Product development (patient exploratory)

Discovered in 2011 by a partnership between Monash University, the University of Nebraska and the STPHI, OZ439 (Fig. 24) is a synthetic trioxolane that possesses the fast-acting curative and transmission-blocking ability, and is active against artemisinin-resistant parasites [106].

Fig. 24

Key biological and physical properties of OZ439. In vivo ED₉₀ kindly provided by S. Wittlin, personal communication, 2018.

The discovery of OZ439 (a.k.a. Artefenomel) stems from the lead optimization of the pre-existing trioxolane OZ277 (O1, Fig. 25, a.k.a. Arterolane), which was discovered in 2004 [107, 108]. As previous studies have shown that the adamantane-peroxide moiety is essential for anti-malarial activity, studies were focused on the eastern portion of the molecule [109, 110, 111]. Notably, analogues with variation in this area were found to have largely similar in vitro activities. In order to improve upon the original lead compound OZ277 (mean survival 11 days, 0 from 5 mice cured), replacement of the amide linker with a phenyl ether linker (O2) led to improved exposure and single-dose curative efficacy (mean survival > 30 days, 5 from 5 mice cured when applied in a single dose of 30 mg/kg 1 day after infection). An attempt to further improve the exposure saw the replacement of the alkylamine chain with a terminal piperazine unit (O3), however, this led to a significant reduction in curative efficacy (mean survival 17 days, 0 from 5 mice cured). Encouragingly, using morpholine (OZ439) resulted in a mean survival of > 30 days, with 5 from 5 mice cured [112]. While OZ439 has shown excellent curative efficacy, no clear correlations were found between in vitro and in vivo activity or physiochemical properties and exposure. As a result, this new lead compound possesses significantly lower solubility and slightly lower potency than OZ277.

Fig. 25

Key stages in the hit to lead pathway of OZ439. Initial replacement of the amide linker with a phenyl ether linker resulted in improved exposure while maintaining potency (O2). The exposure was further improved by changing the alkylamine chain to a piperazine ring (O3). Final replacement of the piperazine ring with a morpholine unit led to the optimized compound OZ439, which possessed better curative efficacy in vivo.

Unlike other peroxide-containing antimalarials (e.g. artemisinin), OZ439 was found to be curative at a single dose of 20 mg/kg in the P. berghei mouse model and possessed prophylactic activity superior to mefloquine when a 30 mg/kg dose was applied 24 h prior to malaria infection. OZ439 also showed full anti-malarial protection when administered 96 h before infection at a dose of 100 mg/kg. Only minor signs of toxicity were observed in a rat model when five consecutive doses of 300 mg/kg were administered with 3 days interval. Conversely, O2 resulted in the death of 9 from 12 animals when used at the same dose [112]. OZ439 has shown a significantly longer half-life in humans (about 46–62 h) when compared to the hit compound OZ277 (3h) [113, 114, 115].

Much like the current peroxide-containing antimalarials, the precise MoA for OZ439 has yet to be discovered but it is believed that oxidative stress plays a major role [116, 117]. First-in-human results for OZ439 were published in 2013 [118] and since then the molecule has progressed into Phase IIb clinical trials with planned completion in 2019 (NCT02497612) [119].

KAF156 (Fig. 26) was identified in 2008 by Novartis and The Scripps Research Institute and is part of the second-generation of imidazolopiperazine antimalarials that potentially possesses a novel MoA and performs as a rapid-acting drug [120].

Fig. 26

Key biological and physical properties of KAF156. Solubility and logD kindly provided by T. Diagana, personal communication, 2018.

A high-throughput screen of 1.7 million compounds in a P. falciparum proliferation assay led to the identification of hit compound K1 (Fig. 27). In order to protect the metabolically vulnerable positions in the lead compound and to optimize the potency, the benzodioxole and phenyl fragments were replaced with 4-fluorobenzene groups (K2). Installation of a dimethyl group in the piperazine ring led to the twofold more potent compound KAF156 [121, 122]. Notably, removal of the glycine residue from the piperazine ring was shown to increase the in vitro potency further (EC₅₀ = 5 nM), but this was accompanied by a lower parasitaemia reduction, indicating the importance of the amino acid side chain for in vivo efficacy [121].

Fig. 27

Key stages in the hit to lead pathway of KAF156. Potential metabolic stability issues were addressed through fluorine bioisosteres giving K2. Introduction of a dimethyl group on the piperazine ring resulted in increased potency and led to the optimized compound.

In the causal prophylactic rodent malaria model, a single oral dose of 10 mg/kg administered 2h before intravenous infection with P. berghei sporozoites were shown to be fully protective. KAF156 has also shown transmission-blocking ability in the P. berghei model.

The MoA for KAF156 remains unknown. Through the culturing of resistant strains, mutations have been identified in three genes: P. falciparum Cyclic Amine Resistance Locus (Pf CARL), UDP-galactose and Acetyl-CoA transporters [58, 123]. KAF156 is currently in Phase IIb clinical trials, administered in combination with Lumefantrine (NCT03167242).

KAE609/Cipargamin/NITD609 (Fig. 28) was discovered by a partnership between Novartis, the STPHI and the Wellcome Trust [124, 125].

