexual 917
blood stage malaria parasite has led to the discovery of a number of compounds with 918
mechanisms of action that are different from currently administered antimalarial drugs 919
(Rottmann et al., 2010, Jimenez-Diaz et al., 2014, Hameed et al., 2015, Golldack et al., 920
2017, Hapuarachchi et al., 2017), some of which are promising clinical candidates 921
(Jimenez-Diaz et al., 2014, Hameed et al., 2015). In terms of generating clinical 922
candidates, the asexual stage parasite high-throughput screens have been successful, 923
however, it is possible that the entirety of currently accessible compound space has 924
been explored (Guiguemde et al., 2010, Guiguemde et al., 2012, Diagana, 2015) and it 925
is likely that many of the potent drug-like compounds have their growth inhibitory 926
affect by accessing one of a limited number of targets (Diagana, 2015). These include 927
the transporter PfATP4, the lipid kinase phosphatidylinositol-4-OH kinase, the 928
pyrimidine biosynthesis enzyme dihydroorotate dehydrogenase, the respiratory chain 929
complex cytochrome bc1 and the folate recycling enzyme dihydrofolate reductase 930
(Diagana, 2015). 931
932
One drawback of the high-throughput screening systems, both target-based and whole 933
cell, is that molecules that need to be biologically activated in vivo will not be detected 934
in an in vitro screen (Flannery et al., 2013). 935
936
Ideally a compound will target multiple stages of the malaria parasite lifecycle, and so, 937
whilst clearing an asexual stage infection, also block transmission and/or remove the 938
reservoir of parasites in the liver (Leroy et al., 2014, Smith et al., 2014). High- 939
throughput screens of gametocyte-stage killing activity have uncovered compounds 940
with transmission-blocking potential (Duffy and Avery, 2013, Plouffe et al., 2016). 941
Furthermore, the development of a method to screen for compounds that kill the liver 942
stage parasite has led to the discovery of a class of compounds - the 943
imidazolopiperazines - with potent activity against liver and blood-stage parasites 944
(Meister et al., 2011). Lead optimisation of the imidazolopiperazine compound led to 945
the development of the clinical candidate KAF156 (Nagle et al., 2012) which has shown 946
promising activity against P. falciparum and P. vivax in early stage clinical trials (White 947
et al., 2016). The mechanism of action of KAF156 is yet to be determined (Kuhen et 948
al., 2014). 949
950
Many compounds that have been identified in whole-cell high-throughput screens 951
affect parasite ion homeostasis (Rottmann et al., 2010, Jimenez-Diaz et al., 2014, 952
Lehane et al., 2014, Flannery et al., 2015). These compounds are discussed further in 953
Sections 1.6. 954
955