Fig. 28

Key biological and physical properties of KAE609. *Significant inhibition at 50 and 500 nM doses.

A high-throughput P. falciparum proliferation assay identified a racemic spiroazepineindole compound (K′1, Fig. 29) which possessed moderate potency against K1 and NF54 strains, as well as potent in vivo efficacy. Confirmation of the hit result by resynthesis of K′1 resulted in the isolation of a ∼ 9:1 diastereomeric ratio favouring the (1R,3S) and (1S,3R) pair of enantiomers. Chiral separation of this major pair identified the (1R,3S) enantiomer K′2 as being > 250-fold more potent than the (1S,3R) enantiomer. Reduction of the ring size (K′3) facilitated a diastereoselective Pictet–Spengler reaction, which ultimately led to the production of highly potent spiroindolone KAE609.

Fig. 29

Key stages in the hit to lead pathway of KAE609. Resolution of the initial racemic hit (K′1) gave the significantly more potent stereoisomer (K′2). Reducing the ring size further increased the potency and final halogenation of the indole ring led to the optimized compound KAE609.

KAE609 is equipotent against drug-resistant strains and was found to be as effective as artesunate against P. falciparum and P. vivax isolates [126]. It shows a good safety profile, with low cytotoxicity, cardiotoxicity and mutagenic activity, and is able to clear parasitaemia rapidly in adults with uncomplicated P. falciparum or P. vivax malaria at a dose of 30 mg/day for 3 days. KAE609 displays low clearance from the body has a long half-life and excellent bioavailability.

KAE609 is one of the key novel compounds that has been found to act through inhibition of Pf ATP4 (vide infra). Along with a number of other structurally distinct compounds that are also thought to inhibit Pf ATP4 as their MoA, KAE609 has shown great promise as a new anti-malarial candidate and is currently in Phase IIb trials (NCT03334747).

DSM265 (Fig. 30) was discovered through a collaboration between the University of Texas Southwestern, the University of Washington and Monash University [127].

Fig. 30

Key biological and physical properties of DSM265.

Identified in a high-throughput screen of P. falciparum dihydroorotate dehydrogenase (Pf DHODH)-based enzyme activity, the selective inhibitor D′1 (Fig. 31) was found. The compound showed high potency but was rapidly metabolized and was inactive in vivo. Substitution of the naphthyl ring with a trifluoromethylphenyl group led to a metabolically stable analogue (D′2). Examination of the bound crystal structures of D′1 and D′2 with Pf DHODH identified key residue interactions which eventually led to the installation of a difluoroethyl group (D′3) which significantly improved the potency and solubility. Final modification of the trifluoromethyl group with a pentafluorosulfur group led to the optimized compound DSM265.

Fig. 31

Key stages in the hit to lead pathway of DSM265. Replacement of the second phenyl ring in the naphthyl group with a trifluoromethyl group improved solubility. Addition of a 1,1-difluoroethyl group significantly increased the potency and final replacement of the trifluoromethyl group in D′2 with a pentafluorosulfur moiety led to the optimized compound DSM265.

DSM265 has been shown to be a highly selective inhibitor of malarial DHODH and is potent against both blood and liver stages of P. falciparum and also drug-resistant parasite isolates. It has an excellent safety profile (provides therapeutic concentrations after 8 days of a single oral dose, tolerated in repeat-doses, non-mutagenic, inactive against human enzymes/receptors) and has a very low clearance rate and a long half-life in humans [128]. DSM265 has completed Phase IIa trials in Peru in patients with P. falciparum or P. vivax (NCT02123290) and also completed a controlled human malaria infection study in combination with OZ439 (NCT02389348).

Identified in 2012 by a team at the University of Cape Town, South Africa in the same campaign as UCT943, MMV048 (Fig. 32) is another compound in the 3,5-diaryl-2-aminopyridine class that possesses good prophylactic activity against Plasmodium cynomolgi in vivo and has the potential to act as a transmission-blocking drug [84].

Fig. 32

Key biological and physical properties of MMV048.

The initial hit (M″1, Fig. 33) showed potent in vitro activity against the NF54 (drug-sensitive) strain of P. falciparum (IC₅₀ = 49 nM) but suffered from poor solubility and high metabolic clearance. The metabolically labile 3-methoxy-4-hydroxyphenyl moiety was replaced by a methoxypyridyl ring giving the more stable and equipotent compound M″2. Further SAR studies revealed that replacement of the methoxy group by a trifluoromethyl group led to the improved potency and stability of MMV048.

Fig. 33

Key stages in the hit to lead pathway of MMV048. Initial replacement of the 3-methoxy-4-hydroxyphenyl moiety helped to improve in vivo stability and solubility. Further replacement of the methoxy group with a trifluoromethyl group improved potency and metabolic stability leading to the optimized compound.

MMV048 has shown 99.3% reduction in parasitaemia in the P. berghei mouse model at a single dose of 30 mg/kg with no signs of parasites after 30 days (ED₉₀ = 1.7 mg/kg). This highlights the potential of MMV048 to act as a single dose treatment.

The target of MMV048 is Pf PI4K, which was recently revealed as a new MoA for anti-malarial drugs (vide infra) [87]. MMV048 is currently in Phase IIa clinical trials in Ethiopia [58].